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Marcos Sotomayor - One of the best experts on this subject based on the ideXlab platform.
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Structural determinants of Protocadherin-15 mechanics and function in hearing and balance perception.
Proceedings of the National Academy of Sciences of the United States of America, 2020Co-Authors: Deepanshu Choudhary, Yoshie Narui, Brandon L. Neel, Lahiru N. Wimalasena, Carissa F. Klanseck, Pedro De-la-torre, Conghui Chen, Raul Araya-secchi, Elakkiya Tamilselvan, Marcos SotomayorAbstract:The vertebrate inner ear, responsible for hearing and balance, is able to sense minute mechanical stimuli originating from an extraordinarily broad range of sound frequencies and intensities or from head movements. Integral to these processes is the tip-link protein complex, which conveys force to open the inner-ear transduction channels that mediate sensory perception. Protocadherin-15 and cadherin-23, two atypically large cadherins with 11 and 27 extracellular cadherin (EC) repeats, are involved in deafness and balance disorders and assemble as parallel homodimers that interact to form the tip link. Here we report the X-ray crystal structure of a Protocadherin-15 + cadherin-23 heterotetrameric complex at 2.9-A resolution, depicting a parallel homodimer of Protocadherin-15 EC1-3 molecules forming an antiparallel complex with two cadherin-23 EC1-2 molecules. In addition, we report structures for 10 Protocadherin-15 fragments used to build complete high-resolution models of the monomeric Protocadherin-15 ectodomain. Molecular dynamics simulations and validated crystal contacts are used to propose models for the complete extracellular Protocadherin-15 parallel homodimer and the tip-link bond. Steered molecular dynamics simulations of these models suggest conditions in which a structurally diverse and multimodal Protocadherin-15 ectodomain can act as a stiff or soft gating spring. These results reveal the structural determinants of tip-link-mediated inner-ear sensory perception and elucidate Protocadherin-15's structural and adhesive properties relevant in disease.
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identification of an adhesive interface for the non clustered δ1 Protocadherin 1 involved in respiratory diseases
Communications biology, 2019Co-Authors: Debadrita Modak, Marcos SotomayorAbstract:: Cadherins form a large family of calcium-dependent adhesive proteins involved in morphogenesis, cell differentiation, and neuronal connectivity. Non-clustered δ1 Protocadherins form a cadherin subgroup of proteins with seven extracellular cadherin (EC) repeats and cytoplasmic domains distinct from those of classical cadherins. Non-clustered δ1 Protocadherins mediate homophilic adhesion and have been implicated in various diseases including asthma, autism, and cancer. Here we present X-ray crystal structures of human Protocadherin-1 (PCDH1), a δ1-Protocadherin member essential for New World Hantavirus infection that is typically expressed in the brain, airway epithelium, skin keratinocytes, and lungs. The structures suggest a binding mode that involves antiparallel overlap of repeats EC1 to EC4. Mutagenesis combined with binding assays and biochemical experiments validated this mode of adhesion. Overall, these results reveal the molecular mechanism underlying adhesiveness of PCDH1 and δ1-Protocadherins, also shedding light on PCDH1's role in maintaining airway epithelial integrity, the loss of which causes respiratory diseases.
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interaction specificity of clustered Protocadherins inferred from sequence covariation and structural analysis
Proceedings of the National Academy of Sciences of the United States of America, 2019Co-Authors: John M Nicoludis, Marcos Sotomayor, Anna G Green, Sanket Walujkar, Debora S Marks, Rachelle GaudetAbstract:Clustered Protocadherins, a large family of paralogous proteins that play important roles in neuronal development, provide an important case study of interaction specificity in a large eukaryotic protein family. A mammalian genome has more than 50 clustered Protocadherin isoforms, which have remarkable homophilic specificity for interactions between cellular surfaces. A large antiparallel dimer interface formed by the first 4 extracellular cadherin (EC) domains controls this interaction. To understand how specificity is achieved between the numerous paralogs, we used a combination of structural and computational approaches. Molecular dynamics simulations revealed that individual EC interactions are weak and undergo binding and unbinding events, but together they form a stable complex through polyvalency. Strongly evolutionarily coupled residue pairs interacted more frequently in our simulations, suggesting that sequence coevolution can inform the frequency of interaction and biochemical nature of a residue interaction. With these simulations and sequence coevolution, we generated a statistical model of interaction energy for the clustered Protocadherin family that measures the contributions of all amino acid pairs at the interface. Our interaction energy model assesses specificity for all possible pairs of isoforms, recapitulating known pairings and predicting the effects of experimental changes in isoform specificity that are consistent with literature results. Our results show that sequence coevolution can be used to understand specificity determinants in a protein family and prioritize interface amino acid substitutions to reprogram specific protein–protein interactions.
