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Comparison of spinodal decomposition in TEM photomicrographs of S. canicula during spermiogenesis (left column) [86], with polarized optical photomicrographs of the time evolution of a thermally quenched polymer dispersed liquid crystal (PDLC) undergoing either spinodal decomposition (center column: 40/60 wt %) [[91], Fig. 2] or nucleation and growth (right column: 60/40 wt % ) [[91], Fig. 3]. A sketch of 1/interaction parameter χ (chi)-concentration C, the phase diagram [[93], Fig. 3.5] for a system of two components (A and B) is shown on the left, where χ is the energy change "when a molecule of A is taken from an environment of pure A and put into an environment of pure B" [[93], p.28], so that χ "expresses the strength of the energetic interaction between the components". C is the concentration, represented as the mole fraction of A running from 0 to 1 ([79], Fig. 10). In the phase diagram on the left, the composition S changes from a value of 1/χ (sub 2) for a single phase to a value of 1/χ (sub 1) for compositions P1 (B-rich phase) and P2 (A-rich phase) that places the system into the unstable region where it separates into two phases by spinodal decomposition [[79], Fig. 10]. Such a separation can occur isothermally during spermiogenesis in S. canicula (left column; TEM photomicrographs here have been magnified from (Fig. (3), white boxes), or during the time evolution of a cold quenched 40/60 wt % PDLC (middle column) [[91], Fig. 3]. For the latter 1/χ can be replaced by T (absolute temperature) in the phase diagram on the left. During a cold quench of a 60/40 wt % PDLC into the metastable regions of the phase diagram, nucleation and growth occur (right column) [[91], Fig. 2]. The polarized optical photomicrographs in the middle and right columns have been taken from the 1996 paper of Kyu et al. [91], with the permission of the American Chemical Society.
Source publication
Despite their relatively arginine-rich composition, protamines exhibit a high degree of structural variation. Indeed, the primary structure of these histone H1-related sperm nuclear basic proteins (SNBPs) is not random and is the depository of important phylogenetic information. This appears to be the result of their fast rate of evolution driven b...
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... look first at the appearance of bicontinuity in TEM photomicrographs of dogfish spermatids (Fig. (4, left col- umn)), magnified from (Fig. (3)), as well as at the time evo- lution of a polymer dispersed liquid crystal (PDLC; (Fig. (4, center ...
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... look first at the appearance of bicontinuity in TEM photomicrographs of dogfish spermatids (Fig. (4, left col- umn)), magnified from (Fig. (3)), as well as at the time evo- lution of a polymer dispersed liquid crystal (PDLC; (Fig. (4, center ...
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... 1978 paper of Gusse and Chevaillier [86], Fig. 1, 6, 15, 18, 24, 25]. Note the concomitant appearance (right) of histones (H) [13], intermediate proteins (S1, S2) [88,89] and protamines (Z1, Z2, Z3, S4) [88][89][90]. Symbols to the left of the TEM photomicrographs are defined in (Fig. (5, left). White boxes are shown at higher magnification in Fig. (4)). (Fig. (3E, F) tem of two components (Fig. (4, left)). We plot 1/ on the vertical axis rather than the more classical temperature be- cause, according to Harrison et al. [ [79], Fig. 10]: "Similar behavior can occur in a system of more than two compo- nents, and as a result of, e.g., a chemical change in the sys- tem rather than ...
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... 1, 6, 15, 18, 24, 25]. Note the concomitant appearance (right) of histones (H) [13], intermediate proteins (S1, S2) [88,89] and protamines (Z1, Z2, Z3, S4) [88][89][90]. Symbols to the left of the TEM photomicrographs are defined in (Fig. (5, left). White boxes are shown at higher magnification in Fig. (4)). (Fig. (3E, F) tem of two components (Fig. (4, left)). We plot 1/ on the vertical axis rather than the more classical temperature be- cause, according to Harrison et al. [ [79], Fig. 10]: "Similar behavior can occur in a system of more than two compo- nents, and as a result of, e.g., a chemical change in the sys- tem rather than change of temperature." Thus, spinodal de- composition can ...
