Figure 6 - uploaded by David Fletcher Smith
Content may be subject to copyright.
Scanning electron micrographs of Hessian fly larvae on wheat plants. A, One-day-old avirulent biotype L larva oriented perpendicular to leaf sheath veins of H9 -Iris wheat (resistant plant). B, One-day-old virulent biotype L larva oriented parallel to leaf sheath veins of Newton wheat (susceptible plant nearly isogenic to H9 -Iris). C, Three-day-old avirulent biotype L larva showing a lack of growth and development on the leaf sheath of a H9 -Iris wheat plant. D, Three-day- old virulent biotype L larva showing increased size on the leaf sheath of a Newton wheat plant. E, Frontal view of writhing 1-d-old avirulent biotype L larva oriented perpendicular to leaf sheath veins. F, Lateral view of the anterior end of a 3-d-old virulent biotype L larva showing mouth parts firmly attached to the plant tissue. Note the parallel orientation of the virulent larvae in the epidermal groove of the leaf sheath of the susceptible plants and the apparent disorientation of the avirulent larvae on the resistant plants.
Source publication
We previously cloned and characterized a novel jacalin-like lectin gene from wheat (Triticum aestivum) plants that responds to infestation by Hessian fly (Mayetiola destructor) larvae, a major dipteran pest of this crop. The infested resistant plants accumulated higher levels of Hfr-1 (for Hessian fly-responsive gene 1) transcripts compared with un...
Contexts in source publication
Context 1
... in extracts from larvae that had fed on plants were not of larval origin. In addition, data- base searches never identified insect genes or proteins with similarity to the HFR1 sequence. Thus, bands detected in samples from virulent and avirulent larvae corresponded to ingested wheat HFR1 and its degradation products. To determine whether HFR1 had detrimental effects on insect growth and development, we carried out an insect feeding bioassay using the recombinant His 6 - HFR1 protein. Since Hessian fly larvae are obligate parasites of the plant and cannot be cultured in vitro, we used another dipteran insect, Drosophila melanogaster (fruit fly), in a diet incorporation bioassay. Compari- son of larval length at 5 d after egg hatch showed no significant differences between the larvae fed the control diet and the lowest concentration of 0.3 m g of His 6 -HFR1 per gram of diet ( P 5 0.09), but did show a significant inverse correlation ( P , 0.0001) with the concentration of His 6 -HFR1 in all other treatments (Fig. 5A). The developmental time and mortality of D. melanogaster larvae increased with higher concentrations of His -HFR1 in the diet. At the lowest concentrations of His 6 -HFR1 (0.3 and 1.5 m g g ), the times to pupation and to eclosion as adults were not significantly different ( P . 0.1) from those of the controls (Fig. 5B). Interme- diate concentrations of His 6 -HFR1 (3, 6, and 9 m g g 2 1 ) significantly lengthened the time to pupation ( P , 0.001) and increased mortality (Fig. 5B; Table I). The highest concentrations (12 and 15 m g g 2 1 ) led to death prior to pupation (Fig. 5B; Table I). Adult flies emerged approximately 4.3 d after pupation on the control diet as well as on the diet containing 0.3, 1.5, and 3 m g of His 6 -HFR1 per gram of diet. However, His 6 -HFR1 concentrations of 6 and 9 m g g 2 1 resulted in arrested development in the puparium and inability to eclose as adults. At the highest concentrations tested (12 and 15 m g g 2 1 ), on days 2 through 11, we observed larvae crawling up the walls of their tubes, presumably to avoid contact with the diet or to search for an alterna- tive food source. These larvae remained in the first instar, did not pupate, and appeared to die slowly of starvation. In contrast, larvae remained in the diet until just prior to pupation when fed lower concentrations of His 6 -HFR1 or the control diet (data not shown). Table I shows the proportion of D. melanogaster larvae that survived to pupation and to adult eclosion when feeding on progressively higher concentrations of His 6 - HFR1. The percentage of larvae that reached pupation and eclosed as adults differed significantly ( P , 0.01) from the control at a concentration of 3.0 m g of His 6 - HFR1 per gram of diet. The concentration at which 50% of the larvae died of His 6 -HFR1 for the larval stage was calculated to be 6.55 6 0.15 m g g 2 1 diet. We compared the behaviors of developing biotype L Hessian fly larvae during the period when the defense response was activated and wheat HFR1 lectin was being produced in resistant but not in susceptible plants. The observed behaviors are defined in detail in ‘‘Materials and Methods,’’ whereas the frequency and duration of these behaviors, on which statistical analyses are based, are summarized in Table II. During the first 6 to 12 h after egg hatch, both virulent and avirulent larvae crawled to the base of the wheat seedling among the leaf sheaths and exhibited identical searching behavior and body contractions (Supplemental Video S1); no significant differences were detected in the frequency of either the searching events or body contraction patterns ( P $ 0.05). Neither avirulent nor virulent larvae appeared to be oriented with respect to veins of the leaf sheath through 12 h after egg hatch. During this period, defense mechanisms in resistant plants, includ- ing the Hfr-1 gene, are just being activated by the presence of avirulent larvae and low levels of Hfr-1 mRNA are detectable (Subramanyam et al., 2006). One day after egg hatch (24-h time point), when HFR1 protein is clearly detectable in resistant plants (Fig. 3), the avirulent and virulent larvae did not differ in size or shape (Fig. 6, A and B) but behavioral differences were evident. During this time, the virulent larvae showed signs of settling (Table II), indicated by fewer searching events per minute compared with the avirulent larvae ( P , 0.00001) and the initiation of gut contractions ( P , 0.00001; Supplemental Video S2; Table II). The virulent larvae aligned themselves in rows within the grooves corresponding to veins of the leaf sheath (Fig. 6B). This sessile behavior suggested that the larvae had begun to feed. In contrast, the avirulent larvae by 24 h after hatch exhibited more searching events per minute than the virulent larvae ( P , 0.00001) and produced body contractions associated with locomotion. These avirulent larvae were often positioned perpendicular to the leaf veins (Fig. 6, A and E). By 24 h after egg hatch, a time when HFR1 protein was detectable and defenses were fully established in resistant plants, the avirulent larvae displayed writhing and head-rearing behaviors that were never exhibited by the virulent larvae during any developmental stage. Larvae that were rearing their heads did not display the searching behavior. This writhing (Fig. 6E; Supplemental Video S3) and head rearing (Supplemental Fig. S3) lasted until 72 and 96 h after hatch, respectively, after which these behaviors ceased. The avirulent larvae continued to appear disoriented with respect to their positions relative to leaf veins at 3 d