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Cheryl Y. Hayashi - One of the best experts on this subject based on the ideXlab platform.
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Spidroins and silk fibers of aquatic spiders
Scientific Reports, 2019Co-Authors: Sandra M Correagarhwal, Thomas H. Clarke, Cheryl Y. Hayashi, Marc Janssen, Luc Crevecoeur, Bryce N Mcquillan, Angela Simpson, Cor J VinkAbstract:: Spiders are commonly found in terrestrial environments and many rely heavily on their silks for fitness related tasks such as reproduction and dispersal. Although rare, a few species occupy aquatic or semi-aquatic habitats and for them, silk-related specializations are also essential to survive in aquatic environments. Most spider silks studied to date are from cob-web and orb-web weaving species, leaving the silks from many other terrestrial spiders as well as water-associated spiders largely undescribed. Here, we characterize silks from three Dictynoidea species: the aquatic spiders Argyroneta aquatica and Desis marina as well as the terrestrial Badumna longinqua. From silk gland RNA-Seq libraries, we report a total of 47 different homologs of the spidroin (spider fibroin) gene family. Some of these 47 Spidroins correspond to known spidroin types (aciniform, ampullate, cribellar, pyriform, and tubuliform), while other Spidroins represent novel branches of the spidroin gene family. We also report a hydrophobic amino acid motif (GV) that, to date, is found only in the Spidroins of aquatic and semi-aquatic spiders. Comparison of spider silk sequences to the silks from other water-associated arthropods, shows that there is a diversity of strategies to function in aquatic environments.
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Silk genes and silk gene expression in the spider Tengella perfuga (Zoropsidae), including a potential cribellar spidroin (CrSp).
PLOS ONE, 2018Co-Authors: Sandra M. Correa-garhwal, Thomas H. Clarke, R. Crystal Chaw, Liliana G. Alaniz, Fanny S. Chan, Rachael E. Alfaro, Cheryl Y. HayashiAbstract:Most spiders spin multiple types of silk, including silks for reproduction, prey capture, and draglines. Spiders are a megadiverse group and the majority of spider silks remain uncharacterized. For example, nothing is known about the silk molecules of Tengella perfuga, a spider that spins sheet webs lined with cribellar silk. Cribellar silk is a type of adhesive capture thread composed of numerous fibrils that originate from a specialized plate-like spinning organ called the cribellum. The predominant components of spider silks are Spidroins, members of a protein family synthesized in silk glands. Here, we use silk gland RNA-Seq and cDNA libraries to infer T. perfuga silks at the protein level. We show that T. perfuga spiders express 13 silk transcripts representing at least five categories of spider silk proteins (Spidroins). One category is a candidate for cribellar silk and is thus named cribellar spidroin (CrSp). Studies of ontogenetic changes in web construction and spigot morphology in T. perfuga have documented that after sexual maturation, T. perfuga females continue to make capture webs but males halt web maintenance and cease spinning cribellar silk. Consistent with these observations, our candidate CrSp was expressed only in females. The other four spidroin categories correspond to paralogs of aciniform, ampullate, pyriform, and tubuliform Spidroins. These Spidroins are associated with egg sac and web construction. Except for the tubuliform spidroin, the Spidroins from T. perfuga contain novel combinations of amino acid sequence motifs that have not been observed before in these spidroin types. Characterization of T. perfuga silk genes, particularly CrSp, expand the diversity of the spidroin family and inspire new structure/function hypotheses.
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Genomic perspectives of spider silk genes through target capture sequencing: Conservation of stabilization mechanisms and homology-based structural models of spidroin terminal regions.
International Journal of Biological Macromolecules, 2018Co-Authors: Matthew A. Collin, Nadia A Ayoub, Thomas H. Clarke, Cheryl Y. HayashiAbstract:Abstract A powerful system for studying protein aggregation, particularly rapid self-assembly, is spider silk. Spider silks are proteinaceous and silk proteins are synthesized and stored within silk glands as liquid dope. As needed, liquid dope is near-instantaneously transformed into solid fibers or viscous adhesives. The dominant constituents of silks are Spidroins (spider fibroins) and their terminal domains are vital for the tight control of silk self-assembly. To better understand spidroin termini, we used target capture and deep sequencing to identify spidroin gene sequences from six species representing the araneoid families of Araneidae, Nephilidae, and Theridiidae. We obtained 145 terminal regions, of which 103 are newly annotated here, as well as novel variants within nine diverse spidroin types. Our comparative analyses demonstrated the conservation of acidic, basic, and cysteine amino acid residues across spidroin types that had been proposed to be important for monomer stability, dimer formation, and self-assembly from a limited sampling of Spidroins. Computational, protein homology modeling revealed areas of spidroin terminal regions that are highly conserved in three-dimensions despite sequence divergence across spidroin types. Analyses of our dense sampling of terminal regions suggest that most Spidroins share stabilization mechanisms, dimer formation, and tertiary structure, despite producing functionally distinct materials.