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... temperature be- cause, according to Harrison et al. [ [79], Fig. 10]: "Similar behavior can occur in a system of more than two compo- nents, and as a result of, e.g., a chemical change in the sys- tem rather than change of temperature." Thus, spinodal de- composition can occcur isothermally [83,84] in the unstable region of the phase diagram ( Fig. (4, left)), as seen by TEM photomicrographs for spermiogenesis in S. canicula in (Fig. (4, left column)). It can also be observed in optical light pho- tomicrographs for a 40/60 wt % PDLC after thermal quench- ing in (Fig. (4, center column)). This is a PDLC that consists of 40 wt % polymethyl methacrylate with hydroxyl groups in the liquid form ...
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... can occur in a system of more than two compo- nents, and as a result of, e.g., a chemical change in the sys- tem rather than change of temperature." Thus, spinodal de- composition can occcur isothermally [83,84] in the unstable region of the phase diagram ( Fig. (4, left)), as seen by TEM photomicrographs for spermiogenesis in S. canicula in (Fig. (4, left column)). It can also be observed in optical light pho- tomicrographs for a 40/60 wt % PDLC after thermal quench- ing in (Fig. (4, center column)). This is a PDLC that consists of 40 wt % polymethyl methacrylate with hydroxyl groups in the liquid form / 60 wt % E7, a mixture of nematic liquid crystals (that includes derivatives of ...
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... of temperature." Thus, spinodal de- composition can occcur isothermally [83,84] in the unstable region of the phase diagram ( Fig. (4, left)), as seen by TEM photomicrographs for spermiogenesis in S. canicula in (Fig. (4, left column)). It can also be observed in optical light pho- tomicrographs for a 40/60 wt % PDLC after thermal quench- ing in (Fig. (4, center column)). This is a PDLC that consists of 40 wt % polymethyl methacrylate with hydroxyl groups in the liquid form / 60 wt % E7, a mixture of nematic liquid crystals (that includes derivatives of cyanobiphenyl, oxycya- nobiphenyl and cyanoterphenyl) with a broad nematic tem- perature range [ Fig. 3.5] for a system of two components (A and B) is ...
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... typical appearance of bicontinuity [83] is readily apparent for both the magnification of step D in the pattern- ing stage of spermiogenesis in S. canicula (Fig. (4, left col- umn)), as well as at 120 minutes for the 40/60 PDLC ( Fig. (4, center column)). Such bicontinuity lasts for a few hours in the PDLC [[91], p.208]. How long it lasts in S. canicula spermiogenesis is not ...
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... typical appearance of bicontinuity [83] is readily apparent for both the magnification of step D in the pattern- ing stage of spermiogenesis in S. canicula (Fig. (4, left col- umn)), as well as at 120 minutes for the 40/60 PDLC ( Fig. (4, center column)). Such bicontinuity lasts for a few hours in the PDLC [[91], p.208]. How long it lasts in S. canicula spermiogenesis is not ...
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... bicontinuity is, however, typical for spinodal de- composition, as compared to nucleation and growth for the 60/40 PDLC after thermal quenching, where droplets are observed in (Fig. (4, right column)) due to Ostwald's ripen- ing mechanism [93]. As Kyu et al. state, regarding biconti- nuity in the 40/60 PDLC [[91], p.206], "This interconnected structure is reminiscent of a spinodal structure. The length scale of this structure increases with elapsed time and even- tually the pattern transforms into droplet morphology proba- bly ...
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... we can see in a transverse section of step D of S. canicula spermiogenesis in both (Fig. (3)) and its magnifica- tion in (Fig. (4)), that all lamellae are oriented to the nuclear envelope in a parallel manner. This is different than in the muricid snail M. brandaris [76][77][78], where all lamellae in a transverse section run perpendicular [79], Fig. 9] to both the nuclear envelope and the axoneme. This is also the case for another muricid snail, N. lamellosa ...
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... steps B, C, D in the patterning stage of S. canicula spermiogenesis in (Fig. (3), as magnified in (Fig. (4)), appear to show a constancy of spacing from the middle of one la- mella to the middle of an adjacent one, as diagrammed in (Fig. (5, left.)) This is , the unit of pattern that, according to Cahn's [81] thermodynamic analysis of spinodal decomposi- tion, shows a maximal value when plotted against a growth rate constant [[79], Fig. 8]. ...
