Figure 1
Cytoskeletal BtubA/B-candidate structures imaged in Prosthecobacter . Prosthecobacter vanneervenii cells showing tube-like BtubA/B-candidate structures occurring (A) individually or (B) in a bundle. Shown are 11-nm thick slices through cryotomograms. Arrows indicate cytoskeletal structures, which are also shown enlarged below. Asterisk in panel A identifies a sub-tomographic average. Upper-left insets show low- magnification overviews of the cells; rectangles indicate areas imaged in 3-D. Bottom: 3-D segmentation of the bundle of panel B shown from two views (four tubes are present). Scale bars are 100 nm. See Figure S3 for further examples of BtubA/B structures. doi:10.1371/journal.pbio.1001213.g001
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Microtubules play crucial roles in cytokinesis, transport, and motility, and are therefore superb targets for anti-cancer drugs. All tubulins evolved from a common ancestor they share with the distantly related bacterial cell division protein FtsZ, but while eukaryotic tubulins evolved into highly conserved microtubule-forming heterodimers, bacteri...
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... [11,12,24]. To begin, we verified that BtubA and BtubB proteins are in fact expressed in the species where the genes are present (Figures S1 and S2). Western hybridization and PCR also confirmed the absence of BtubA and BtubB in P. fluviatilis ( Figure S2) [24]. Next, Prosthecobacter cells were grown under different conditions and plunge-frozen across EM grids. A total of 589 cells were then imaged in 3-D by ECT. The spindle-shaped cells were polymorphic and exhibited prosthecae (cellular stalks) of different lengths. As seen in other bacterial phyla [25], multiple classes of cytoskeletal structures were seen, but one class had a tube-like morphology and was frequently found in the harboring species, but never in the btubAB -lacking strain (Figure 1). The abundance of these tube-like structures was dependent on the species imaged as well as the growth conditions and growth stage, and was found to be highest in P. vanneervenii cells grown directly on EM grids (67% of cells imaged). In sum, the tube-like structures were found in 48 of 176 P. vanneervenii , 9 of 111 P. dejongeii , 15 of 151 P. debontii , and 0 of 151 P. fluviatilis cells. The tube-like structures were 200– 1,200 nm long, always parallel to the cytoplasmic membrane, almost always localized in the stalk or in the transition zone between stalk and cell body, and occurred either individually or in bundles of two, three, or four (Figure 1, Figure S3, Movie S1). Chemical fixatives were found to degrade the structures (Figure S4), explaining why they were likely missed in previous conventional EM studies [11,22]. Since genetic tools are not yet available for prosthecobacters, we applied labeling and heterologous expression approaches to test whether the candidate structures were in fact composed of BtubA/ B as expected by their correlation with the presence of the genes. Recombinant Escherichia coli cells co-expressing BtubA and BtubB were imaged by ECT and exhibited strikingly similar tube-like structures running the length of the cells (Figure 2A) with the same localization as had been reported for BtubA/B from immuno- fluorescence [19]. Tube-like structures were not seen in control E. coli cells not expressing ButbA/B. Nearly identical tube-like structures were also seen when recombinant BtubA/B was polymerized in vitro and imaged by ECT (Figure 2B). The diameters and subunit repeat distances of all three structures (in Prosthecobacter , recombinant E. coli , and in vitro) were similar (7.6, 7.7, and 7.6 nm diameters, and 4.4, 4.4, and 4.2 nm repeat distances, respectively) (Figures 1, 2, and S3). Finally, immuno- gold-staining using anti-BtubB antibodies localized the proteins to the same region of Prosthecobacter cells as the candidate structures seen by ECT (Figures S5 and S6). We conclude therefore that the tube-like structures are composed of BtubA/B, and the slight differences in repeat distance, straightness, and bundling in the three samples were due to differences in protein concentrations and/or the absence of other interacting proteins in vitro and in E. coli . We have described the BtubA/B structures so far as ‘‘tube-like’’ because when acquiring a cryo-tomographic tilt-series, images of samples tilted beyond , 65 u cannot generally be included, so there is a missing ‘‘wedge’’ of data in reciprocal space that reduces the resolution in the direction of the electron beam. As a result, the ‘‘top’’ and ‘‘bottom’’ boundaries of cylindrical objects (considering the electron beam to be ‘‘vertical’’) are smeared, leaving the sidewalls to appear like two arcs facing each other (Figure 3A–D). Because the opposing arcs observed here were always in this orientation (facing each other and the beam path), it was clear that the structures must have been complete tubes distorted by the missing wedge rather than, for instance, parallel protofilaments, which would not be expected to