after hatch (Fig. 6C). All virulent larvae were sessile, with heads attached to feeding sites from 48 h after egg hatch until pupation (Fig. 6, D and F), whereas the avirulent larvae became sessile only after 96 h. At no time did the avirulent larvae exhibit visible gut contractions indicative of food ingestion. By 96 h after hatch, the virulent larvae had successfully established feeding sites and fed voraciously (Supplemental Video S4). With careful observation near the end of this video, one sees evidence of saliva being expelled onto the leaf surface and then ingested. By 192 h after hatch, the avirulent larvae appeared flattened and desiccated, which suggested that they were dead (Supplemental Fig. S4), whereas the virulent larvae, now in their second instar, had greatly increased in size. During the first 24 h after egg hatch, first-instar virulent and avirulent Hessian fly larvae did not differ in length ( P 5 0.37; Fig. 7). However, the first-instar virulent larvae showed a significant increase in length compared with the avirulent larvae ( P 5 0.003) by 48 h after egg hatch and 72 h (Fig. 6, C and D). By 120 h after hatch, the virulent larvae had more than doubled in length compared with the avirulent larvae ( P , 0.00001). The virulent larvae were in the second-instar stage by 192 h after hatch and had grown to over 2 mm in length. In contrast, at 192 h after hatch, the avirulent larvae were dead. They had not developed past the first-instar stage and had not increased in length ( P 5 0.58, 0.41 6 0.02 mm) since hatching from the egg. Developmental outcomes diverge quickly in compatible and incompatible interactions for both the wheat plants and the Hessian fly larvae. Initially, virulent and avirulent larvae exhibit identical behaviors during the 12 h following egg hatch (Table II) as they migrate to the base of the plant searching for a permanent feeding site. During the migration, larvae exhibit searching behavior (Table II), puncture leaf sheath epidermal cells with their minute mandibles (0.5 m m long 3 0.1 m m diameter; Harris et al., 2006), and apply salivary secretions containing elicitors of plant responses. However, the plant responses diverge within about 6 h, with an induction of the first stages of susceptibility. During day 1 of compatible interactions, cell walls at the attack site of susceptible plants begin to thin and develop 4- to 10- m m ruptures (Harris et al., 2006) through which cellular contents flow. Virulent larvae rapidly became sessile and fed on the contents of these lysing cells (Table II; Harris et al., 2006) days before the nutritive tissue that will sustain larval development becomes well established. Two days after egg hatch, virulent larvae were noticeably larger than avirulent larvae (Figs. 6 and 7). Over the next few days, tran- script levels from wheat genes involved in sugar trans- port (Liu et al., 2007; C.E. Williams, S. Subramanyam, K.D. Saltzmann, J.A. Nemacheck, and C. Zheng, unpublished data), amino acid synthesis (K.D. Saltzmann and C.E. Williams, unpublished data), HFR2 (a pore- forming protein; Puthoff et al., 2005), and genes involved in polyamine biosynthesis (S. Subramanyam and C.E. Williams, unpublished data) increased in susceptible plants. These changes in wheat mRNA levels occur at the same time that physical changes in leaf sheath epidermal cells become evident in susceptible plants, resulting in the formation of nutritive tissue (Harris et al., 2006). This tissue provides the virulent larvae with a diet rich in soluble proteins and sugars that sustain their growth and development until pupation. In contrast to the rapid initiation of feeding by the virulent larvae, the avirulent larvae exhibited prolonged activity (Table II), as a gene-for-gene recognition event triggers increased mRNA levels of defense genes (Sardesai et al., 2005b) along with genes encoding lectins, such as Hfr-1 (Subramanyam et al., 2006) and Hfr-3 (Giovanini et al., 2007). The specific elicita- tion of these wheat lectins, which previously have not been linked to defense, suggests the targeted involve- ment of this class of proteins during resistance to Hessian fly. In response to these changes in the resistant wheat plant, avirulent larvae exhibit a number of stress responses. Avirulent larvae have a nearly 8-fold ...
Context 2
... mass of 37.5 kD, corresponding to the size of the intact HFR1 protein. The smaller, more abundant bands appeared to be proteolytic products of HFR1 (Fig. 4). Only two very faint low molecular mass bands were seen in lanes containing extracts from avirulent larvae (Fig. 4). The high levels of HFR1 in resistant plants (Fig. 3) and the corresponding absence of intact HFR1 protein in the avirulent larvae that reside on but do not grow on these plants suggested that the avirulent larvae feed minimally. The anti-HFR1 antibodies did not detect HFR1, its degradation products, or any other bands on blots containing extract from neonate larvae (data not shown) that were allowed to emerge in water and had never fed on wheat plants. This result indicated that the bands detected in extracts from larvae that had fed on plants were not of larval origin. In addition, data- base searches never identified insect genes or proteins with similarity to the HFR1 sequence. Thus, bands detected in samples from virulent and avirulent larvae corresponded to ingested wheat HFR1 and its degradation products. To determine whether HFR1 had detrimental effects on insect growth and development, we carried out an insect feeding bioassay using the recombinant His 6 - HFR1 protein. Since Hessian fly larvae are obligate parasites of the plant and cannot be cultured in vitro, we used another dipteran insect, Drosophila melanogaster (fruit fly), in a diet incorporation bioassay. Compari- son of larval length at 5 d after egg hatch showed no significant differences between the larvae fed the control diet and the lowest concentration of 0.3 m g of His 6 -HFR1 per gram of diet ( P 5 0.09), but did show a significant inverse correlation ( P , 0.0001) with the concentration of His 6 -HFR1 in all other treatments (Fig. 5A). The developmental time and mortality of D. melanogaster larvae increased with higher concentrations of His -HFR1 in the diet. At the lowest concentrations of His 6 -HFR1 (0.3 and 1.5 m g g ), the times to pupation and to eclosion as adults were not significantly different ( P . 