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... order to visualize this more readily, one of us (H.E.K.) has measured in every panel in (Fig. (4)) using a finely calibrated Staedtler steel ruler. In (Fig. (5, left column)), we can readily see from the histograms that m for S. canicula spermiogenesis (shaded bars) is constant for steps A-D (3540 nm, 30-35 nm, 30-35 nm, 30-35 nm) during the pattern- ing stage, then diminishes to 10-15 nm in step E (measured from [[86], Fig. 26, ...
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... histograms of the 40/60 PDLC (Fig. 5, center col- umn) also show relatively constant values for m for the opti- cal photomicrographs of (Fig. 4, center column) from 3 min- utes to 120 minutes (2.5-3 μm, 2.5-3 μm, 3.5-4 μm, 3.5-4 μm). After this time, heterogeneous droplets begin to ap- pear at 300 minutes and particularly at 1020 minutes in (Fig. (5, center column)), yielding "twin towers" (3.5-4 μm and 4.5-5μm), and then a very dispersed histogram (3-13.5 μm). This final step ...
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... dogfish S. canicula and polar- ized optical photomicrographs of PDLC. Features are indicated in the diagram (on the left) for which (lambda, a unit of pattern) can be measured during spermiogenesis from the middle of the black chromatin stripe, dot or doughnut-like formation, across the white nucleo- plasm, to another formation of chromatin in (Fig. (4)), along with the diameter (d) of the chromatin fiber, the width (w) of the nucleoplasm and the diameter of the nucleus (n.d.). can also be measured during the time evolution of a PDLC undergoing either spinodal decomposi- tion or nucleation and growth. The shaded bar in each histogram represents m (lambda maximum, the dominant pattern ...
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... We speculate that the species in (Table (I)) displaying phase separation by spi- nodal decomposition during spermiogenesis are just a small subset of internally fertilizing species that happen to have the appropriate biochemistry in their histone-to-protamine tran- sition to fall into the unstable portion of the phase diagram ( Fig. (4, ...
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... example, amongst the internally fertilizing chon- drichthyan fish (Table (I)), we have already seen (Fig. (4)) that the spermatid of the elasmobranch dogfish S. canicula displays the bicontinuous lamellar chromatin/nucleoplasm patterning typical of phase separation by spinodal decompo- sition, while undergoing a transition from histones to inter- mediate proteins to protamine and keratinous protamines (Fig. (3)). The spermatid of the ...
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... even here small differences in the sequence of "simple" protamines makes spermiogenesis in the externally fertilizing bony fish Dicentrarchus labrax [[55], Fig. 5] look more like nucleation and growth (Fig. (4, right column)), while externally fertilizing rainbow trout (Salmo gairdneri) [[107], Fig. 18] is more complex than that, developing from coarse granules into thick fibers. However, both species definitely lack the bicontinuous lamellae characteristic of spinodal decomposition (Fig. (4, left and center ...
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... labrax [[55], Fig. 5] look more like nucleation and growth (Fig. (4, right column)), while externally fertilizing rainbow trout (Salmo gairdneri) [[107], Fig. 18] is more complex than that, developing from coarse granules into thick fibers. However, both species definitely lack the bicontinuous lamellae characteristic of spinodal decomposition (Fig. (4, left and center ...
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... bony fish, display features of nucleation and growth of chromatin granules during spermiogenesis. In mammalian species chromatin forms 60- 100 nm toroids [1,56,109] rather than bicontinuous pat- terns. While a complete explanation for this is not as yet available, there are several considerations worth noting. First, spinodal decomposition ( Fig. (4, left and center columns)), and nucleation and growth (Fig. (4, right col- umn)) may represent the extremes of a single mechanism of phase separation rather than separate processes. As Gunton et al. [[110], p.364] point out: "Thus in the classical pictures of nucleation and of spinodal decomposition, there is a sharp transition between metastable and unstable ...