always orient themselves in the same direction with respect to the electron beam. Nevertheless different orientations of tubes with respect to the tilt axis aggravate the missing wedge artifact differently [26,27], so to explore this effect tomograms of a known, tubular input structure consisting of BtubA/B crystal structures (see below) were simulated at different angles with respect to the tilt axis. These simulations recapitulated the experimental results well, since the density patterns (Figure 3H) were highly similar to those seen in experimental tomograms. To further confirm that the BtubA/B structures were in fact complete tubes and to obtain clearer cross-sectional views, btubAB harboring Prosthecobacter cells, recombinant E. coli cells, and purified BtubA/B polymerized in vitro were all high-pressure-frozen, cryosectioned, and imaged (Figure 3E–G). Cryosections through BtubA/B tubes appeared pentagonal, suggesting five-protofilament tubes. Using the heterodimeric BtubA/B crystal structure [17], we produced tube models with four, five, and six protofilaments for comparison. To maintain reasonable lateral interactions in such small tubes, protofilaments had to be spaced slightly closer (4.6 nm) than protofilaments in eukaryotic microtubules (5 nm), and this resulted in tube diameters of 6.7, 7.8, and 9.2 nm, respectively, for four-, five-, and six-protofilament tubes. Thus only the five-protofilament model was consistent with the 7.6-nm diameter measured in the tomograms, and the five- protofilament model fit the density of the BtubA/B tubes compellingly well (Figure 3I). Cross-sectional views of BtubA/B tubes in cryo-tomograms of whole cells and sub-tomogram averages often showed a left-right asymmetry (arrowheads in Figure 3A–C). Such an asymmetry can only arise from an uneven number of protofilaments, as demonstrated by simulated tomograms (Figure S7), further suggesting five rather than four or six protofilaments. Because the left-right asymmetries in computa- tional projections and in sub-tomographic averages at different positions along the tube axis remained consistent, the five protofilaments must be straight rather than twisting around the tube (Figure S8). Previous EM images of negatively stained, recombinant BtubA/ B polymerized in vitro were not described as tubes, but as protofilament bundles or twisted pairs [17,19,21]. We obtained similar-looking images staining our own purified BtubA/B (Figure S9), but having observed clear tubes in vivo and noting the frequent pairing of parallel densities , 7.6 nm apart in both our negatively stained images and the previously published images, we believe all these samples contained five-protofilament tubes as well. The alternative (two protofilaments 7.6 nm apart) seems unlikely since BtubA/B protofilaments are known to be only 4 nm in diameter [17], and would therefore have to be closer together to interact. Slight helical twists in the tubes in vitro may have caused the appearance of twisted pairs [17]. While the number of protofilaments in eukaryotic microtubules can vary, the lateral interactions between them are conserved [28] such that each protofilament is shifted 0.93 nm along the tube axis relative to its neighbors. In 13-protofilament microtubules, this shift results in a three-start helix around the microtubule and a seam where a - and b -subunits interact [29]. Because the loops that are involved in these interactions are also present in BtubA and BtubB [17], we expect BtubA/B protofilaments to be shifted similarly. The sum of five such shifts (4.65 nm) is similar to the subunit repeat distance measured in BtubA/B tubes (4.2 and 4.4 nm, respectively) and suggests that BtubA/B form one-start helical tubes (Figure 4). The difference could be accommodated by a slightly different lateral interaction (a stagger of 0.84–0.88 nm instead of 0.93 nm). In support of this model, the major features of Fourier transforms of BtubA/B tube images matched those of a one-start five-protofilament helix model (Figures 5 and S10), but did not clarify whether BtubA/B tubes have an ‘‘A-lattice’’ without seam or a ‘‘B-lattice’’ with seam [30]. The latter seems more likely, however, since the B-lattice has been resolved in eukaryotic 13-protofilament microtubules, and is therefore depict- ed in Figure 4. Based on our data, the BtubA/B crystal structure [17], and the known structural features of the eukaryotic microtubule, we conclude therefore that BtubA/B heterodimers form five-protofilament, one-start helical tubes in vivo with lateral and longitudinal interactions like their eukaryotic counterparts. Since BtubA/B are true homologs of eukaryotic tubulin [11,12,17] and they form closely related structures differing mainly in the number of protofilaments, we suggest they be referred to as ‘‘bacterial microtubules’’ (bMTs). It has been suggested that BtubA and BtubB evolved from modern eukaryotic a - and/or b -tubulins [11,17,19,20]. If this were true, a phylogenetic association linking BtubA and BtubB to a - and/or b -tubulin would be expected. As shown ...