0.1) from those of the controls (Fig. 5B). Interme- diate concentrations of His 6 -HFR1 (3, 6, and 9 m g g 2 1 ) significantly lengthened the time to pupation ( P , 0.001) and increased mortality (Fig. 5B; Table I). The highest concentrations (12 and 15 m g g 2 1 ) led to death prior to pupation (Fig. 5B; Table I). Adult flies emerged approximately 4.3 d after pupation on the control diet as well as on the diet containing 0.3, 1.5, and 3 m g of His 6 -HFR1 per gram of diet. However, His 6 -HFR1 concentrations of 6 and 9 m g g 2 1 resulted in arrested development in the puparium and inability to eclose as adults. At the highest concentrations tested (12 and 15 m g g 2 1 ), on days 2 through 11, we observed larvae crawling up the walls of their tubes, presumably to avoid contact with the diet or to search for an alterna- tive food source. These larvae remained in the first instar, did not pupate, and appeared to die slowly of starvation. In contrast, larvae remained in the diet until just prior to pupation when fed lower concentrations of His 6 -HFR1 or the control diet (data not shown). Table I shows the proportion of D. melanogaster larvae that survived to pupation and to adult eclosion when feeding on progressively higher concentrations of His 6 - HFR1. The percentage of larvae that reached pupation and eclosed as adults differed significantly ( P , 0.01) from the control at a concentration of 3.0 m g of His 6 - HFR1 per gram of diet. The concentration at which 50% of the larvae died of His 6 -HFR1 for the larval stage was calculated to be 6.55 6 0.15 m g g 2 1 diet. We compared the behaviors of developing biotype L Hessian fly larvae during the period when the defense response was activated and wheat HFR1 lectin was being produced in resistant but not in susceptible plants. The observed behaviors are defined in detail in ‘‘Materials and Methods,’’ whereas the frequency and duration of these behaviors, on which statistical analyses are based, are summarized in Table II. During the first 6 to 12 h after egg hatch, both virulent and avirulent larvae crawled to the base of the wheat seedling among the leaf sheaths and exhibited identical searching behavior and body contractions (Supplemental Video S1); no significant differences were detected in the frequency of either the searching events or body contraction patterns ( P $ 0.05). Neither avirulent nor virulent larvae appeared to be oriented with respect to veins of the leaf sheath through 12 h after egg hatch. During this period, defense mechanisms in resistant plants, includ- ing the Hfr-1 gene, are just being activated by the presence of avirulent larvae and low levels of Hfr-1 mRNA are detectable (Subramanyam et al., 2006). One day after egg hatch (24-h time point), when HFR1 protein is clearly detectable in resistant plants (Fig. 3), the avirulent and virulent larvae did not differ in size or shape (Fig. 6, A and B) but behavioral differences were evident. During this time, the virulent larvae showed signs of settling (Table II), indicated by fewer searching events per minute compared with the avirulent larvae ( P , 0.00001) and the initiation of gut contractions ( P , 0.00001; Supplemental Video S2; Table II). The virulent larvae aligned themselves in rows within the grooves corresponding to veins of the leaf sheath (Fig. 6B). This sessile behavior suggested that the larvae had begun to feed. In contrast, the avirulent larvae by 24 h after hatch exhibited more searching events per minute than the virulent larvae ( P , 0.00001) and produced body contractions associated with locomotion. These avirulent larvae were often positioned perpendicular to the leaf veins (Fig. 6, A and E). By 24 h after egg hatch, a time when HFR1 protein was detectable and defenses were fully established in resistant plants, the avirulent larvae displayed writhing and head-rearing behaviors that were never exhibited by the virulent larvae during any developmental stage. Larvae that were rearing their heads did not display the searching behavior. This writhing (Fig. 6E; Supplemental Video S3) and head rearing (Supplemental Fig. S3) lasted until 72 and 96 h after hatch, respectively, after which these behaviors ceased. The avirulent larvae continued to appear disoriented with respect to their positions relative to leaf veins at 3 d after hatch (Fig. 6C). All virulent larvae were sessile, with heads attached to feeding sites from 48 h after egg hatch until pupation (Fig. 6, D and F), whereas the avirulent larvae became sessile only after 96 h. At no time did the avirulent larvae exhibit visible gut contractions indicative of food ingestion. By 96 h after hatch, the virulent larvae had successfully established feeding sites and fed voraciously (Supplemental Video S4). With careful observation near the end of this video, one sees evidence of saliva being expelled onto the leaf surface and then ingested. By 192 h after hatch, the avirulent larvae appeared flattened and desiccated, which suggested that they were dead (Supplemental Fig. S4), whereas the virulent larvae, now in their second instar, had greatly increased in size. During the first 24 h after egg hatch, first-instar virulent and avirulent Hessian fly larvae did not differ in length ( P 5 0.37; Fig. 7). However, the first-instar virulent larvae showed a significant increase in length compared with the avirulent larvae ( P 5 0.003) by 48 h after egg hatch and 72 h (Fig. 6, C and D). By 120 h after hatch, the virulent larvae had more than doubled in length compared with the avirulent larvae ( P , 0.00001). The virulent larvae were in the second-instar stage by 192 h after hatch and had grown to over 2 mm in length. In contrast, at 192 h after hatch, the avirulent larvae were dead. They had not developed past the first-instar stage and had not increased in length ( P 5 0.58, 0.41 6 0.02 mm) since hatching from the egg. Developmental outcomes diverge quickly in compatible and incompatible interactions for both the wheat plants and the Hessian fly larvae. Initially, virulent and avirulent larvae exhibit identical behaviors during the 12 h following egg hatch (Table II) as they migrate to the base of the plant searching for a permanent feeding site. During the migration, larvae exhibit searching behavior (Table II), puncture leaf sheath epidermal cells with their minute mandibles (0.5 m m long 3 0.1 m m diameter; Harris et al., 2006), and apply salivary secretions containing elicitors of plant responses. However, the plant responses diverge within about 6 h, with an induction of the first stages of susceptibility. During day 1 of compatible interactions, cell walls at the attack site of susceptible plants begin to thin and develop 4- to 10- m m ruptures (Harris et al., 2006) through which cellular contents flow. Virulent larvae rapidly became sessile and fed on the contents of these lysing cells (Table II; Harris et al., 2006) days before the nutritive tissue that will sustain larval development becomes well established. Two days after egg hatch, virulent larvae were noticeably larger than avirulent larvae (Figs. 6 and 7). Over the next few days, tran- script levels from wheat genes involved in sugar trans- port (Liu et al., 2007; C.E. Williams, S. Subramanyam, K.D. Saltzmann, J.A. Nemacheck, and C. Zheng, unpublished data), amino acid synthesis (K.D. Saltzmann and C.E. Williams, unpublished data), HFR2 (a pore- forming protein; Puthoff et al., 2005), and genes involved in polyamine biosynthesis (S. Subramanyam and C.E. Williams, unpublished data) increased in susceptible plants. These changes in wheat mRNA levels occur at the same time that physical changes in leaf sheath epidermal cells become evident in susceptible plants, resulting in the formation of nutritive tissue (Harris et al., 2006). This tissue provides the virulent larvae with a diet rich in soluble proteins ...