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The Nephila clavipes genome highlights the diversity of spider silk genes and their complex expression
Nature Genetics, 2017Co-Authors: Paul L Babb, Sandra M. Correa-garhwal, Cheryl Y. Hayashi, Nicholas F Lahens, David N Nicholson, John B Hogenesch, Matjaž Kuntner, Linden Higgins, Ingi Agnarsson, Benjamin F VoightAbstract:Spider silks are the toughest known biological materials, yet are lightweight and virtually invisible to the human immune system, and they thus have revolutionary potential for medicine and industry. Spider silks are largely composed of Spidroins, a unique family of structural proteins. To investigate spidroin genes systematically, we constructed the first genome of an orb-weaving spider: the golden orb-weaver ( Nephila clavipes ), which builds large webs using an extensive repertoire of silks with diverse physical properties. We cataloged 28 Nephila Spidroins, representing all known orb-weaver spidroin types, and identified 394 repeated coding motif variants and higher-order repetitive cassette structures unique to specific Spidroins. Characterization of spidroin expression in distinct silk gland types indicates that glands can express multiple spidroin types. We find evidence of an alternatively spliced spidroin, a spidroin expressed only in venom glands, evolutionary mechanisms for spidroin diversification, and non-spidroin genes with expression patterns that suggest roles in silk production. Benjamin Voight and colleagues report the annotated genome of the golden orb-weaver spider. They describe 28 spider silk genes (Spidroins), characterize their expression in distinct silk gland types and identify non-spidroin genes with expression patterns suggesting potential roles in silk production.
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the nephila clavipes genome highlights the diversity of spider silk genes and their complex expression
Nature Genetics, 2017Co-Authors: Paul L Babb, Cheryl Y. Hayashi, Nicholas F Lahens, David N Nicholson, John B Hogenesch, Matjaž Kuntner, Linden Higgins, Ingi Agnarsson, Sandra M Correagarhwal, Benjamin F VoightAbstract:Benjamin Voight and colleagues report the annotated genome of the golden orb-weaver spider. They describe 28 spider silk genes (Spidroins), characterize their expression in distinct silk gland types and identify non-spidroin genes with expression patterns suggesting potential roles in silk production.
Anna Rising - One of the best experts on this subject based on the ideXlab platform.
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production and properties of triple chimeric Spidroins
Biomacromolecules, 2018Co-Authors: Yizhong Zhou, Anna Rising, Jan Johansson, Qing MengAbstract:All spider silk proteins (Spidroins) are composed of N- and C-terminal domains (NT and CT) that act as regulators of silk solubility and assembly and a central repetitive region, which confers mechanical properties to the fiber. Among the seven types of spider silks, aciniform silk has the highest toughness. Herein, we fused NT and CT domains from major and minor ampullate Spidroins (MaSps and MiSps), respectively, to 1–4 repeat domains (W) from another type of spidroin, aciniform spidroin 1(AcSp1). Although the three domains originate from distantly related spidroin types, they keep their respective characteristics in the chimeric Spidroins. Furthermore, all chimeric Spidroins could form silk-like fibers by manual-drawing. In contrast to fibers made in the same manner from W domains only, NTW1–4CT fibers show superior mechanical properties. Our results suggest that chimeric Spidroins with NT, CT, and repeat domains can be designed to form fibers with various mechanical properties.
