KONO Lab
Bioinformatics, Systems biology, Synthetic biology
Graduate School of Media and Governance, Keio University
Faculty of Environment and Information Studies, Keio University
Material x Omics
Darwin's bark spider shares a spidroin repertoire with Caerostris extrusa but achieves extraordinary silk toughness through gene expression
Spider silk is a protein-based material whose toughness suggests possible novel applications. A particularly fascinating example of silk toughness is provided by Darwin's bark spider (Caerostris darwini) found in Madagascar. This spider produces extraordinarily tough silk, with an average toughness of 350 MJ m−1 and over 50% extensibility, and can build river-bridging webs with a size of 2.8 m2. Recent studies have suggested that specific spidroins expressed in C. darwini are responsible for the mechanical properties of its silk. Therefore, a more comprehensive investigation of spidroin sequences, silk thread protein contents and phylogenetic conservation among closely related species is required. Here, we conducted genomic, transcriptomic and proteomic analyses of C. darwini and its close relative Caerostris extrusa. A variety of spidroins and low-molecular-weight proteins were found in the dragline silk of these species; all of the genes encoding these proteins were conserved in both genomes, but their genes were more expressed in C. darwini. The potential to produce very tough silk is common in the genus Caerostris, and our results may suggest the existence of plasticity allowing silk mechanical properties to be changed by optimizing related gene expression in response to the environment
Multicomponent nature underlies the extraordinary mechanical properties of spider dragline silk
Significance: Artificial synthesis of spider silk has been actively pursued. However, until now, the natural mechanical properties of spider silk have been largely unreproducible. We thoroughly investigated the genomes and transcripts of four related species of orb-weaver spiders as well as the proteins in their silk threads. Then, in addition to spidroin, we found several low-molecular-weight proteins in common. Interestingly, the low-molecular-weight protein component of spider dragline silk doubled the tensile strength of artificial silk–based material. This discovery will greatly advance the industry and research on the use of protein-based materials.
Dragline silk of golden orb-weaver spiders (Nephilinae) is noted for its unsurpassed toughness, combining extraordinary extensibility and tensile strength, suggesting industrial application as a sustainable biopolymer material. To pinpoint the molecular composition of dragline silk and the roles of its constituents in achieving its mechanical properties, we report a multiomics approach, combining high-quality genome sequencing and assembly, silk gland transcriptomics, and dragline silk proteomics of four Nephilinae spiders. We observed the consistent presence of the MaSp3B spidroin unique to this subfamily as well as several nonspidroin SpiCE proteins. Artificial synthesis and the combination of these components in vitro showed that the multicomponent nature of dragline silk, including MaSp3B and SpiCE, along with MaSp1 and MaSp2, is essential to realize the mechanical properties of spider dragline silk.
The balance of crystalline and amorphous regions in the fibroin structure underpins the tensile strength of bagworm silk
Protein-based materials are considered versatile biomaterials, and their biodegradability is an advantage for sustainable development. Bagworm produces strong silk for use in unique situations throughout its life stages. Rigorous molecular analyses of Eumeta variegata suggested that the particular mechanical properties of its silk are due to the coexistence of poly-A and GA motifs. However, little molecular information on closely related species is available, and it is not understood how these properties were acquired evolutionarily or whether the motif combination is a conserved trait in other bagworms. Here, we performed a transcriptome analysis of two other bagworm species (Canephora pungelerii and Bambalina sp.) belonging to the family Psychidae to elucidate the relationship between the fibroin gene and silk properties. The obtained transcriptome assemblies and tensile tests indicated that the motif combination and silk properties were conserved among the bagworms. Furthermore, our analysis showed that C. pungelerii produces extraordinarily strong silk (breaking strength of 1.4 GPa) and indicated that the cause may be the C. pungelerii-specific balance of crystalline/amorphous regions in the H-fibroin repetitive domain. This particular H-fibroin architecture may have been evolutionarily acquired to produce strong thread to maintain bag stability during the relatively long development period of Canephora species relative to other bagworms.