Context 3
... fed the control diet and the lowest concentration of 0.3 m g of His 6 -HFR1 per gram of diet ( P 5 0.09), but did show a significant inverse correlation ( P , 0.0001) with the concentration of His 6 -HFR1 in all other treatments (Fig. 5A). The developmental time and mortality of D. melanogaster larvae increased with higher concentrations of His -HFR1 in the diet. At the lowest concentrations of His 6 -HFR1 (0.3 and 1.5 m g g ), the times to pupation and to eclosion as adults were not significantly different ( P . 0.1) from those of the controls (Fig. 5B). Interme- diate concentrations of His 6 -HFR1 (3, 6, and 9 m g g 2 1 ) significantly lengthened the time to pupation ( P , 0.001) and increased mortality (Fig. 5B; Table I). The highest concentrations (12 and 15 m g g 2 1 ) led to death prior to pupation (Fig. 5B; Table I). Adult flies emerged approximately 4.3 d after pupation on the control diet as well as on the diet containing 0.3, 1.5, and 3 m g of His 6 -HFR1 per gram of diet. However, His 6 -HFR1 concentrations of 6 and 9 m g g 2 1 resulted in arrested development in the puparium and inability to eclose as adults. At the highest concentrations tested (12 and 15 m g g 2 1 ), on days 2 through 11, we observed larvae crawling up the walls of their tubes, presumably to avoid contact with the diet or to search for an alterna- tive food source. These larvae remained in the first instar, did not pupate, and appeared to die slowly of starvation. In contrast, larvae remained in the diet until just prior to pupation when fed lower concentrations of His 6 -HFR1 or the control diet (data not shown). Table I shows the proportion of D. melanogaster larvae that survived to pupation and to adult eclosion when feeding on progressively higher concentrations of His 6 - HFR1. The percentage of larvae that reached pupation and eclosed as adults differed significantly ( P , 0.01) from the control at a concentration of 3.0 m g of His 6 - HFR1 per gram of diet. The concentration at which 50% of the larvae died of His 6 -HFR1 for the larval stage was calculated to be 6.55 6 0.15 m g g 2 1 diet. We compared the behaviors of developing biotype L Hessian fly larvae during the period when the defense response was activated and wheat HFR1 lectin was being produced in resistant but not in susceptible plants. The observed behaviors are defined in detail in ‘‘Materials and Methods,’’ whereas the frequency and duration of these behaviors, on which statistical analyses are based, are summarized in Table II. During the first 6 to 12 h after egg hatch, both virulent and avirulent larvae crawled to the base of the wheat seedling among the leaf sheaths and exhibited identical searching behavior and body contractions (Supplemental Video S1); no significant differences were detected in the frequency of either the searching events or body contraction patterns ( P $ 0.05). Neither avirulent nor virulent larvae appeared to be oriented with respect to veins of the leaf sheath through 12 h after egg hatch. During this period, defense mechanisms in resistant plants, includ- ing the Hfr-1 gene, are just being activated by the presence of avirulent larvae and low levels of Hfr-1 mRNA are detectable (Subramanyam et al., 2006). One day after egg hatch (24-h time point), when HFR1 protein is clearly detectable in resistant plants (Fig. 3), the avirulent and virulent larvae did not differ in size or shape (Fig. 6, A and B) but behavioral differences were evident. During this time, the virulent larvae showed signs of settling (Table II), indicated by fewer searching events per minute compared with the avirulent larvae ( P , 0.00001) and the initiation of gut contractions ( P , 0.00001; Supplemental Video S2; Table II). The virulent larvae aligned themselves in rows within the grooves corresponding to veins of the leaf sheath (Fig. 6B). This sessile behavior suggested that the larvae had begun to feed. In contrast, the avirulent larvae by 24 h after hatch exhibited more searching events per minute than the virulent larvae ( P , 0.00001) and produced body contractions associated with locomotion. These avirulent larvae were often positioned perpendicular to the leaf veins (Fig. 6, A and E). By 24 h after egg hatch, a time when HFR1 protein was detectable and defenses were fully established in resistant plants, the avirulent larvae displayed writhing and head-rearing behaviors that were never exhibited by the virulent larvae during any developmental stage. Larvae that were rearing their heads did not display the searching behavior. This writhing (Fig. 6E; Supplemental Video S3) and head rearing (Supplemental Fig. S3) lasted until 72 and 96 h after hatch, respectively, after which these behaviors ceased. The avirulent larvae continued to appear disoriented with respect to their positions relative to leaf veins at 3 d after hatch (Fig. 6C). All virulent larvae were sessile, with heads attached to feeding sites from 48 h after egg hatch until pupation (Fig. 6, D and F), whereas the avirulent larvae became sessile only after 96 h. At no time did the avirulent larvae exhibit visible gut contractions indicative of food ingestion. By 96 h after hatch, the virulent larvae had successfully established feeding sites and fed voraciously (Supplemental Video S4). With careful observation near the end of this video, one sees evidence of saliva being expelled onto the leaf surface and then ingested. By 192 h after hatch, the avirulent larvae appeared flattened and desiccated, which suggested that they were dead (Supplemental Fig. S4), whereas the virulent larvae, now in their second instar, had greatly increased in size. During the first 24 h after egg hatch, first-instar virulent and avirulent Hessian fly larvae did not differ in length ( P 5 0.37; Fig. 7). However, the first-instar virulent larvae showed a significant increase in length compared with the avirulent larvae ( P 5 0.003) by 48 h after egg hatch and 72 h (Fig. 6, C and D). By 120 h after hatch, the virulent larvae had more than doubled in length compared with the avirulent larvae ( P , 0.00001). The virulent larvae were in the second-instar stage by 192 h after hatch and had grown to over 2 mm in length. In contrast, at 192 h after hatch, the avirulent larvae were dead. They had not developed past the first-instar stage and had not increased in length ( P 5 0.58, 0.41 6 0.02 mm) since hatching from the egg. Developmental outcomes diverge quickly in compatible and incompatible interactions for both the wheat plants and the Hessian fly larvae. Initially, virulent and avirulent larvae exhibit identical behaviors during the 12 h following egg hatch (Table II) as they migrate to the base of the plant searching for a permanent feeding site. During the migration, larvae exhibit searching behavior (Table II), puncture leaf sheath epidermal cells with their minute mandibles (0.5 m m long 3 0.1 m m diameter; Harris et al., 2006), and apply salivary secretions containing elicitors of plant responses. However, the plant responses diverge within about 6 h, with an induction of the first stages of susceptibility. During day 1 of compatible interactions, cell walls at the attack site of susceptible plants begin to thin and develop 4- to 10- m m ruptures (Harris et al., 2006) through which cellular contents flow. Virulent larvae rapidly became sessile and fed on the contents of these lysing cells (Table II; Harris et al., 2006) days before the nutritive tissue that will sustain larval development becomes well established. Two days after egg hatch, virulent larvae were noticeably larger than avirulent larvae (Figs. 6 and 7). Over the next few days, tran- script levels from wheat genes involved in sugar trans- port (Liu et al., 2007; C.E. Williams, S. Subramanyam, K.D. Saltzmann, J.A. Nemacheck, and C. Zheng, unpublished data), amino acid synthesis (K.D. Saltzmann and C.E. Williams, unpublished data), HFR2 (a pore- forming protein; Puthoff et al., 2005), and genes involved in polyamine biosynthesis (S. Subramanyam and C.E. Williams, unpublished data) increased in susceptible plants. These changes in wheat mRNA levels occur at the same time that physical changes in leaf sheath epidermal cells become evident in susceptible plants, resulting in the formation of nutritive tissue (Harris et al., 2006). This tissue provides the virulent larvae with a diet rich in soluble proteins and sugars that sustain their growth and development until pupation. In contrast to the rapid initiation of feeding by the virulent larvae, the avirulent larvae exhibited prolonged activity (Table II), as a gene-for-gene recognition event triggers increased mRNA levels of defense genes (Sardesai et al., 2005b) along with genes encoding lectins, such as Hfr-1 (Subramanyam et al., 2006) and Hfr-3 (Giovanini et al., 2007). The specific elicita- tion of these wheat lectins, which previously have not been linked to defense, suggests the targeted involve- ment of this class of proteins during resistance to Hessian fly. In response to these changes in the resistant wheat plant, avirulent larvae exhibit a number of stress responses. Avirulent larvae have a nearly 8-fold increase in levels of mRNA encoding d -glutathione S -transferase, which persists for several days and is presumed to function in an attempt to detoxify wheat allelochemicals (Mittapalli et al., 2007b). In addition, avirulent larvae experience oxidative stress, manifested as elevated levels of superoxide dismutase mRNA (Giovanini et al., 2006) along with phospholipid glutathione peroxidase mRNA (Mittapalli et al., 2007a). By 24 h after egg hatch, avirulent larvae exhibited two unusual behaviors that were never seen in virulent larvae: writhing (Supplemental Video S3) and head rearing (Table II; Supplemental Fig. S3). These behaviors may be another response to stresses from plant chemicals as well as to the stress of starvation. Avirulent larvae never settled or exhibited sustained ...