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diversified structural basis of a conserved molecular mechanism for ph dependent dimerization in spider silk n terminal domains
ChemBioChem, 2015Co-Authors: Mārtiņs Otikovs, Qing Meng, Michael Landreh, Kerstin Nordling, Gefei Chen, Hans Jornvall, Nina Kronqvist, Anna RisingAbstract:: Conversion of spider silk proteins from soluble dope to insoluble fibers involves pH-dependent dimerization of the N-terminal domain (NT). This conversion is tightly regulated to prevent premature precipitation and enable rapid silk formation at the end of the duct. Three glutamic acid residues that mediate this process in the NT from Euprosthenops australis major ampullate spidroin 1 are well conserved among Spidroins. However, NTs of minor ampullate Spidroins from several species, including Araneus ventricosus ((Av)MiSp NT), lack one of the glutamic acids. Here we investigate the pH-dependent structural changes of (Av)MiSp NT, revealing that it uses the same mechanism but involves a non-conserved glutamic acid residue instead. Homology modeling of the structures of other MiSp NTs suggests that these harbor different compensatory residues. This indicates that, despite sequence variations, the molecular mechanism underlying pH-dependent dimerization of NT is conserved among different silk types.
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Toward spinning artificial spider silk
Nature Chemical Biology, 2015Co-Authors: Anna Rising, Jan JohanssonAbstract:Spider silks have enormous potential as strong yet flexible biomaterials, but obtaining artificial silk polymers has proven challenging. Recent advances in our understanding of natural silk processing may inform techniques for silk production. Spider silk is strong and extensible but still biodegradable and well tolerated when implanted, making it the ultimate biomaterial. Shortcomings that arise in replicating spider silk are due to the use of recombinant spider silk proteins (Spidroins) that lack native domains, the use of denaturing conditions under purification and spinning and the fact that the understanding of how spiders control silk formation is incomplete. Recent progress has unraveled the molecular mechanisms of the spidroin N- and C-terminal nonrepetitive domains (NTs and CTs) and revealed the pH and ion gradients in spiders' silk glands, clarifying how spidroin solubility is maintained and how silk is formed in a fraction of a second. Protons and CO_2, generated by carbonic anhydrase, affect the stability and structures of the NT and CT in different ways. These insights should allow the design of conditions and devices for the spinning of recombinant Spidroins into native-like silk.
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morphology and composition of the spider major ampullate gland and dragline silk
Biomacromolecules, 2013Co-Authors: Marlene Andersson, Jan Johansson, Lena Holm, Yvonne Ridderstrale, Anna RisingAbstract:Spider silk is made of unique proteins—Spidroins—secreted and stored as a protein solution (dope) in specialized glands. The major ampullate gland, source of the dragline silk, is composed of a tail, a sac and an elongated duct. For this gland, several different types of epithelial cells and granules have been described, but it is largely unknown how they correlate with spidroin production. It is also not settled what parts of the large Spidroins end up in the final silk, and it has been suggested that the N-terminal domain (NT) is lacking. Here we show that NT is present in the dope and throughout dragline silk fibers, including the skin layer, and that the major ampullate tail and sac consist of three different and sharply demarcated zones (A–C), each with a distinct epithelial cell type. Finally, we show that Spidroins are produced in the A and B zone epithelia, while the C zone granules lack Spidroins.
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Full-length minor ampullate spidroin gene sequence.
PLOS ONE, 2012Co-Authors: Gefei Chen, Anna Rising, Jan Johansson, Yunlong Zhang, Zijiang Yang, Qing MengAbstract:Spider silk includes seven protein based fibers and glue-like substances produced by glands in the spider's abdomen. Minor ampullate silk is used to make the auxiliary spiral of the orb-web and also for wrapping prey, has a high tensile strength and does not supercontract in water. So far, only partial cDNA sequences have been obtained for minor ampullate Spidroins (MiSps). Here we describe the first MiSp full-length gene sequence from the spider species Araneus ventricosus, using a multidimensional PCR approach. Comparative analysis of the sequence reveals regulatory elements, as well as unique spidroin gene and protein architecture including the presence of an unusually large intron. The spliced full-length transcript of MiSp gene is 5440 bp in size and encodes 1766 amino acid residues organized into conserved nonrepetitive N- and C-terminal domains and a central predominantly repetitive region composed of four units that are iterated in a non regular manner. The repeats are more conserved within A. ventricosus MiSp than compared to repeats from homologous proteins, and are interrupted by two nonrepetitive spacer regions, which have 100% identity even at the nucleotide level.
Jan Johansson - One of the best experts on this subject based on the ideXlab platform.