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... granules during spermiogenesis. In mammalian species chromatin forms 60- 100 nm toroids [1,56,109] rather than bicontinuous pat- terns. While a complete explanation for this is not as yet available, there are several considerations worth noting. First, spinodal decomposition ( Fig. (4, left and center columns)), and nucleation and growth (Fig. (4, right col- umn)) may represent the extremes of a single mechanism of phase separation rather than separate processes. As Gunton et al. [[110], p.364] point out: "Thus in the classical pictures of nucleation and of spinodal decomposition, there is a sharp transition between metastable and unstable states as charac- terized by the classical spinodal ...
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... species, Therefore, we put forward the hypothesis that lamellar chromatin/nucleoplasm patterning during spermiogenesis may be the visual manifestation of the mechanism of liquid-liquid phase separation by spinodal decomposition when the appropriate concentrations of com- plex protamines and DNA fall into the unstable region of the phase diagram (Fig. (4. left)). This occurs during histone-to- protamine replacement, and SNBP processing and side chain modification of spermiogenic chromatin. Such an hypothesis complements our present knowledge of phase separation in the metastable portion of the phase diagram by nucleation and growth during mammalian spermiogenesis ...
Citations
... Still, they exhibit much more structural variability than histones and can range in mass between 6 and 40 kDa. Finally, protamines (Kasinsky et al., 2012) are highly arginine-rich. ...
... This is a dynamic process that, at the biochemical level involves a massive degradation of most of the genomic histones and post-translational processing of both chromosomal proteins involved (histone acetylation and protamine phosphorylation and cleavage) (Fig.4). Hence, this process is not only highly dynamic but it is also very complex (Kasinsky et al., 2012). The difficulty in the study of the biochemical events involved in the chromatin transitions at the different stages of spermiogenesis, particularly in invertebrates, where only low amounts of sample are usually available, arises from the difficulty in fractionating the different cell types at each stage. ...
While there is extensive information about sperm nuclear basic proteins (SNBP) in vertebrates, there is very little information about Arthropoda by comparison. This paper aims to contribute to filling this gap by analyzing these proteins in the sperm of the noble false widow spider Steatoda nobilis (Order Araneae, Family Theridiidae). To this end, we have developed a protein extraction method that allows the extraction of cysteine-containing protamines suitable for the preparation and analysis of SNBPs from samples where the amount of starting tissue material is limited. We carried out top-down mass spectrometry sequencing and molecular phylogenetic analyses to characterize the protamines of S. nobilis and other spiders. We also used electron microscopy to analyze the chromatin organization of the sperm, and we found it to exhibit liquid-liquid phase spinodal decomposition during the late stages of spermiogenesis. These studies further our knowledge of the distribution of SNBPs within the animal kingdom and provide additional support for a proposed evolutionary origin of many protamines from a histone H1 (H5) replication-independent precursor.
... Although of a different nature, a similar lack of techniques to analyze, in this case, the minute amounts of SNBPs from the small amounts of sperm obtained from insects hindered the steady pace of the work. The high abundance of arginine in canonical protamines (Balhorn 2007;Kasinsky et al. 2012), similar to ribosomal proteins, additionally complicated the issue. Only with the advent of powerful techniques such as intact protein sequencing by liquid chromatography-nanoelectrospray ionization-mass spectrometry (LCMS) (Coon et al. 2005) has it been ultimately possible to overcome such difficulties (D'Ippolito et al. 2019). ...
... Regardless of all the difficulties encountered in their identification, the SNBP P1 and P2 from A. mellifera are very similar both in size and composition to the arginine-rich protamines of other invertebrate and vertebrate organisms (Kasinsky et al. 2012), and they are likely the products of extensive protein precursor processing. It is this processing, that might ultimately be responsible for the complex chromatin organization transitions observed during spermiogenesis (Fig. 2F) which are characteristic of insects (Kasinsky et al. 2021). ...
... As shown in Fig. 7E, the two bands in Fig. 7B PC consist of a mixture of at least four different proteins with very strong sequence similarity and several amino acid substitutions, suggesting the existence of multiple genes encoding for protamines with sequence microheterogeneity. Their true protamine nature is revealed by their high arginine clusters, as observed in other invertebrate and vertebrate protamines (Lewis et al. 2003;Kasinsky et al. 2012). ...