Context 4
... His 6 -HFR1 per gram of diet. However, His 6 -HFR1 concentrations of 6 and 9 m g g 2 1 resulted in arrested development in the puparium and inability to eclose as adults. At the highest concentrations tested (12 and 15 m g g 2 1 ), on days 2 through 11, we observed larvae crawling up the walls of their tubes, presumably to avoid contact with the diet or to search for an alterna- tive food source. These larvae remained in the first instar, did not pupate, and appeared to die slowly of starvation. In contrast, larvae remained in the diet until just prior to pupation when fed lower concentrations of His 6 -HFR1 or the control diet (data not shown). Table I shows the proportion of D. melanogaster larvae that survived to pupation and to adult eclosion when feeding on progressively higher concentrations of His 6 - HFR1. The percentage of larvae that reached pupation and eclosed as adults differed significantly ( P , 0.01) from the control at a concentration of 3.0 m g of His 6 - HFR1 per gram of diet. The concentration at which 50% of the larvae died of His 6 -HFR1 for the larval stage was calculated to be 6.55 6 0.15 m g g 2 1 diet. We compared the behaviors of developing biotype L Hessian fly larvae during the period when the defense response was activated and wheat HFR1 lectin was being produced in resistant but not in susceptible plants. The observed behaviors are defined in detail in ‘‘Materials and Methods,’’ whereas the frequency and duration of these behaviors, on which statistical analyses are based, are summarized in Table II. During the first 6 to 12 h after egg hatch, both virulent and avirulent larvae crawled to the base of the wheat seedling among the leaf sheaths and exhibited identical searching behavior and body contractions (Supplemental Video S1); no significant differences were detected in the frequency of either the searching events or body contraction patterns ( P $ 0.05). Neither avirulent nor virulent larvae appeared to be oriented with respect to veins of the leaf sheath through 12 h after egg hatch. During this period, defense mechanisms in resistant plants, includ- ing the Hfr-1 gene, are just being activated by the presence of avirulent larvae and low levels of Hfr-1 mRNA are detectable (Subramanyam et al., 2006). One day after egg hatch (24-h time point), when HFR1 protein is clearly detectable in resistant plants (Fig. 3), the avirulent and virulent larvae did not differ in size or shape (Fig. 6, A and B) but behavioral differences were evident. During this time, the virulent larvae showed signs of settling (Table II), indicated by fewer searching events per minute compared with the avirulent larvae ( P , 0.00001) and the initiation of gut contractions ( P , 0.00001; Supplemental Video S2; Table II). The virulent larvae aligned themselves in rows within the grooves corresponding to veins of the leaf sheath (Fig. 6B). This sessile behavior suggested that the larvae had begun to feed. In contrast, the avirulent larvae by 24 h after hatch exhibited more searching events per minute than the virulent larvae ( P , 0.00001) and produced body contractions associated with locomotion. These avirulent larvae were often positioned perpendicular to the leaf veins (Fig. 6, A and E). By 24 h after egg hatch, a time when HFR1 protein was detectable and defenses were fully established in resistant plants, the avirulent larvae displayed writhing and head-rearing behaviors that were never exhibited by the virulent larvae during any developmental stage. Larvae that were rearing their heads did not display the searching behavior. This writhing (Fig. 6E; Supplemental Video S3) and head rearing (Supplemental Fig. S3) lasted until 72 and 96 h after hatch, respectively, after which these behaviors ceased. The avirulent larvae continued to appear disoriented with respect to their positions relative to leaf veins at 3 d after hatch (Fig. 6C). All virulent larvae were sessile, with heads attached to feeding sites from 48 h after egg hatch until pupation (Fig. 6, D and F), whereas the avirulent larvae became sessile only after 96 h. At no time did the avirulent larvae exhibit visible gut contractions indicative of food ingestion. By 96 h after hatch, the virulent larvae had successfully established feeding sites and fed voraciously (Supplemental Video S4). With careful observation near the end of this video, one sees evidence of saliva being expelled onto the leaf surface and then ingested. By 192 h after hatch, the avirulent larvae appeared flattened and desiccated, which suggested that they were dead (Supplemental Fig. S4), whereas the virulent larvae, now in their second instar, had greatly increased in size. During the first 24 h after egg hatch, first-instar virulent and avirulent Hessian fly larvae did not differ in length ( P 5 0.37; Fig. 7). However, the first-instar virulent larvae showed a significant increase in length compared with the avirulent larvae ( P 5 0.003) by 48 h after egg hatch and 72 h (Fig. 6, C and D). By 120 h after hatch, the virulent larvae had more than doubled in length compared with the avirulent larvae ( P , 0.00001). The virulent larvae were in the second-instar stage by 192 h after hatch and had grown to over 2 mm in length. In contrast, at 192 h after hatch, the avirulent larvae were dead. They had not developed past the first-instar stage and had not increased in length ( P 5 0.58, 0.41 6 0.02 mm) since hatching from the egg. Developmental outcomes diverge quickly in compatible and incompatible interactions for both the wheat plants and the Hessian fly larvae. Initially, virulent and avirulent larvae exhibit identical behaviors during the 12 h following egg hatch (Table II) as they migrate to the base of the plant searching for a permanent feeding site. During the migration, larvae exhibit searching behavior (Table II), puncture leaf sheath epidermal cells with their minute mandibles (0.5 m m long 3 0.1 m m diameter; Harris et al., 2006), and apply salivary secretions containing elicitors of plant responses. However, the plant responses diverge within about 6 h, with an induction of the first stages of susceptibility. During day 1 of compatible interactions, cell walls at the attack site of susceptible plants begin to thin and develop 4- to 10- m m ruptures (Harris et al., 2006) through which cellular contents flow. Virulent larvae rapidly became sessile and fed on the contents of these lysing cells (Table II; Harris et al., 2006) days before the nutritive tissue that will sustain larval development becomes well established. Two days after egg hatch, virulent larvae were noticeably larger than avirulent larvae (Figs. 6 and 7). Over the next few days, tran- script levels from wheat genes involved in sugar trans- port (Liu et al., 2007; C.E. Williams, S. Subramanyam, K.D. Saltzmann, J.A. Nemacheck, and C. Zheng, unpublished data), amino acid synthesis (K.D. Saltzmann and C.E. Williams, unpublished data), HFR2 (a pore- forming protein; Puthoff et al., 2005), and genes involved in polyamine biosynthesis (S. Subramanyam and C.E. Williams, unpublished data) increased in susceptible plants. These changes in wheat mRNA levels occur at the same time that physical changes in leaf sheath epidermal cells become evident in susceptible plants, resulting in the formation of nutritive tissue (Harris et al., 2006). This tissue provides the virulent larvae with a diet rich in soluble proteins and sugars that sustain their growth and development until pupation. In contrast to the rapid initiation of feeding by the virulent larvae, the avirulent larvae exhibited prolonged activity (Table II), as a gene-for-gene recognition event triggers increased mRNA levels of defense genes (Sardesai et al., 2005b) along with genes encoding lectins, such as Hfr-1 (Subramanyam et al., 2006) and Hfr-3 (Giovanini et al., 2007). The specific elicita- tion of these wheat lectins, which