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production and properties of triple chimeric Spidroins
Biomacromolecules, 2018Co-Authors: Yizhong Zhou, Anna Rising, Jan Johansson, Qing MengAbstract:All spider silk proteins (Spidroins) are composed of N- and C-terminal domains (NT and CT) that act as regulators of silk solubility and assembly and a central repetitive region, which confers mechanical properties to the fiber. Among the seven types of spider silks, aciniform silk has the highest toughness. Herein, we fused NT and CT domains from major and minor ampullate Spidroins (MaSps and MiSps), respectively, to 1–4 repeat domains (W) from another type of spidroin, aciniform spidroin 1(AcSp1). Although the three domains originate from distantly related spidroin types, they keep their respective characteristics in the chimeric Spidroins. Furthermore, all chimeric Spidroins could form silk-like fibers by manual-drawing. In contrast to fibers made in the same manner from W domains only, NTW1–4CT fibers show superior mechanical properties. Our results suggest that chimeric Spidroins with NT, CT, and repeat domains can be designed to form fibers with various mechanical properties.
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Toward spinning artificial spider silk
Nature Chemical Biology, 2015Co-Authors: Anna Rising, Jan JohanssonAbstract:Spider silks have enormous potential as strong yet flexible biomaterials, but obtaining artificial silk polymers has proven challenging. Recent advances in our understanding of natural silk processing may inform techniques for silk production. Spider silk is strong and extensible but still biodegradable and well tolerated when implanted, making it the ultimate biomaterial. Shortcomings that arise in replicating spider silk are due to the use of recombinant spider silk proteins (Spidroins) that lack native domains, the use of denaturing conditions under purification and spinning and the fact that the understanding of how spiders control silk formation is incomplete. Recent progress has unraveled the molecular mechanisms of the spidroin N- and C-terminal nonrepetitive domains (NTs and CTs) and revealed the pH and ion gradients in spiders' silk glands, clarifying how spidroin solubility is maintained and how silk is formed in a fraction of a second. Protons and CO_2, generated by carbonic anhydrase, affect the stability and structures of the NT and CT in different ways. These insights should allow the design of conditions and devices for the spinning of recombinant Spidroins into native-like silk.
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morphology and composition of the spider major ampullate gland and dragline silk
Biomacromolecules, 2013Co-Authors: Marlene Andersson, Jan Johansson, Lena Holm, Yvonne Ridderstrale, Anna RisingAbstract:Spider silk is made of unique proteins—Spidroins—secreted and stored as a protein solution (dope) in specialized glands. The major ampullate gland, source of the dragline silk, is composed of a tail, a sac and an elongated duct. For this gland, several different types of epithelial cells and granules have been described, but it is largely unknown how they correlate with spidroin production. It is also not settled what parts of the large Spidroins end up in the final silk, and it has been suggested that the N-terminal domain (NT) is lacking. Here we show that NT is present in the dope and throughout dragline silk fibers, including the skin layer, and that the major ampullate tail and sac consist of three different and sharply demarcated zones (A–C), each with a distinct epithelial cell type. Finally, we show that Spidroins are produced in the A and B zone epithelia, while the C zone granules lack Spidroins.
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Full-length minor ampullate spidroin gene sequence.
PLOS ONE, 2012Co-Authors: Gefei Chen, Anna Rising, Jan Johansson, Yunlong Zhang, Zijiang Yang, Qing MengAbstract:Spider silk includes seven protein based fibers and glue-like substances produced by glands in the spider's abdomen. Minor ampullate silk is used to make the auxiliary spiral of the orb-web and also for wrapping prey, has a high tensile strength and does not supercontract in water. So far, only partial cDNA sequences have been obtained for minor ampullate Spidroins (MiSps). Here we describe the first MiSp full-length gene sequence from the spider species Araneus ventricosus, using a multidimensional PCR approach. Comparative analysis of the sequence reveals regulatory elements, as well as unique spidroin gene and protein architecture including the presence of an unusually large intron. The spliced full-length transcript of MiSp gene is 5440 bp in size and encodes 1766 amino acid residues organized into conserved nonrepetitive N- and C-terminal domains and a central predominantly repetitive region composed of four units that are iterated in a non regular manner. The repeats are more conserved within A. ventricosus MiSp than compared to repeats from homologous proteins, and are interrupted by two nonrepetitive spacer regions, which have 100% identity even at the nucleotide level.