Context 2
... [11,12,24]. To begin, we verified that BtubA and BtubB proteins are in fact expressed in the species where the genes are present (Figures S1 and S2). Western hybridization and PCR also confirmed the absence of BtubA and BtubB in P. fluviatilis ( Figure S2) [24]. Next, Prosthecobacter cells were grown under different conditions and plunge-frozen across EM grids. A total of 589 cells were then imaged in 3-D by ECT. The spindle-shaped cells were polymorphic and exhibited prosthecae (cellular stalks) of different lengths. As seen in other bacterial phyla [25], multiple classes of cytoskeletal structures were seen, but one class had a tube-like morphology and was frequently found in the harboring species, but never in the btubAB -lacking strain (Figure 1). The abundance of these tube-like structures was dependent on the species imaged as well as the growth conditions and growth stage, and was found to be highest in P. vanneervenii cells grown directly on EM grids (67% of cells imaged). In sum, the tube-like structures were found in 48 of 176 P. vanneervenii , 9 of 111 P. dejongeii , 15 of 151 P. debontii , and 0 of 151 P. fluviatilis cells. The tube-like structures were 200– 1,200 nm long, always parallel to the cytoplasmic membrane, almost always localized in the stalk or in the transition zone between stalk and cell body, and occurred either individually or in bundles of two, three, or four (Figure 1, Figure S3, Movie S1). Chemical fixatives were found to degrade the structures (Figure S4), explaining why they were likely missed in previous conventional EM studies [11,22]. Since genetic tools are not yet available for prosthecobacters, we applied labeling and heterologous expression approaches to test whether the candidate structures were in fact composed of BtubA/ B as expected by their correlation with the presence of the genes. Recombinant Escherichia coli cells co-expressing BtubA and BtubB were imaged by ECT and exhibited strikingly similar tube-like structures running the length of the cells (Figure 2A) with the same localization as had been reported for BtubA/B from immuno- fluorescence [19]. Tube-like structures were not seen in control E. coli cells not expressing ButbA/B. Nearly identical tube-like structures were also seen when recombinant BtubA/B was polymerized in vitro and imaged by ECT (Figure 2B). The diameters and subunit repeat distances of all three structures (in Prosthecobacter , recombinant E. coli , and in vitro) were similar (7.6, 7.7, and 7.6 nm diameters, and 4.4, 4.4, and 4.2 nm repeat distances, respectively) (Figures 1, 2, and S3). Finally, immuno- gold-staining using anti-BtubB antibodies localized the proteins to the same region of Prosthecobacter cells as the candidate structures seen by ECT (Figures S5 and S6). We conclude therefore that the tube-like structures are composed of BtubA/B, and the slight differences in repeat distance, straightness, and bundling in the three samples were due to differences in protein concentrations and/or the absence of other interacting proteins in vitro and in E. coli . We have described the BtubA/B structures so far as ‘‘tube-like’’ because when acquiring a cryo-tomographic tilt-series, images of samples tilted beyond , 65 u cannot generally be included, so there is a missing ‘‘wedge’’ of data in reciprocal space that reduces the resolution in the direction of the electron beam. As a result, the ‘‘top’’ and ‘‘bottom’’ boundaries of cylindrical objects (considering the electron beam to be ‘‘vertical’’) are smeared, leaving the sidewalls to appear like two arcs facing each other (Figure 3A–D). Because the opposing arcs observed here were always in this orientation (facing each other and the beam path), it was clear that the structures must have been complete tubes distorted by the missing wedge rather than, for instance, parallel protofilaments, which would not be expected to always orient themselves in the same direction with respect to the electron beam. Nevertheless different orientations of tubes with respect to the tilt axis aggravate the missing wedge artifact differently [26,27], so to explore this effect tomograms of a known, tubular input structure consisting of BtubA/B crystal structures (see below) were simulated at different angles with respect to the tilt axis. These simulations recapitulated the experimental results well, since the density patterns (Figure 3H) were highly similar to those seen in experimental tomograms. To further confirm