previously have not been linked to defense, suggests the targeted involve- ment of this class of proteins during resistance to Hessian fly. In response to these changes in the resistant wheat plant, avirulent larvae exhibit a number of stress responses. Avirulent larvae have a nearly 8-fold increase in levels of mRNA encoding d -glutathione S -transferase, which persists for several days and is presumed to function in an attempt to detoxify wheat allelochemicals (Mittapalli et al., 2007b). In addition, avirulent larvae experience oxidative stress, manifested as elevated levels of superoxide dismutase mRNA (Giovanini et al., 2006) along with phospholipid glutathione peroxidase mRNA (Mittapalli et al., 2007a). By 24 h after egg hatch, avirulent larvae exhibited two unusual behaviors that were never seen in virulent larvae: writhing (Supplemental Video S3) and head rearing (Table II; Supplemental Fig. S3). These behaviors may be another response to stresses from plant chemicals as well as to the stress of starvation. Avirulent larvae never settled or exhibited sustained feeding and did not increase in size after hatching from the egg. They appeared to die slowly of starvation (death occurs at approximately day 5; Gallun, 1977; Supplemental Fig. S4) rather than being killed quickly by the consumption of highly toxic plant substances. Although the avirulent larvae may be able to feed on individual cells that they puncture with their mandibles, plant defenses inhibit them from ingesting life-sustaining quantities of nutrients. Experiments examining the feeding habits of Hessian fly larvae on 32 P-labeled resistant and susceptible wheat seedlings showed that virulent Hessian fly larvae had ingested six times more radiation than avirulent larvae at 2 d after hatch (Gallun and Langston, 1963). Avirulent larvae stopped uptake of 32 P by 3 d, whereas virulent larvae continued uptake for 14 d. Feeding or lack of feeding can result in physical changes within the insect midgut. The peritrophic matrix of the lepidopteran midgut is synthesized ...
Citations
... Several DIR proteins have already been structurally studied in several species, with the majority of them are linked to the fighting against diseases, insect infestation, abiotic stress resilience, or indeed combinatorial defense mechanisms. A DIR1 gene in Pisum sativum (Hadwiger et al., 1992), 6 Dirrigent genes in Pinus tree (Ralph et al. 2006), 4 homologous genes including 1 BrDIR2 in turnip (Thamil et al., 2013), DIR1 in Gossypium hirsutum (Shi et al., 2012), while in Triticum aestivum (Subramanyam et al., 2006(Subramanyam et al., , 2008Song et al., 2013;Ma and Liu, 2015) Hfr1, TaDIR13 and having Dir domain in JRL genes, Another DIR1 in Boea hygrometrica , DIR22 in Glycine max , DIR factor in Tamarix androssowii (Gao et al., 2010) and another DIR factor in Sugarcane (Guo et al., 2012). Various Dirrigent proteins, including SHDIR11 and SHDIR16 transcription in Saccharum officinarum triggered by salicylic acid, Jasmonic acid, and Methyl Jasmonate, also react to the production of diseaseassociated hormonal substances (Damaj et al., 2010). ...
Dirrigent proteins are required for the synthesis of lignans, a distinct and widely distributed class of plant-derived secondary metabolites with promising pharmacologic characteristics and a potential role in plant defense. However, no detailed data on the Dirrigent family of Solanum tuberosum is known. Comparatively, 33 DIR sequences were explored in Solanum tuberosum in this research study. There are no introns in the majority of StDIR genes. A single Dirigent domain is found in all StDIR proteins. The abundance of amino acids, sequence similarity analysis, phylogenetic analysis, genomic location, gene structure, conserved domain analysis, evolutionary analysis, and gene promoter assessment are all studied. This study lays a foundation for potential plant genetic engineering and crop improvement research by providing an in-depth and thorough explanation of the detailed molecular mechanism and structural characterization of StDIR proteins in the genome of Solanum tuberosum. This work will provide useful data for enabling the proper selection of dirrigent proteins in higher plants and will support future studies on the DIR gene family.
... In Arabidopsis, four antagonistic JRLs, namely, JAL31, ATJAL23, PBP1/AtJAL30, and AtJAL22, regulate the size of ER body-type b-glucosidase complexes [9], and At-JAC1 functions in flowering time control [14]. Some JRLs were related to biotic stress responses [15][16][17][18]. The expression of OsJRL genes was induced by Magnaporthe grisea infection [19]. ...
The jacalin-related lectins (JRLs) are widely distributed in plants and are involved in plant development and multiple stress responses. However, the characteristics of the HvJRL gene family at the genome-wide level and the roles of JRLs in barley’s response to low-nitrogen (LN) stress have been rarely reported. In this study, 32 HvJRL genes were identified and unevenly distributed at both ends of the seven chromosomes in barley. HvJRL proteins generally exhibited low sequence similarity but shared conserved jacalin domains by multiple sequence analysis. These proteins were classified into seven subfamilies based on phylogenetic analysis, with a similar gene structure and conserved motifs in the same subfamily. The HvJRL promoters contained a large number of diverse cis-elements associated with hormonal response and stress regulation. Based on the phylogenetic relationships and functionally known JRL homologs, it was predicted that some HvJRLs have the potential to serve functions in multiple stress responses but not nutrition deficiency stress. Subsequently, nine differentially expressed genes (DEGs) encoding eight HvJRL proteins were identified in two barley genotypes with different LN tolerance by transcriptome analysis. Furthermore, 35S:HvHorcH transgenic Arabidopsis seedlings did enhance LN tolerance, which indicated that HvHorcH may be an important regulator of LN stress response (LNSR). The HvJRL DEGs identified herein could provide new candidate genes for LN tolerance studies.
... Of the eight associated with 'biotic stress', six coded for previously described papain-like cysteine proteases, a dicer protein and LOL1 isoform X2. Two additional downregulated transcripts coded for binding to TOMV RNA 1 (BTR1) [34] ('transcription' GO-group) and functionally characterised non-classified T. aestivum hessian fly response gene 1 protein (TaHfr1, [35]; not classified in GO-group). ...
The wheat NAC transcription factor TaNACL-D1 enhances resistance to the economically devastating Fusarium head blight (FHB) disease. The objective of this study was to decipher the alterations in gene expression, pathways and biological processes that led to enhanced resistance as a result of the constitutive expression of TaNACL-D1 in wheat. Transcriptomic analysis was used to determine the genes and processes enhanced in wheat due to TaNACL-D1 overexpression, both in the presence and absence of the causal agent of FHB, Fusarium graminearum (0- and 1-day post-treatment). The overexpression of TaNACL-D1 resulted in more pronounced transcriptional reprogramming as a response to fungal infection, leading to the enhanced expression of genes involved in detoxification, immune responses, secondary metabolism, hormone biosynthesis, and signalling. The regulation and response to JA and ABA were differentially regulated between the OE and the WT. Furthermore, the results suggest that the OE may more efficiently: (i) regulate the oxidative burst; (ii) modulate cell death; and (iii) induce both the phenylpropanoid pathway and lignin synthesis. Thus, this study provides insights into the mode of action and downstream target pathways for this novel NAC transcription factor, further validating its potential as a gene to enhance FHB resistance in wheat.