Insects are the largest group of animals when it comes to the number and diversity of species. Yet, with the exception of Drosophila, no information is currently available on the primary structure of their sperm nuclear basic proteins (SNBPs). This paper represents the first attempt in this regard and provides information about six species of Mecoptera: Poecillimon thessalicus, Graptosaltria nigrofuscata, Apis mellifera, Nasonia vitripennis, Parachauliodes continentalis, and Tribolium castaneum. The SNBPs of these species were characterized by acetic acid urea gel electrophoresis (AU-PAGE) and HPLC fractionated. Protein sequencing was obtained using a combination of mass spectrometry sequencing, Edman N-terminal degradation sequencing and genome mining. While the SNBPs of several of these species exhibit a canonical arginine-rich protamine nature, a few of them exhibit a protamine-like (PL) composition. They appear to be the products of extensive cleavage processing from a precursor protein which are sometimes further processed by other post-translational modifications (PTMs) that are likely involved in the chromatin transitions observed during spermiogenesis in these organisms.
... This is known to occur in sperm of echinoderms (Poccia and Green 1992;Poccia 1995). Protamines (P type) are relatively small arginine-rich proteins that replace the somatic histone complement during spermiogenesis (Balhorn 2007;Kasinsky et al. 2012) resulting in small nuclei with highly compacted chromatin. Protamine-like proteins (PL-type) are enriched in both lysine and arginine. ...
Sperm nuclear basic proteins (SNBPs) were isolated from extracted antheridia-rich male gametophytes raised from spores of the swordfern, Polystichum munitum. Electrophoretic (acetic acid-urea PAGE and SDS-PAGE) and chromatographic (rp-HPLC) characterization of the nuclear proteins exhibited the characteristics of the histone (H-type). In both types of gel electrophoresis, Histones H1, H2A, and H2B showed an altered electrophoretic mobility corresponding to that which is routinely observed for the histones in other plants. Histones present during spermatogenesis of the fern P. munitum were compared with the few current SNBPs known to be present in higher and lower evolutionary plant clades. A transition from an early protamine (P-type) SNBPS in charophytes and bryophytes to the (H-type) SNBP observed here is reminiscent of similar reversions observed in the animal kingdom.
... [25][26][27][28] In the early-to-middle stages of the spermatogenesis before completing the DNA condensation, some species exhibit patterns reminiscent of spinodaldecomposition-type phase separation. [29][30][31][32] Such patterning is most likely a biomolecular condensation induced either by changes of interaction upon histone-to-protamine replacement or by posttranslational modification of protamine such as a series of phosphorylation-dephosphorylation cycles occurring to organize protamine on DNA in human. [33][34][35] Both mechanisms would effectively modulate the R +/ ratio. ...
Protamine, a small, strongly positively-charged protein, plays a key role in achieving chromatin condensation inside sperm cells and is also involved in the formulation of nanoparticles for gene therapy and...
... We originally speculated that PSR might evade self-elimination by not participating in the histone-to-protamine transition, as is the case for certain chromosomal regions and whole chromosomes in other organisms (Raychaudhuri et al. 2012;Rathke et al. 2014). It was not possible to test this hypothesis by manipulating the wasp's protamines because they have not yet been identified; efforts to bioinformatically identify the protamine gene(s) have been hindered presumably because SNBPs are known to evolve rapidly, obscuring signatures of homology (Retief et al. 1993;Kasinsky et al. 2011;Oliva and Dixon 1991;Wyckoff et al. 2000). However, as an alternative, we post-transcriptionally targeted the gene hira, whose encoded protein facilitates the addition of the transitional histone H3.3 to the paternal DNA following removal of protamines in the egg (Bonnefoy et al. 2007). ...