that the BtubA/B structures were in fact complete tubes and to obtain clearer cross-sectional views, btubAB harboring Prosthecobacter cells, recombinant E. coli cells, and purified BtubA/B polymerized in vitro were all high-pressure-frozen, cryosectioned, and imaged (Figure 3E–G). Cryosections through BtubA/B tubes appeared pentagonal, suggesting five-protofilament tubes. Using the heterodimeric BtubA/B crystal structure [17], we produced tube models with four, five, and six protofilaments for comparison. To maintain reasonable lateral interactions in such small tubes, protofilaments had to be spaced slightly closer (4.6 nm) than protofilaments in eukaryotic microtubules (5 nm), and this resulted in tube diameters of 6.7, 7.8, and 9.2 nm, respectively, for four-, five-, and six-protofilament tubes. Thus only the five-protofilament model was consistent with the 7.6-nm diameter measured in the tomograms, and the five- protofilament model fit the density of the BtubA/B tubes compellingly well (Figure 3I). Cross-sectional views of BtubA/B tubes in cryo-tomograms of whole cells and sub-tomogram averages often showed a left-right asymmetry (arrowheads in Figure 3A–C). Such an asymmetry can only arise from an uneven number of protofilaments, as demonstrated by simulated tomograms (Figure S7), further suggesting five rather than four or six protofilaments. Because the left-right asymmetries in computa- tional projections and in sub-tomographic averages at different positions along the tube axis remained consistent, the five protofilaments must be straight rather than twisting around the tube (Figure S8). Previous EM images of negatively stained, recombinant BtubA/ B polymerized in vitro were not described as tubes, but as protofilament bundles or twisted pairs [17,19,21]. We obtained similar-looking images staining our own purified BtubA/B (Figure S9), but having observed clear tubes in vivo and noting the frequent pairing of parallel densities , 7.6 nm apart in both our negatively stained images and the previously published images, we believe all these samples contained five-protofilament tubes as well. The alternative (two protofilaments 7.6 nm apart) seems unlikely since BtubA/B protofilaments are known to be only 4 nm in diameter [17], and would therefore have to be closer together to interact. Slight helical twists in the tubes in vitro may have caused the appearance of twisted pairs [17]. While the number of protofilaments in eukaryotic microtubules can vary, the lateral interactions between them are conserved [28] such that each protofilament is shifted 0.93 nm along the tube axis relative to its neighbors. In 13-protofilament microtubules, this shift results in a three-start helix around the microtubule and a seam where a - and b -subunits interact [29]. Because the loops that are involved in these interactions are also present in BtubA and BtubB [17], we expect BtubA/B protofilaments to be shifted similarly. The sum of five such shifts (4.65 nm) is similar to the subunit repeat distance measured in BtubA/B tubes (4.2 and 4.4 nm, respectively) and suggests that BtubA/B form one-start helical tubes (Figure 4). The difference could be accommodated by a slightly different lateral interaction (a stagger of 0.84–0.88 nm instead of 0.93 nm). In support of this model, the major features of Fourier transforms of BtubA/B tube images matched those of a one-start five-protofilament helix model (Figures 5 and S10), but did not clarify whether BtubA/B tubes have an ‘‘A-lattice’’ without seam or a ‘‘B-lattice’’ with seam [30]. The latter seems more likely, however, since the B-lattice has been resolved in eukaryotic 13-protofilament microtubules, and is therefore depict- ed in Figure 4. Based on our data, the BtubA/B crystal structure [17], and the known structural features of the eukaryotic microtubule, we conclude therefore that BtubA/B heterodimers form five-protofilament, one-start helical tubes in vivo with lateral and longitudinal interactions like their eukaryotic counterparts. Since BtubA/B are true homologs of eukaryotic tubulin [11,12,17] and they form closely related structures differing mainly in the number of protofilaments, we suggest they be referred to as ‘‘bacterial microtubules’’ (bMTs). It has been suggested that BtubA and BtubB evolved from modern eukaryotic a - and/or b -tubulins [11,17,19,20]. If this were true, a phylogenetic association linking BtubA and BtubB to a - and/or b -tubulin would be expected. As shown ...