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ph dependent dimerization of spider silk n terminal domain requires relocation of a wedged tryptophan side chain
Journal of Molecular Biology, 2012Co-Authors: Kristaps Jaudzems, My Hedhammar, Anna Rising, Michael Landreh, Kerstin Nordling, Stefan D Knight, Hans Jornvall, Gelareh Askarieh, Jan JohanssonAbstract:Formation of spider silk from its constituent proteins—Spidroins—involves changes from soluble helical/coil conformations to insoluble β-sheet aggregates. This conversion needs to be regulated to avoid precocious aggregation proximally in the silk gland while still allowing rapid silk assembly in the distal parts. Lowering of pH from about 7 to 6 is apparently important for silk formation. The spidroin N-terminal domain (NT) undergoes stable dimerization and structural changes in this pH region, but the underlying mechanisms are incompletely understood. Here, we determine the NMR and crystal structures of Euprosthenops australis NT mutated in the dimer interface (A72R). Also, the NMR structure of wild‐type (wt) E. australis NT at pH 7.2 and 300 mM sodium chloride was determined. The wt NT and A72R structures are monomers and virtually identical, but they differ from the subunit structure of dimeric wt NT mainly by having a tryptophan (W10) buried between helix 1 and helix 3, while W10 is surface exposed in the dimer. Wedging of the W10 side chain in monomeric NT tilts helix 3 approximately 5–6 A into a position that is incompatible with that of the observed dimer structure. The structural differences between monomeric and dimeric NT domains explain the tryptophan fluorescence patterns of NT at pH 7 and pH 6 and indicate that the biological function of NT depends on conversion between the two conformations.
Nadia A Ayoub - One of the best experts on this subject based on the ideXlab platform.
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Genomic perspectives of spider silk genes through target capture sequencing: Conservation of stabilization mechanisms and homology-based structural models of spidroin terminal regions.
International Journal of Biological Macromolecules, 2018Co-Authors: Matthew A. Collin, Nadia A Ayoub, Thomas H. Clarke, Cheryl Y. HayashiAbstract:Abstract A powerful system for studying protein aggregation, particularly rapid self-assembly, is spider silk. Spider silks are proteinaceous and silk proteins are synthesized and stored within silk glands as liquid dope. As needed, liquid dope is near-instantaneously transformed into solid fibers or viscous adhesives. The dominant constituents of silks are Spidroins (spider fibroins) and their terminal domains are vital for the tight control of silk self-assembly. To better understand spidroin termini, we used target capture and deep sequencing to identify spidroin gene sequences from six species representing the araneoid families of Araneidae, Nephilidae, and Theridiidae. We obtained 145 terminal regions, of which 103 are newly annotated here, as well as novel variants within nine diverse spidroin types. Our comparative analyses demonstrated the conservation of acidic, basic, and cysteine amino acid residues across spidroin types that had been proposed to be important for monomer stability, dimer formation, and self-assembly from a limited sampling of Spidroins. Computational, protein homology modeling revealed areas of spidroin terminal regions that are highly conserved in three-dimensions despite sequence divergence across spidroin types. Analyses of our dense sampling of terminal regions suggest that most Spidroins share stabilization mechanisms, dimer formation, and tertiary structure, despite producing functionally distinct materials.
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Silk gene expression of theridiid spiders: implications for male-specific silk use
Zoology, 2017Co-Authors: Sandra M. Correa-garhwal, Nadia A Ayoub, Thomas H. Clarke, R. Crystal Chaw, Cheryl Y. HayashiAbstract:Abstract Spiders (order Araneae) rely on their silks for essential tasks, such as dispersal, prey capture, and reproduction. Spider silks are largely composed of Spidroins, members of a protein family that are synthesized in silk glands. As needed, silk stored in silk glands is extruded through spigots on the spinnerets. Nearly all studies of spider silks have been conducted on females; thus, little is known about male silk biology. To shed light on silk use by males, we compared silk gene expression profiles of mature males to those of females from three cob-web weaving species (Theridiidae). We de novo assembled species-specific male transcriptomes from Latrodectus hesperus , Latrodectus geometricus , and Steatoda grossa followed by differential gene expression analyses. Consistent with their complement of silk spigots, male theridiid spiders express appreciable amounts of aciniform, major ampullate, minor ampullate, and pyriform spidroin genes but not tubuliform spidroin genes. The relative expression levels of particular spidroin genes varied between sexes and species. Because mature males desert their prey-capture webs and become cursorial in their search for mates, we anticipated that major ampullate (dragline) spidroin genes would be the silk genes most highly expressed by males. Indeed, major ampullate spidroin genes had the highest expression in S. grossa males. However, minor ampullate spidroin genes were the most highly expressed spidroin genes in L. geometricus and L. hesperus males. Our expression profiling results suggest species-specific adaptive divergence of silk use by male theridiids.