B chromosomes are non-essential, extra chromosomes that can exhibit transmission-enhancing behaviors, including meiotic drive, mitotic drive, and induction of genome elimination, in plants and animals. A fundamental but poorly understood question is what characteristics allow B chromosomes to exhibit these extraordinary behaviors. The jewel wasp, Nasonia vitripennis , harbors a heterochromatic, paternally transmitted B chromosome known as paternal sex ratio (PSR), which causes complete elimination of the sperm-contributed half of the genome during the first mitotic division of fertilized embryos. This genome elimination event may result from specific, previously observed alterations of the paternal chromatin. Due to the haplo-diploid reproduction of the wasp, genome elimination by PSR causes female-destined embryos to develop as haploid males that transmit PSR. PSR does not undergo self-elimination despite its presence with the paternal chromatin until the elimination event. Here we performed fluorescence microscopic analyses aimed at understanding this unexplained property. Our results show that PSR, like the rest of the genome, participates in the histone-to-protamine transition, arguing that PSR does not avoid this transition to escape self-elimination. In addition, PSR partially escapes the chromatin-altering activity of the intracellular bacterium, Wolbachia , demonstrating that this ability to evade chromatin alteration is not limited to PSR’s own activity. Finally, we observed that the rDNA locus and other unidentified heterochromatic regions of the wasp’s genome also seem to evade chromatin disruption by PSR, suggesting that PSR’s genome-eliminating activity does not affect heterochromatin. Thus, PSR may target an aspect of euchromatin to cause genome elimination.
... Via BRDTs, acetylated histones are evicted and replaced on DNA by Transition Proteins (TPs) and Protamines in mammals or Transition protein-like (TPL) and Protamine-like proteins (PLs) in Drosophila. These SNBPs are highly basic, fast-evolving small proteins 24,25 . In addition, Protamines and PLs usually feature multiple cysteine residues, which can form intra-and/or inter-molecular covalent disulfide bonds that participate in Protamines and PLs stability on DNA [26][27][28] . ...
Spermiogenesis is a radical process of differentiation whereby sperm cells acquire a compact and specialized morphology to cope with the constraints of sexual reproduction while preserving their main cargo, an intact copy of the paternal genome. In animals, this often involves the replacement of most histones by sperm-specific nuclear basic proteins (SNBPs). Yet, how the SNBP-structured genome achieves compaction and accommodates shaping remain largely unknown. Here, we exploit confocal, electron and super-resolution microscopy, coupled with polymer modeling to identify the higher-order architecture of sperm chromatin in the needle-shaped nucleus of the emerging model cricket Gryllus bimaculatus. Accompanying spermatid differentiation, the SNBP-based genome is strikingly reorganized as ~25nm-thick fibers orderly coiled along the elongated nucleus axis. This chromatin spool is further found to achieve large-scale helical twisting in the final stages of spermiogenesis, favoring its ultracompaction. We reveal that these dramatic transitions may be recapitulated by a surprisingly simple biophysical principle based on a nucleated rigidification of chromatin linked to the histone-to-SNBP transition within a confined nuclear space. Our work highlights a unique, liquid crystal-like mode of higher-order genome organization in ultracompact cricket sperm, and establishes a multidisciplinary methodological framework to explore the diversity of non-canonical modes of DNA organization.
... Specifically, Adam2 encodes ADAM 2 (a disintegrin and metallopeptidase domain 2), a rapidly evolving, sperm membrane-specific protein with a yet-unclear function in sperm-egg interaction [91,92], and Zp2 encodes ZP2, an egg-specific zona pellucida glycoprotein that mediates sperm adhesion during fertilization [24,63,93,94]. A third germ cell-specific comparison gene, Prm1, has served extensively as a model for rapid evolution of reproductive genes, and encodes protamine 1, which replaces histones in chromatin condensation during spermatogenesis [95][96][97][98][99]. For a somatic, nuclear comparison gene, we chose Tecta, which encodes α-tectorin, a tectorial membrane protein comprising tandem VWD domains paralogous to zonadhesin D0-D4 [100][101][102]. ...
Background
Speciation genes contribute disproportionately to species divergence, but few examples exist, especially in vertebrates. Here we test whether Zan , which encodes the sperm acrosomal protein zonadhesin that mediates species-specific adhesion to the egg’s zona pellucida, is a speciation gene in placental mammals.
Results
Genomic ontogeny reveals that Zan arose by repurposing of a stem vertebrate gene that was lost in multiple lineages but retained in Eutheria on acquiring a function in egg recognition. A 112-species Zan sequence phylogeny, representing 17 of 19 placental Orders, resolves all species into monophyletic groups corresponding to recognized Orders and Suborders, with <5% unsupported nodes. Three other rapidly evolving germ cell genes ( Adam2 , Zp2 , and Prm1 ), a paralogous somatic cell gene ( TectA ), and a mitochondrial gene commonly used for phylogenetic analyses ( Cytb ) all yield trees with poorer resolution than the Zan tree and inferior topologies relative to a widely accepted mammalian supertree. Zan divergence by intense positive selection produces dramatic species differences in the protein’s properties, with ordinal divergence rates generally reflecting species richness of placental Orders consistent with expectations for a speciation gene that acts across a wide range of taxa. Furthermore, Zan ’s combined phylogenetic utility and divergence exceeds those of all other genes known to have evolved in Eutheria by positive selection, including the only other mammalian speciation gene, Prdm9 .