Citations
... In addition, prokaryotic tubulin homologs can form various extended structures; for example, bacterial tubulins A and B (BtubA and BtubB) from Prosthecobacter dejoneii, which are closely related to eukaryotic α-and β-tubulins, assemble as a heterodimer [74]. BtubA/B tubular pentagonal structures were found in Prosthecobacter vanneervenii; these structures are 200 nm to 1200 nm long and present individually or in bundles of up to four [75]. The four- Our work supports the idea that the appearance of the permanent tubulin cytoskeleton as a system of microtubules occurred much later during evolution than the appearance of all the necessary components in the cells, not only did the last common ancestor of all eukaryotes already have microtubules and microtubule motors [14]. ...
... In addition, prokaryotic tubulin homologs can form various extended structures; for example, bacterial tubulins A and B (BtubA and BtubB) from Prosthecobacter dejoneii, which are closely related to eukaryotic αand β-tubulins, assemble as a heterodimer [74]. BtubA/B tubular pentagonal structures were found in Prosthecobacter vanneervenii; these structures are 200 nm to 1200 nm long and present individually or in bundles of up to four [75]. The four-stranded and two-stranded filament structures were described in Bacillus thuringiensis and are assembled from another tubulin homolog, TubZ [76]. ...
Microtubules are an indispensable component of all eukaryotic cells due to their role in mitotic spindle formation, yet their organization and number can vary greatly in the interphase. The last common ancestor of all eukaryotes already had microtubules and microtubule motor proteins moving along them. Sponges are traditionally regarded as the oldest animal phylum. Their body does not have a clear differentiation into tissues, but it contains several distinguishable cell types. The choanocytes stand out among them and are responsible for creating a flow of water with their flagella and increasing the filtering and feeding efficiency of the sponge. Choanocyte flagella contain microtubules, but thus far, observing a developed system of cytoplasmic microtubules in non-flagellated interphase sponge cells has been mostly unsuccessful. In this work, we combine transcriptomic analysis, immunofluorescence, and electron microscopy with time-lapse recording to demonstrate that microtubules appear in the cytoplasm of sponge cells only when transdifferentiation processes are activated. We conclude that dynamic cytoplasmic microtubules in the cells of sponges are not a persistent but rather a transient structure, associated with cellular plasticity.
... However, there are indications that microtubules are also crucial for quantum-mechanical processing of information [4,5]. This could be another reason why starting with bacteria [6], they are carefully preserved during the evolution of living organisms. ...
... However, there are indications that microtubules are also crucial for quantum-mechanical processing of information [4,5]. This could be another reason why starting with bacteria [6], they are carefully preserved during the evolution of living organisms. ...
... Interestingly, tyrosinase is not the only example of a Veruccomicrobium protein with traits of eukaryotic origin. These bacteria do, for instance, contain tubulin, which is found in all eukaryotes as a component of the cytoskeleton, but rarely in bacteria, and in contrast to other bacterial tubulins, it has been argued against being the result of lateral gene transfer from a eukaryote due to its distinct biochemical characteristics [70]. Thus, the tyrosinase only adds to the mysteries surrounding the evolution of the PVC superphylum, composed of the bacterial phyla Planctomycetes, Verrucomicrobia, Chlamydiae (PVC) and other species of related ancestry but very diverse pheno-and genotypes [71]. ...