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Duplication and concerted evolution of MiSp-encoding genes underlie the material properties of minor ampullate silks of cobweb weaving spiders
BMC Evolutionary Biology, 2017Co-Authors: Jannelle M. Vienneau-hathaway, Amanda Kelly Lane, Thomas H. Clarke, Jessica E. Garb, Sandra M. Correa-garhwal, Matthew A. Collin, Cheryl Y. Hayashi, Elizabeth R. Brassfield, Evelyn E. Schwager, Nadia A AyoubAbstract:Background Orb-web weaving spiders and their relatives use multiple types of task-specific silks. The majority of spider silk studies have focused on the ultra-tough dragline silk synthesized in major ampullate glands, but other silk types have impressive material properties. For instance, minor ampullate silks of orb-web weaving spiders are as tough as draglines, due to their higher extensibility despite lower strength. Differences in material properties between silk types result from differences in their component proteins, particularly members of the spidroin (spider fibroin) gene family. However, the extent to which variation in material properties within a single silk type can be explained by variation in spidroin sequences is unknown. Here, we compare the minor ampullate Spidroins (MiSp) of orb-weavers and cobweb weavers. Orb-web weavers use minor ampullate silk to form the auxiliary spiral of the orb-web while cobweb weavers use it to wrap prey, suggesting that selection pressures on minor ampullate Spidroins (MiSp) may differ between the two groups. Results We report complete or nearly complete MiSp sequences from five cobweb weaving spider species and measure material properties of minor ampullate silks in a subset of these species. We also compare MiSp sequences and silk properties of our cobweb weavers to published data for orb-web weavers. We demonstrate that all our cobweb weavers possess multiple MiSp loci and that one locus is more highly expressed in at least two species. We also find that the proportion of β-spiral-forming amino acid motifs in MiSp positively correlates with minor ampullate silk extensibility across orb-web and cobweb weavers. Conclusions MiSp sequences vary dramatically within and among spider species, and have likely been subject to multiple rounds of gene duplication and concerted evolution, which have contributed to the diverse material properties of minor ampullate silks. Our sequences also provide templates for recombinant silk proteins with tailored properties.
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Evidence from Multiple Species that Spider Silk Glue Component ASG2 is a Spidroin
Scientific Reports, 2016Co-Authors: Matthew A. Collin, Nadia A Ayoub, Thomas H. Clarke, Cheryl Y. HayashiAbstract:Spiders in the superfamily Araneoidea produce viscous glue from aggregate silk glands. Aggregate glue coats prey-capture threads and hampers the escape of prey from webs, thereby increasing the foraging success of spiders. cDNAs for Aggregate Spider Glue 1 (ASG1) and 2 (ASG2) have been previously described from the golden orb-weaver, Nephila clavipes and Western black widow, Latrodectus hesperus . To further investigate aggregate glues, we assembled ASG1 and ASG2 from genomic target capture libraries constructed from three species of cob-web weavers and three species of orb-web weavers, all araneoids. We show that ASG1 is unlikely to be a glue, but rather is part of a widespread arthropod gene family, the peritrophic matrix proteins. For ASG2, we demonstrate its remarkable architectural and sequence similarities to spider silk fibroins, indicating that ASG2 is a member of the spidroin gene family. Thus, Spidroins have diversified into glues in addition to task-specific, high performance fibers.
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Proteomic Evidence for Components of Spider Silk Synthesis from Black Widow Silk Glands and Fibers.
Journal of Proteome Research, 2015Co-Authors: R. Crystal Chaw, Nadia A Ayoub, Thomas H. Clarke, Sandra M. Correa-garhwal, Cheryl Y. HayashiAbstract:Spider silk research has largely focused on Spidroins, proteins that are the primary components of spider silk fibers. Although a number of Spidroins have been characterized, other types of proteins associated with silk synthesis are virtually unknown. Previous analyses of tissue-specific RNA-seq libraries identified 647 predicted genes that were differentially expressed in silk glands of the Western black widow, Latrodectus hesperus. Only ∼5% of these silk-gland specific transcripts (SSTs) encode Spidroins; although the remaining predicted genes presumably encode other proteins associated with silk production, this is mostly unverified. Here, we used proteomic analysis of multiple silk glands and dragline silk fiber to investigate the translation of the differentially expressed genes. We find 48 proteins encoded by the differentially expressed transcripts in L. hesperus major ampullate, minor ampullate, and tubuliform silk glands and detect 17 SST encoded proteins in major ampullate silk fibers. The obse...