... The absence of this genefor-gene recognition yields a compatible interaction where the plants are susceptible in response to HF larval attack. During incompatible interactions, the resistant plants mount defense responses (Liu et al., 2007;Sardesai et al., 2005;Subramanyam et al., 2006Subramanyam et al., , 2008Subramanyam et al., , 2013) that prevent the avirulent larvae from feeding, leading to their death within 4-5 days after egg hatch (DAH), and the wheat seedlings show normal growth (Gallun, 1977). In contrast, during compatible interactions, the larval salivary secretions suppress the plant immune response and alter host plant physiology, resulting in development of a nutritive tissue that is a source of diet rich in proteins and sugars, allowing the virulent larvae to complete their development, whereas the growth of the susceptible plants is stunted (Harris et al., 2006;Puthoff et al., 2005;Subramanyam & Nemacheck, 2021b;Subramanyam et al., 2015Subramanyam et al., , 2018Subramanyam et al., , 2021Williams et al., 2011). ...
Hessian fly (HF) (Mayetiola destructor Say) is an obligate destructive pest of wheat (Triticum aestivum L.) causing severe economic losses worldwide. Deployment of resistant wheat cultivars harboring HF resistance (H) genes is still the most effective and economical method to manage this insect pest. However, extensive use of H genes can impose selection pressure on HF populations, leading to the development of virulent biotypes that can breakdown plant resistance. Further, increase in environmental temperatures to 25–30°C during the wheat‐growing season can also negatively impact HF resistance, thereby necessitating the identification of new and novel sources of HF resistance and evaluate them for temperature sensitivity. In the current study, we evaluated the phenotypic response of 254 Triticum turgidum (L.) wheat accessions of African origin to Biotype L HF infestation. Of these, 10 accessions exhibited >70% resistance to Biotype L HF infestation. These 10 resistant accessions also showed >70% resistance when screened with vH13 HF biotype. Additionally, these HF resistant lines were evaluated for expression of resistance at a higher temperature of 30°C, and three tetraploid wheat accessions that maintained 100% resistance to Biotype L HF at the increased temperature were identified. These newly identified HF resistant lines offer valuable tools to breeders and farmers that can be used in breeding programs to develop cultivars for durable resistance to HF and efficient crop management.
... As its name suggests, it is a fusion of two proteins, a jacalin-related lectin (JRL) and a dirigent protein (DIR) at the N-terminus. Many representatives of this protein class were found in different monocot plants, such as AsCrs-1 [4] in creeping bentgrass (Agrostis stolonifera), PeD-J [5] in Moso bamboo (Phyllostachys edulis), TaVer2 [6] and TaHFR1 [7] in wheat (Triticum aestivum), SbSL [8] in sorghum (Sorghum bicolor), ZmBGAF [9] in maize (Zea mays), ShDJ [10] in sugarcane (Saccharum hybrid cultivar), and OsJAC1 [11] in rice (Oryza sativa). Recently, Ma and Han (2021) identified 46 MCJ genes from the wheat genome and divided the class into three subfamilies [12]. ...
... Among the two MCJ domains, the JRL domain is better characterized, particularly regarding its carbohydrate binding properties. Previously identified interactions displayed specificity toward mannose [7,11,12,15], galactose [9,12], N-acetylgalactosamine [8], or N-acetylglucosamine [6]. ...
Pesticides are routinely used to prevent severe losses in agriculture. This practice is under debate because of its potential negative environmental impact and selection of resistances in pathogens. Therefore, the development of disease resistant plants is mandatory. It was shown that the rice (Oryza sativa) protein OsJAC1 enhances resistance against different bacterial and fungal plant pathogens in rice, barley, and wheat. Recently we reported possible carbohydrate interaction partners for both domains of OsJAC1 (a jacalin-related lectin (JRL) and a dirigent (DIR) domain), however, a mechanistic understanding of its function is still lacking. Here, we report crystal structures for both individual domains and the complex of galactobiose with the DIR domain, which revealed a new carbohydrate binding motif for DIR proteins. Docking studies of the two domains led to a model of the full-length protein. Our findings offer insights into structure and binding properties of OsJAC1 and its possible function in pathogen resistance.
... The downregulation of DMR6 and dirigent protein genes suggests that heat stress may delay the process of producing defensive secondary metabolites and weaken cell wall fortification. Hessian fly responsive gene 1 was induced in wheat plants during the incompatible interaction, and its protein possesses lectin-like domains and insecticide activity [61]. The downregulation of Hessian fly responsive gene 1 by heat stress in our study may suggest weaker antibiotic resistance in heat-stressed wheat plants to HF infestation. ...
Heat stress compromises wheat (Triticum aestivium) resistance to Hessian fly (HF, Mayetiola destructor (Say)). This study aimed to investigate the impact of heat stress on transcript expression of wheat genes associated with resistance to HF infestation under normal and heat-stressed conditions. To this end, ‘Molly’, a wheat cultivar containing the resistance gene H13, was subjected to HF infestation, heat stress, and the combination of HF infestation and heat stress. Our RNA-Seq approach identified 21 wheat genes regulated by HF infestation under normal temperatures (18 °C) and 155 genes regulated by HF infestation when plants were exposed to 35 °C for 6 h. Three differentially expressed genes (DEGs) from the RNA-Seq analysis were selected to validate the gene function of these DEGs using the RT-qPCR approach, indicating that these DEGs may differentially contribute to the expression of wheat resistance during the early stage of wheat–HF interaction under various stresses. Moreover, the jasmonate ZIM domain (JAZ) gene was also significantly upregulated under these treatments. Our results suggest that the genes in heat-stressed wheat plants are more responsive to HF infestation than those in plants growing under normal temperature conditions, and these genes in HF-infested wheat plants are more responsive to heat stress than those in plants without infestation.
... Lectin proteins that bind to carbohydrates, monosaccharides and complex glycans also show inducible character by various stresses, including herbivory in wheat (Subramanyam et al., 2008) and tobacco (Vandenborre et al., 2009). Accordingly, lectins have broad insecticidal activity in plants (reviewed by Michiels et al., 2010), as originally reported by Janzen et al. (1976), using phytohemagglutinin from black beans (P. ...