Tyrosinases belong to the type-III copper enzyme family, which is involved in melanin production in a wide range of organisms. Despite similar overall characteristics and functions, their structures, activities, substrate specificities and regulation vary. The tyrosinase from the bacterium Verrucomicrobium spinosum (vsTyr) is produced as a pre-pro-enzyme in which a C-terminal extension serves as an inactivation domain. It does not require a caddie protein for copper ion incorporation, which makes it similar to eukaryotic tyrosinases. To gain an understanding of the catalytic machinery and regulation of vsTyr activity, we determined the structure of the catalytically active “core domain” of vsTyr by X-ray crystallography. The analysis showed that vsTyr is an atypical bacterial tyrosinase not only because it is independent of a caddie protein but also because it shows the highest structural (and sequence) similarity to plant-derived members of the type-III copper enzyme family and is more closely related to fungal tyrosinases regarding active site features. By modelling the structure of the pre-pro-enzyme using AlphaFold, we observed that Phe453, located in the C-terminal extension, is appropriately positioned to function as a “gatekeeper” residue. Our findings raise questions concerning the evolutionary origin of vsTyr.
... F. johnsoniae strain CJ2618 was grown as described (4). P. vanneervenii cells were grown as described (33). A. asiaticus strain 452471 was grown as described (34). ...
The bacterial flagellar type III secretion system (fT3SS) is a suite of membrane-embedded and cytoplasmic proteins responsible for building the flagellar motility machinery. Homologous nonflagellar (NF-T3SS) proteins form the injectisome machinery that bacteria use to deliver effector proteins into eukaryotic cells, and other family members were recently reported to be involved in the formation of membrane nanotubes. Here, we describe a novel, evolutionarily widespread, hat-shaped structure embedded in the inner membranes of bacteria, of yet-unidentified function, that is present in species containing fT3SS. Mutant analysis suggests a relationship between this novel structure and the fT3SS, but not the NF-T3SS. While the function of this novel structure remains unknown, we hypothesize that either some of the fT3SS proteins assemble within the hat-like structure, perhaps including the fT3SS core complex, or that fT3SS components regulate other proteins that form part of this novel structure.
IMPORTANCE The type III secretion system (T3SS) is a fascinating suite of proteins involved in building diverse macromolecular systems, including the bacterial flagellar motility machine, the injectisome machinery that bacteria use to inject effector proteins into host cells, and probably membrane nanotubes which connect bacterial cells. Here, we accidentally discovered a novel inner membrane-associated complex related to the flagellar T3SS. Examining our lab database, which is comprised of more than 40,000 cryo-tomograms of dozens of species, we discovered that this novel structure is both ubiquitous and ancient, being present in highly divergent classes of bacteria. Discovering a novel, widespread structure related to what are among the best-studied molecular machines in bacteria will open new venues for research aiming at understanding the function and evolution of T3SS proteins.
... Plant microtubules, unlike their animal counterparts, appear to be nucleated in a decentralized manner at the nuclear envelope and possibly in the cell cortex (Kost, Bao, & Chua, 2002;Lloyd, 1991). All tubulins, motor proteins, chaperones, and MAPs were known to evolve from a common bacterial ancestor (Ludueñ a, 2013;Pilhofer, Ladinsky, McDowall, Petroni, & Jensen, 2011). Further studies suggest that tubulin superfamily with α, β, γ, δ, ε, ζ, and η as the members, which are highly conserved in eukaryotes (McKean, Vaughan, & Gull, 2001). ...
... Actin-chromobody, a cameloid nanobody directed against actin that has been genetically encoded and fused to TagGFP2 nanobodies (Melak, Plessner, & Grosse, 2017). Actin-chromobodies have been successfully used in plant and animal systems (Panza, Maier, Schmees, Rothbauer, & Sollner, 2015;Plessner, Melak, Chinchilla, Baarlink, & Grosse, 2015;Rocchetti, Hawes, & Kriechbaumer, 2014). Though Actin-chromobody also poses problems of a high background similar to Life-Act, Actin-chromobody does not cause changes in actin dynamics. ...
The cytoskeleton is the network of polymeric structures which is a highly dynamic component of the plant cells. Cytoskeletal networks have been indicated for almost every intracellular activity, from cell division to cell movement, cell morphogenesis, apoptosis, and cell signaling. In plants, this filamentous network is comprised of microtubules and actin filaments. Cytoskeletal elements are also involved in many signaling pathways, including abiotic stress responses. Deciphering the role of the cytoskeleton requires efficient omics technologies, advanced imaging methods, and molecular genetics. In this chapter, we discuss the structure of actin filaments and microtubules, the available methodologies to study these structures and finally the perception and cytoskeletal response to abiotic stresses such as salinity, drought, or temperature.