Jessica E. Garb - One of the best experts on this subject based on the ideXlab platform.
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Duplication and concerted evolution of MiSp-encoding genes underlie the material properties of minor ampullate silks of cobweb weaving spiders
BMC Evolutionary Biology, 2017Co-Authors: Jannelle M. Vienneau-hathaway, Amanda Kelly Lane, Thomas H. Clarke, Jessica E. Garb, Sandra M. Correa-garhwal, Matthew A. Collin, Cheryl Y. Hayashi, Elizabeth R. Brassfield, Evelyn E. Schwager, Nadia A AyoubAbstract:Background Orb-web weaving spiders and their relatives use multiple types of task-specific silks. The majority of spider silk studies have focused on the ultra-tough dragline silk synthesized in major ampullate glands, but other silk types have impressive material properties. For instance, minor ampullate silks of orb-web weaving spiders are as tough as draglines, due to their higher extensibility despite lower strength. Differences in material properties between silk types result from differences in their component proteins, particularly members of the spidroin (spider fibroin) gene family. However, the extent to which variation in material properties within a single silk type can be explained by variation in spidroin sequences is unknown. Here, we compare the minor ampullate Spidroins (MiSp) of orb-weavers and cobweb weavers. Orb-web weavers use minor ampullate silk to form the auxiliary spiral of the orb-web while cobweb weavers use it to wrap prey, suggesting that selection pressures on minor ampullate Spidroins (MiSp) may differ between the two groups. Results We report complete or nearly complete MiSp sequences from five cobweb weaving spider species and measure material properties of minor ampullate silks in a subset of these species. We also compare MiSp sequences and silk properties of our cobweb weavers to published data for orb-web weavers. We demonstrate that all our cobweb weavers possess multiple MiSp loci and that one locus is more highly expressed in at least two species. We also find that the proportion of β-spiral-forming amino acid motifs in MiSp positively correlates with minor ampullate silk extensibility across orb-web and cobweb weavers. Conclusions MiSp sequences vary dramatically within and among spider species, and have likely been subject to multiple rounds of gene duplication and concerted evolution, which have contributed to the diverse material properties of minor ampullate silks. Our sequences also provide templates for recombinant silk proteins with tailored properties.
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Ancient Properties of Spider Silks Revealed by the Complete Gene Sequence of the Prey-Wrapping Silk Protein (AcSp1)
Molecular Biology and Evolution, 2012Co-Authors: Nadia A Ayoub, Jessica E. Garb, Amanda Kuelbs, Cheryl Y. HayashiAbstract:Spider silk fibers have impressive mechanical properties and are primarily composed of highly repetitive structural proteins (termed Spidroins) encoded by a single gene family. Most characterized spidroin genes are incompletely known because of their extreme size (typically >9 kb) and repetitiveness, limiting understanding of the evolutionary processes that gave rise to their unusual gene architectures. The only complete spidroin genes characterized thus far form the dragline in the Western black widow, Latrodectus hesperus. Here, we describe the first complete gene sequence encoding the aciniform spidroin AcSp1, the primary component of spider prey-wrapping fibers. L. hesperus AcSp1 contains a single enormous (∼19 kb) exon. The AcSp1 repeat sequence is exceptionally conserved between two widow species (∼94% identity) and between widows and distantly related orb-weavers (∼30% identity), consistent with a history of strong purifying selection on its amino acid sequence. Furthermore, the 16 repeats (each 371-375 amino acids long) found in black widow AcSp1 are, on average, >99% identical at the nucleotide level. A combination of stabilizing selection on amino acid sequence, selection on silent sites, and intragenic recombination likely explains the extreme homogenization of AcSp1 repeats. In addition, phylogenetic analyses of spidroin paralogs support a gene duplication event occurring concomitantly with specialization of the aciniform glands and the tubuliform glands, which synthesize egg-case silk. With repeats that are dramatically different in length and amino acid composition from dragline Spidroins, our L. hesperus AcSp1 expands the knowledge base for developing silk-based biomimetic technologies.
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Early events in the evolution of spider silk genes.