Success of plants largely depends on their ability to defend against herbivores. Since emergence of the first voracious consumers, plants maintained adapting their structures and chemistry to escape from extinction. The constant pressure was further accelerated by adaptation of herbivores to plant defenses, which all together sparked the rise of a chemical empire comprised of thousands of specialized metabolites currently found in plants. Metabolic diversity in the plant kingdom is truly amazing, and although many plant metabolites have already been identified, a large number of potentially useful chemicals remain unexplored in plant bio‐resources. Similarly, biosynthetic routes for plant metabolites involve many enzymes, some of which still wait for identification and biochemical characterization. Moreover, regulatory mechanisms that control gene expression and enzyme activities in specialized metabolism of plants are scarcely known. Finally, understanding of how plant defense chemicals exert their toxicity and/or repellency against herbivores remains limited to typical examples, such as proteinase inhibitors, cyanogenic compounds and nicotine. In this review, we attempt summarizing the current status quo in metabolic defense of plants that is predominantly based on the survey of ubiquitous examples of plant interactions with chewing herbivores.
... Recent research on plant JRLs has focused on their functions in stress response . For example, the wheat TaHfr-1 gene is induced by Mayetiola destructor feeding (Subramanyam et al., 2008), and the O. sativa Salt gene is induced by drought and hormone treatment and shows high tissue specificity (Jiang et al., 2006). The Arabidopsis RTM1 protein is similar to JRL, and its corresponding gene is induced by tobacco etch virus (TEV), inhibiting its longrange transmission (Chisholm et al., 2001). ...
Jacalin-related lectins (JRLs) are a new subfamily of plant lectins that has recently been recognized and plays an important role in plant growth, development, and abiotic stress response. Although moso bamboo (Phyllostachys edulis) is an economically and industrially important bamboo worldwide, there has been no systematic identification of JRLs in this species. Here, we identified 25 JRL genes in moso bamboo, and these genes are unequally distributed among 10 genome scaffolds. Phylogenetic analysis showed that the moso bamboo JRLs were clustered into four JRL subgroups: I, II, V, and VII. Numerous stress-responsive and hormone-regulated cis-elements were detected in the upstream promoter regions of the JRLs. Genome collinearity analyses showed that the JRL genes of moso bamboo are more closely related to those of Brachypodium distachyon than to those of Oryza sativa and Zea mays. Sixty-four percent of the PeJRL genes are present as segmental and tandem duplicates. qRT-PCR expression analysis showed that JRL genes in the same subgroup were significantly downregulated in response to salicylic acid (SA), abscisic acid (ABA), and methyl jasmonate (MeJA) treatments and significantly upregulated under low temperature, drought, and salt stress; they also exhibited tissue-specific expression patterns. Subcellular localization experiments revealed that PeJRL04 and PeJRL13 were localized to the cell membrane, nucleus, and cytoplasm. Three dimensional structure prediction and yeast two-hybrid assays were used to verify that PeJRL13 exists as a self-interacting homodimer in vivo. These findings provide an important reference for understanding the functions of specific moso bamboo JRL genes and for the effective selection of stress-related genes.
... The recognition of insect avirulence (avr) gene product by plant Hessian fly resistance (H) gene product [3] yields an incompatible interaction (avirulent larvae, resistant plants). In this interaction, the wheat seedling mounts H-gene-mediated resistance, resulting in increased accumulation of defense proteins [4][5][6][7] that disrupt larval midgut microvilli [8]. The avirulent larvae die within 4-5 days after egg-hatch (DAH), and the resistant plants show normal growth. ...
... Chymotrypsin and trypsin serine proteases are demonstrated to be the major digestive enzymes in the gut of Hessian fly larvae [49]. Fewer up-regulated genes and lower expression levels in A3 larvae likely result in poor protease activity, which is correlated with the unsuccessful attempt to establish feeding sites by the avirulent larvae at 3 DAH [6]. In addition to trypsin and chymotrypsin, several genes encoding clip-domain proteases were also significantly up-regulated in V3 and Bd3 larvae that were not observed in A3 larvae (Figure 7a). ...
The Hessian fly is a destructive pest of wheat. Employing additional molecular strategies can complement wheat’s native insect resistance. However, this requires functional characterization of Hessian-fly-responsive genes, which is challenging because of wheat genome complexity. The diploid Brachypodium distachyon (Bd) exhibits nonhost resistance to Hessian fly and displays phenotypic/molecular responses intermediate between resistant and susceptible host wheat, offering a surrogate genome for gene characterization. Here, we compared the transcriptomes of Biotype L larvae residing on resistant/susceptible wheat, and nonhost Bd plants. Larvae from susceptible wheat and nonhost Bd plants revealed similar molecular responses that were distinct from avirulent larval responses on resistant wheat. Secreted salivary gland proteins were strongly up-regulated in all larvae. Genes from various biological pathways and molecular processes were up-regulated in larvae from both susceptible wheat and nonhost Bd plants. However, Bd larval expression levels were intermediate between larvae from susceptible and resistant wheat. Most genes were down-regulated or unchanged in avirulent larvae, correlating with their inability to establish feeding sites and dying within 4–5 days after egg-hatch. Decreased gene expression in Bd larvae, compared to ones on susceptible wheat, potentially led to developmentally delayed 2nd-instars, followed by eventually succumbing to nonhost resistance defense mechanisms.
... The wheat-Hessian fly interaction fits the gene-for-gene model (Hatchett and Gallun 1970) resulting in either a compatible or incompatible interaction. During incompatible interactions, the H (Hessian fly resistance) gene-mediated resistance is accompanied by increase in accumulation of transcripts encoding plant defense proteins Subramanyam et al. 2006Subramanyam et al. , 2008Subramanyam et al. , 2013) that disrupt the insect midgut microvilli (Shukle et al. 2010) resulting in larval death (avirulent larvae) while the resistant plant (harboring the H gene) shows normal growth (Gallun 1977). However, during compatible interactions, larvae inject salivary effector proteins (Chen et al. 2006) that alter the host plant physiology and suppress plant defense responses (Baluch et al. 2012). ...
Insect UDP-glycosyltransferases (UGTs) play an important role in detoxification of substrates such as plant allelochemicals, and cuticle formation by the process of glucosidation. Hessian fly ( Mayetiola destructor ), belonging to the order Diptera (Family: Cecidomyiidae), is a destructive pest of host wheat causing significant economic losses. In the current study, using the assembled genome, we identified thirteen genes in M. destructor that belong to the family of UGTs ( MdesUGT ). Expression profiling revealed differential expression of MdesUGT genes in Hessian fly feeding instars. Further, we report the molecular cloning of MdesUGT1 , designated as UGT301F1, from M. destructor . Characterization of the MdesUGT1 amino acid sequence revealed a conserved signature motif and sugar donor-binding domains characteristic of UGT proteins. Further expression analysis revealed dramatic increase in transcript accumulation of MdesUGT1 in the first and second feeding instars during compatible interactions (susceptible wheat, virulent larvae) but lacked significant upregulation during incompatible wheat Hessian fly interactions. Similar increase in MdesUGT1 transcripts was also observed during interactions of Hessian fly with nonhost, Brachypodium distachyon . These findings suggest the possible early involvement of MdesUGT1 in detoxification of plant toxins, and subsequent role in cuticular formation, thus contributing to the growth and development of this dipteran insect pest. Identification and characterization of insect UGTs could provide valuable insights into the detoxification and growth inhibitory mechanisms and facilitate future plant pest management strategies.