Spider silk self-assembly via modular liquid-liquid phase separation and nanofibrillation
Spider silk fiber rapidly assembles from spidroin protein in soluble state via an incompletely understood mechanism. Here, we present an integrated model for silk formation that incorporates the effects of multiple chemical and physical gradients on the different spidroin functional domains. Central to the process is liquid-liquid phase separation (LLPS) that occurs in response to multivalent anions such as phosphate, mediated by the carboxyl-terminal and repetitive domains. Acidification coupled with LLPS triggers the swift self-assembly of nanofibril networks, facilitated by dimerization of the amino-terminal domain, and leads to a liquid-to-solid phase transition. Mechanical stress applied to the fibril structures yields macroscopic fibers with hierarchical organization and enriched for β-sheet conformations. Studies using native silk gland material corroborate our findings on spidroin phase separation. Our results suggest an intriguing parallel between silk assembly and other LLPS-mediated mechanisms, such as found in intracellular membraneless organelles and protein aggregation disorders.
Spidroin profiling of cribellate spiders provides insight into the evolution of spider prey capture strategies
Orb-weaving spiders have two main methods of prey capture: cribellate spiders use dry, sticky capture threads, and ecribellate spiders use viscid glue droplets. Predation behaviour is a major evolutionary driving force, and it is important on spider phylogeny whether the cribellate and ecribellate spiders each evolved the orb architecture independently or both strategies were derived from an ancient orb web. These hypotheses have been discussed based on behavioural and morphological characteristics, with little discussion on this subject from the perspective of molecular materials of orb web, since there is little information about cribellate spider-associated spidroin genes. Here, we present in detail a spidroin catalogue of six uloborid species of cribellate orb-weaving spiders, including cribellate and pseudoflagelliform spidroins, with transcriptome assembly complemented with long read sequencing, where silk composition is confirmed by proteomics. Comparative analysis across families (Araneidae and Uloboridae) shows that the gene architecture, repetitive domains, and amino acid frequencies of the orb web constituting silk proteins are similar among orb-weaving spiders regardless of the prey capture strategy. Notably, the fact that there is a difference only in the prey capture thread proteins strongly supports the monophyletic origin of the orb web.
Orb-weaving spider Araneus ventricosus genome elucidates the spidroin gene catalogue
Members of the family Araneidae are common orb-weaving spiders, and they produce several types of silks throughout their behaviors and lives, from reproduction to foraging. Egg sac, prey capture thread, or dragline silk possesses characteristic mechanical properties, and its variability makes it a highly attractive material for ecological, evolutional, and industrial fields. However, the complete set of constituents of silks produced by a single species is still unclear, and novel spidroin genes as well as other proteins are still being found. Here, we present the first genome in genus Araneus together with the full set of spidroin genes with unamplified long reads and confirmed with transcriptome of the silk glands and proteome analysis of the dragline silk. The catalogue includes the first full length sequence of a paralog of major ampullate spidroin MaSp3, and several spider silk-constituting elements designated SpiCE. Family-wide phylogenomic analysis of Araneidae suggests the relatively recent acquisition of these genes, and multiple-omics analyses demonstrate that these proteins are critical components in the abdominal spidroin gland and dragline silk, contributing to the outstanding mechanical properties of silk in this group of species.
The bagworm genome reveals a unique fibroin gene that provides high tensile strength
Arthropod silk is known as a versatile tool, and its variability makes it an attractive biomaterial. Eumeta variegata is a bagworm moth (Lepidoptera, Psychidae) that uses silk throughout all life stages. Notably, the bagworm-specific uses of silk include larval development in a bag coated with silk and plant materials and the use of silk attachments to hang pupae. An understanding at the molecular level of bagworm silk, which enables such unique purposes, is an opportunity to expand the possibilities for artificial biomaterial design. However, very little is known about the bagworm fibroin gene and the mechanical properties of bagworm silk. Here, we report the bagworm genome, including a silk fibroin gene. The genome is approximately 700 Mbp in size, and the newly found fibroin gene has a unique repetitive motif. Furthermore, a mechanical property test demonstrates a phylogenetic relationship between the unique motif and tensile strength of bagworm silk.
1000 spider silkomes: Linking sequences to silk physical properties
Spider silks are among the toughest known materials and thus provide models for renewable, biodegradable, and sustainable biopolymers. However, the entirety of their diversity still remains elusive, and silks that exceed the performance limits of industrial fibers are constantly being found. We obtained transcriptome assemblies from 1098 species of spiders to comprehensively catalog silk gene sequences and measured the mechanical, thermal, structural, and hydration properties of the dragline silks of 446 species. The combination of these silk protein genotype-phenotype data revealed essential contributions of multicomponent structures with major ampullate spidroin 1 to 3 paralogs in high-performance dragline silks and numerous amino acid motifs contributing to each of the measured properties. We hope that our global sampling, comprehensive testing, integrated analysis, and open data will provide a solid starting point for future biomaterial designs.