... Elegant studies in artificially fused unicellular microorganisms demonstrated transfer of learned behavior from one cell to the other, suggesting that rudimentary memory may be stored in the cytoskeletal proteins (Vogel and Dussutour, 2016). In addition, information transfer was detected after fusing two bacteria of different species, indicating that microtubules and tubulin, recently identified in microbes, could participate in this process (Pilhofer et al., 2011;Charubin et al., 2020). Interestingly, human tissues, such as the skeletal muscle, fascia and blood cells may process and store information, further implicating microtubules and tubulin in cognition (Moore and Cao, 2008;Tozzi, 2014;Snijders et al., 2020). ...
A growing body of epidemiological and research data has associated neurotropic viruses with accelerated brain aging and increased risk of neurodegenerative disorders. Many viruses replicate optimally in senescent cells, as they offer a hospitable microenvironment with persistently elevated cytosolic calcium, abundant intracellular iron, and low interferon type I. As cell-cell fusion is a major driver of cellular senescence, many viruses have developed the ability to promote this phenotype by forming syncytia. Cell-cell fusion is associated with immunosuppression mediated by phosphatidylserine externalization that enable viruses to evade host defenses. In hosts, virus-induced immune dysfunction and premature cellular senescence may predispose to neurodegenerative disorders. This concept is supported by novel studies that found postinfectious cognitive dysfunction in several viral illnesses, including human immunodeficiency virus-1, herpes simplex virus-1, and SARS-CoV-2. Virus-induced pathological syncytia may provide a unified framework for conceptualizing neuronal cell cycle reentry, aneuploidy, somatic mosaicism, viral spreading of pathological Tau and elimination of viable synapses and neurons by neurotoxic astrocytes and microglia. In this narrative review, we take a closer look at cell-cell fusion and vesicular merger in the pathogenesis of neurodegenerative disorders. We present a “decentralized” information processing model that conceptualizes neurodegeneration as a systemic illness, triggered by cytoskeletal pathology. We also discuss strategies for reversing cell-cell fusion, including, TMEM16F inhibitors, calcium channel blockers, senolytics, and tubulin stabilizing agents. Finally, going beyond neurodegeneration, we examine the potential benefit of harnessing fusion as a therapeutic strategy in regenerative medicine.
... 16,17 Called bacterial tubulin A and B (BtubA/B), these proteins interact to form microtubulelike structures in the presence of guanosine triphosphate (GTP). 18 Bacterial microtubules (bMTs) consist of five 19 or four protofilaments, 20 as reported in in vivo and in vitro studies, respectively. Bacterial microtubules are thus noticeably thinner than eukaryotic microtubules, which consist of 13 protofilaments. ...
... 22 Another distinguishing feature of Prosthecobacter is the presence of narrowed extensions of the cell wall, called prosthecae. 19 Bacterial microtubules seem to be predominately located in these cell stalks, which suggests that they might be involved in their formation. It has also been proposed that BtubA/B filaments may contribute to intracellular organization. ...
... Unlike eukaryotic tubulin, BtubA/B is not dependent on protein chaperones and post-translational modifications, and it can be functionally expressed in Escherichia coli. 19 Because the cytoplasm of Prosthecobacter probably resembles that of E. coli, 18 CFPS platforms derived from E. coli represent a physiologically relevant environment to investigate bMTs. Not only does CFPS allow one to bypass protein purification but it also enables the continuous interrogation of the protein dynamic behavior in the course of its production. ...