PLOS ONE, 2012Co-Authors: James Starrett, Jessica E. Garb, Amanda Kuelbs, Ugochi O. Azubuike, Cheryl Y. HayashiAbstract:Silk spinning is essential to spider ecology and has had a key role in the expansive diversification of spiders. Silk is composed primarily of proteins called Spidroins, which are encoded by a multi-gene family. Spidroins have been studied extensively in the derived clade, Orbiculariae (orb-weavers), from the suborder Araneomorphae (‘true spiders’). Orbicularians produce a suite of different silks, and underlying this repertoire is a history of duplication and spidroin gene divergence. A second class of silk proteins, Egg Case Proteins (ECPs), is known only from the orbicularian species, Lactrodectus hesperus (Western black widow). In L. hesperus, ECPs bond with tubuliform Spidroins to form egg case silk fibers. Because most of the phylogenetic diversity of spiders has not been sampled for their silk genes, there is limited understanding of spidroin gene family history and the prevalence of ECPs. Silk genes have not been reported from the suborder Mesothelae (segmented spiders), which diverged from all other spiders >380 million years ago, and sampling from Mygalomorphae (tarantulas, trapdoor spiders) and basal araneomorph lineages is sparse. In comparison to orbicularians, mesotheles and mygalomorphs have a simpler silk biology and thus are hypothesized to have less diversity of silk genes. Here, we present cDNAs synthesized from the silk glands of six mygalomorph species, a mesothele, and a non-orbicularian araneomorph, and uncover a surprisingly rich silk gene diversity. In particular, we find ECP homologs in the mesothele, suggesting that ECPs were present in the common ancestor of extant spiders, and originally were not specialized to complex with tubuliform Spidroins. Furthermore, gene-tree/species-tree reconciliation analysis reveals that numerous spidroin gene duplications occurred after the split between Mesothelae and Opisthothelae (Mygalomorphae plus Araneomorphae). We use the spidroin gene tree to reconstruct the evolution of amino acid compositions of Spidroins that perform different ecological functions.
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untangling spider silk evolution with spidroin terminal domains
BMC Evolutionary Biology, 2010Co-Authors: Nadia A Ayoub, Jessica E. Garb, Cheryl Y. HayashiAbstract:Background Spidroins are a unique family of large, structural proteins that make up the bulk of spider silk fibers. Due to the highly variable nature of their repetitive sequences, spidroin evolutionary relationships have principally been determined from their non-repetitive carboxy (C)-terminal domains, though they offer limited character data. The few known spidroin amino (N)-terminal domains have been difficult to obtain, but potentially contain critical phylogenetic information for reconstructing the diversification of spider silks. Here we used silk gland expression data (ESTs) from highly divergent species to evaluate the functional significance and phylogenetic utility of spidroin N-terminal domains.
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Untangling spider silk evolution with spidroin terminal domains
BMC Evolutionary Biology, 2010Co-Authors: Jessica E. Garb, Nadia A Ayoub, Cheryl Y. HayashiAbstract:Background Spidroins are a unique family of large, structural proteins that make up the bulk of spider silk fibers. Due to the highly variable nature of their repetitive sequences, spidroin evolutionary relationships have principally been determined from their non-repetitive carboxy (C)-terminal domains, though they offer limited character data. The few known spidroin amino (N)-terminal domains have been difficult to obtain, but potentially contain critical phylogenetic information for reconstructing the diversification of spider silks. Here we used silk gland expression data (ESTs) from highly divergent species to evaluate the functional significance and phylogenetic utility of spidroin N-terminal domains. Results We report 11 additional spidroin N-termini found by sequencing ~1,900 silk gland cDNAs from nine spider species that shared a common ancestor > 240 million years ago. In contrast to their hyper-variable repetitive regions, spidroin N-terminal domains have retained striking similarities in sequence identity, predicted secondary structure, and hydrophobicity. Through separate and combined phylogenetic analyses of N-terminal domains and their corresponding C-termini, we find that combined analysis produces the most resolved trees and that N-termini contribute more support and less conflict than the C-termini. These analyses show that paralogs largely group by silk gland type, except for the major ampullate Spidroins. Moreover, spidroin structural motifs associated with superior tensile strength arose early in the history of this gene family, whereas a motif conferring greater extensibility convergently evolved in two distantly related paralogs. Conclusions A non-repetitive N-terminal domain appears to be a universal attribute of spidroin proteins, likely retained from the origin of spider silk production. Since this time, spidroin N-termini have maintained several features, consistent with this domain playing a key role in silk assembly. Phylogenetic analyses of the conserved N- and C-terminal domains illustrate dramatic radiation of the spidroin gene family, involving extensive duplications, shifts in expression patterns and extreme diversification of repetitive structural sequences that endow spider silks with an unparalleled range of mechanical properties.