Conclusions
Species-specific egg recognition conferred by Zan ’s functional divergence served as a mode of prezygotic reproductive isolation that promoted the extraordinary adaptive radiation and success of Eutheria.
... Protamines P-type were first described in fish and the name was coined by Miescher (1874) to apply to an organic base derived from the sperm nuclei of Rhine salmon (Salmo salar). These are a group of considerably heterogeneous, and diverse arginine-(and in some instances cysteine-) rich proteins (30-70 % arginine), which are usually smaller than H-type, presenting a small size, ranging from 30 to 100 amino acids, with a molecular mass from 4000 to 10,000 Da, and a higher electrophoretic mobility than H-type ( Fig. 2a) (Kasinsky et al. 2011). ...
... In most cases, PL-type proteins occur simultaneously in the spermatozoa with a full H-type complement (Ausió 1999), accounting for 20-25 % of the total SNBPs. In terms of structure, these proteins often maintain the WHD of linker histones, which serves as a reminder of their evolutionary origin (Kasinsky et al. 2011). ...
... Thus, during spermatogenesis in this cartilaginous fish, two new spermatidspecific intermediate proteins appear at the start of nuclear elongation in spermatids (S1 and S2), which gradually replace somatic histones and are eventually replaced by the final SNBPs, which are composed of a typical fish protamine (Z3) and three keratinous protamines (Z1, Z2, and S4) (Gusse and Chevaillier 1978). S1 and S2 are less basic than P-type and have an intermediate molecular mass, between H and P-type (Kasinsky et al. 2011). These appear just before this event, when the nucleus begins to elongate, and display parallel fibrillar reorganization. ...
One of the most astonishing examples of chromatin remodelling occurs during the maturation of male germ cells, where changes in protein structure, as well as chromatin compaction, take place. During the post-meiotic formation of sperm (spermiogenesis), chromatin appears super-condensed, and transcriptionally inactive, allowing for a more hydrodynamic sperm head, and preventing physical and chemical damage to DNA. DNA is closely condensed in the mature sperm nucleus through its linkage with sperm-specific nuclear proteins called sperm nuclear basic proteins, clustered in three categories: histones, protamines and protamine-like proteins. Fish represent a unique group of vertebrates, including species with specific proteins from each type, such as Sparus aurata, Mullus surmuletus, Dicentrarchus labrax, and Scyliorhinus canicula, which display histones, protamine-like proteins, protamines, and keratinous protamines, respectively. Phylogenetically, these proteins are evolutionarily related, presenting a sporadic and non-random distribution, as a result of vertical evolution, where only histones would be found in more primitive species, and protamines would be restricted to those species located at the uppermost branches of the phylogenetic tree. The relative frequency of this transition is almost insignificant during the differentiation of genera, and species, and very small amongst different families, but, very noticeable amongst different orders. Thus, the aim of this study is to gather the existing background related to sperm nuclear basic proteins in fish, showing a general perspective of the state of the art about diversity, and nuclear reorganization of sperm chromatin during spermiogenesis, and the evolution and phylogenetic distribution of these proteins in fish sperm.
... Protamine is a small 33 amino acid long protein that takes part in DNA packing during spermatogenesis (Kasinsky et al., 2011). Over 60% of the salmon protamine sequence is comprised of arginines so that it interacts strongly with negatively charged DNA phosphates in vivo and potentially with any other negatively charged molecules. ...