Genetic control over a cytoskeletal network inside lipid vesicles offers a potential route to controlled shape changes and DNA segregation in synthetic cell biology. Bacterial microtubules (bMTs) are protein filaments found in bacteria of the genus Prosthecobacter. They are formed by the tubulins BtubA and BtubB, which polymerize in the presence of GTP. Here, we show that the tubulins BtubA/B can be functionally expressed from DNA templates in a reconstituted transcription-translation system, thus providing a cytosol-like environment to study their biochemical and biophysical properties. We found that bMTs spontaneously interact with lipid membranes and display treadmilling. When compartmentalized inside liposomes, de novo synthesized BtubA/B tubulins self-organize into cytoskeletal structures of different morphologies. Moreover, bMTs can exert a pushing force on the membrane and deform liposomes, a phenomenon that can be reversed by a light-activated disassembly of the filaments. Our work establishes bMTs as a new building block in synthetic biology. In the context of creating a synthetic cell, bMTs could help shape the lipid compartment, establish polarity or directional transport, and assist the division machinery.
... Les microtubules forment un réseau très conservé chez les eucaryotes (levure, champignons, plantes et animaux) ( Figure 2). Des homologues des tubulines eucaryotes, impliqués dans la division cellulaire, sont également observés chez les procaryotes tels que FTZ retrouvé chez la plupart des bactéries et archées ainsi que BtubA et BtubB plus récemment identifiés chez Prosthecobacter (Pilhofer et al., 2011) (Figure 2G). Selon les espèces, les microtubules adoptent une grande variété d'organisation allant d'une microfibre formée de 4 protofilaments chez la bactérie Prosthecobacter à un complexe axonémal regroupant 40 protofilaments au niveau du spermatozoïde de l'insecte Mantispa perla ( Figure 2F et 2G) (Dallai et al., 2005;Chaaban and Brouhard, 2017). ...
Les microtubules sont des composants essentiels du cytosquelette, dont la dynamique est intiment liée à l’hydrolyse du GTP de la tubuline (sous-unité β) lors de la polymérisation. La paroi des MTs, constituée de tubuline-GDP et métastable, serait stabilisée par l’extrémité en croissance constituée de tubuline-GTP et ainsi nommée coiffe GTP. Cependant, la nature de la coiffe ainsi que les modifications conformationnelles subies par la tubuline lors de l’hydrolyse du GTP restent discutées. Le modèle actuel propose que la tubuline-GTP de la coiffe possèderait une conformation étendue et que son hydrolyse serait accompagnée d’une compaction fragilisant les contacts latéraux inter-tubuline et donc la stabilité de la paroi. Cependant, ce modèle découle d’études structurales réalisées sur des parois de MTs formées à partir de tubuline-GMPCPP, un analogue lentement hydrolysable du GTP, et suppose que cet analogue constituerait le modèle bona fide du GTP. Afin de vérifier cette théorie, nous avons entrepris au cours de ma thèse l’analyse structurale de parois assemblées en présence d’une gamme élargie d’analogues lentement hydrolysables du GTP. Nos résultats montrent notamment qu’un autre analogue du GTP, le GDP-BeF3-, pourrait s’avérer être un meilleur analogue structural que le GMPCPP et suggèrent l’absence de compaction de la tubuline lors de l’hydrolyse du GTP. La compaction de la tubuline ne serait donc pas le mécanisme responsable de l’instabilité de la paroi des MTs.
... The cytoskeleton comprises microtubules (MTs), which are dynamic cytoplasmic tubular polymers, microfilaments, and intermediate filaments [137]. MTs are critical for the innate and adaptive immune responses and the dynamics of inflammatory cells [138][139][140][141]. Colchicine is an orally administered potent anti-inflammatory drug. ...
Coronavirus disease 2019 (COVID-19), which is a respiratory illness associated with high mortality, has been classified as a pandemic. The major obstacles for the clinicians to contain the disease are limited information availability, difficulty in disease diagnosis, predicting disease prognosis, and lack of disease monitoring tools. Additionally, the lack of valid therapies has further contributed to the difficulties in containing the pandemic. Recent studies have reported that the dysregulation of the immune system leads to an ineffective antiviral response and promotes pathological immune response, which manifests as ARDS, myocarditis, and hepatitis. In this study, a novel platform has been described for disseminating information to physicians for the diagnosis and monitoring of patients with COVID-19. An adjuvant approach using compounds that can potentiate antiviral immune response and mitigate COVID-19-induced immune-mediated target organ damage has been presented. A prolonged beneficial effect is achieved by implementing algorithm-based individualized variability measures in the treatment regimen.