Protamine is an arginine-rich peptide that replaces histones in the DNA-protein complex during spermatogenesis. Protamine is clinically used in cardiopulmonary bypass surgery to neutralize the effects of heparin that is required during the treatment. Here we demonstrate that protamine and its 14–22 amino acid long fragments overcome the neurite outgrowth inhibition by chondroitin sulfate proteoglycans (CSPGs) that are generally regarded as major inhibitors of regenerative neurite growth after injuries of the adult central nervous system (CNS). Since the full-length protamine was found to have toxic effects on neuronal cells we used the in vitro neurite outgrowth assay to select a protamine fragment that retains the activity to overcome the neurite outgrowth inhibition on CSPG substrate and ended up in the 14 amino acid fragment, low-molecular weight protamine (LMWP). In contrast to the full-length protamine, LMWP displays very low or no toxicity in our assays in vitro and in vivo. We therefore started studies on LMWP as a possible drug lead in treatment of CNS injuries, such as the spinal cord injury (SCI). LMWP mimicks HB-GAM (heparin-binding growth-associated molecule; pleiotrophin) in that it overcomes the CSPG inhibition on neurite outgrowth in primary CNS neurons in vitro and inhibits binding of protein tyrosine phosphatase (PTP) sigma, an inhibitory receptor in neurite outgrowth, to its CSPG ligand. Furthermore, the chondroitin sulfate (CS) chains of the cell matrix even enhance the LMWP-induced neurite outgrowth on CSPG substrate. In vivo studies using the hemisection and hemicontusion SCI models in mice at the cervical level C5 revealed that LMWP enhances recovery when administered through intracerebroventricular or systemic route. We suggest that LMWP is a promising drug lead to develop therapies for CNS injuries.
... At the biochemical level, spermiogenesis in many animals and plants involves an important chromatin re-organization in which the histones at the onset of the differentiation process get replaced by arginine-rich protamine or protamine-like (PL) proteins in the mature sperm (Ausió, 1999;Kasinsky et al., 2012). Protamines and PLs are members of the important family of intrinsically disordered proteins (IDPs) (Dunker et al., 2001). ...
... While the occurrence of LLPS in somatic chromatin (Gibson et al., 2019) has been recently disputed using both biophysical and in situ cytological approaches , its presence has been often observed in spermiogenic chromatin of different types of organisms, including marine invertebrates, vertebrates (Harrison, 2010;Harrison et al., 2005;Kasinsky et al., 2012;Martens et al., 2009), and in algae (Kasinsky et al., 2014) and plants (D'Ippolito et al., 2019). ...
... We have analyzed published transverse section TEM photomicrographs of spermatids and sperm from entognathus and ectognathus insects. We examined the characteristic features of chromatin/ nucleoplasm patterning due to phase separation by SD or by inversion of a microemulsion [nucleation (Nc)] that we observed previously in our TEM photomicrograph analysis of spermiogenesis in several invertebrate and vertebrate species (Harrison, 2010;Harrison et al., 2005;Kasinsky et al., 2012;Martens et al., 2009).The jewel wasp Nasonia vitripennis, the honey bee Apis mellifera and the fruit fly Drosophila melanogaster, were analyzed in this work. ...
We have examined published transmission electron microscopy (TEM)
photomicrographs of chromatin condensation patterning in developing sperm nuclei from five species of entognathous hexapods within the Classes Protura, Collembola, Diplura and five species of ancestral wingless insects in the Orders Archaeognatha and Zygentoma as well as in fifteen species of the winged insects. Each species reproduces by internal fertilization. Spatially quantitative analysis indicates that spermiogenic chromatin condensation patterning in several of these species may be due to spinodal decomposition (SD) or to microemulsion inversion (chromatin-in-nucleoplasm → nucleoplasm-in-chromatin), also known as nucleation (Nc). These are two different dynamic mechanisms of liquid–liquid phase separation (LLPS). They might either occur independently or co-exist during the chromatin condensation associated with insect spermiogenesis. For example, the chromatin condensation pattern such as that observed in transverse sections of developing sperm nuclei from the wingless insect Anurida maritima (Collembola) is: granules → fibers → lamellae (SD) → nucleation (Nc) → condensed nuclei. Similar transitions are also observed in other more recently evolved species within the Class Insecta. From the limited but comprehensive sample of entognathus and ectognathus hexapods analyzed here, it appears that LLPS of sperm chromatin during spermiogenesis has occurred quite pervasively within the subphylum Hexapoda, including insects.