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an adhesive interface for the non clustered δ1 Protocadherin 1 involved in respiratory diseases
bioRxiv, 2018Co-Authors: Debadrita Modak, Marcos SotomayorAbstract:Cadherins form a large family of calcium-dependent adhesive proteins involved in morphogenesis, cell differentiation, and neuronal connectivity. Non-clustered {delta}1 Protocadherins form a cadherin subgroup of proteins with seven extracellular cadherin (EC) repeats and cytoplasmic domains distinct from those of classical cadherins. The non-clustered {delta}1 Protocadherins mediate homophilic adhesion and have been implicated in various diseases including asthma, autism, and cancer. Here we present X-ray crystal structures of Protocadherin-1 (PCDH1), a {delta}1-Protocadherin member essential for New World hantavirus infection that is typically expressed in the brain, airway epithelium, skin keratinocytes, and lungs. The structures suggest a binding mode that involves antiparallel overlap of repeats EC1 to EC4. Mutagenesis combined with binding assays and biochemical experiments validated this mode of adhesion. Overall, these results reveal the molecular mechanism underlying adhesiveness of PCDH1 and {delta}1-Protocadherins, also shedding light on PCDH1's role in maintaining airway epithelial integrity, the loss of which causes respiratory diseases.
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interaction specificity of clustered Protocadherins inferred from sequence covariation and structural analysis
bioRxiv, 2018Co-Authors: John M Nicoludis, Marcos Sotomayor, Anna G Green, Sanket Walujkar, Debora S Marks, Rachelle GaudetAbstract:Clustered Protocadherins are a large family of paralogous proteins that play important roles in neuronal development. The more than 50 clustered Protocadherin isoforms have remarkable homophilic specificity for interactions between cellular surfaces that is controlled by a large antiparallel dimer interface formed by the first four extracellular cadherin (EC) domains. To understand how specificity is achieved between the numerous paralogs, we used a combination of structural and computational approaches. Molecular dynamics simulations revealed that individual EC interactions are weak and go through binding and unbinding events, but together they form a stable complex through polyvalency. Using sequence coevolution, we generated a statistical model of interaction energy for the clustered Protocadherin family that measures the contributions of all amino acid pairs in the interface. Our interaction energy model assesses specificity for all possible pairs of isoforms, recapitulating known pairings and predicting the effects of experimental changes in isoform specificity that are consistent with literature results. Our results show that sequence coevolution can be used to understand specificity determinants in a protein family and prioritize interface amino acid substitutions to reprogram specific protein-protein interactions.
Yoshie Narui - One of the best experts on this subject based on the ideXlab platform.
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Structural determinants of Protocadherin-15 mechanics and function in hearing and balance perception.
Proceedings of the National Academy of Sciences of the United States of America, 2020Co-Authors: Deepanshu Choudhary, Yoshie Narui, Brandon L. Neel, Lahiru N. Wimalasena, Carissa F. Klanseck, Pedro De-la-torre, Conghui Chen, Raul Araya-secchi, Elakkiya Tamilselvan, Marcos SotomayorAbstract:The vertebrate inner ear, responsible for hearing and balance, is able to sense minute mechanical stimuli originating from an extraordinarily broad range of sound frequencies and intensities or from head movements. Integral to these processes is the tip-link protein complex, which conveys force to open the inner-ear transduction channels that mediate sensory perception. Protocadherin-15 and cadherin-23, two atypically large cadherins with 11 and 27 extracellular cadherin (EC) repeats, are involved in deafness and balance disorders and assemble as parallel homodimers that interact to form the tip link. Here we report the X-ray crystal structure of a Protocadherin-15 + cadherin-23 heterotetrameric complex at 2.9-A resolution, depicting a parallel homodimer of Protocadherin-15 EC1-3 molecules forming an antiparallel complex with two cadherin-23 EC1-2 molecules. In addition, we report structures for 10 Protocadherin-15 fragments used to build complete high-resolution models of the monomeric Protocadherin-15 ectodomain. Molecular dynamics simulations and validated crystal contacts are used to propose models for the complete extracellular Protocadherin-15 parallel homodimer and the tip-link bond. Steered molecular dynamics simulations of these models suggest conditions in which a structurally diverse and multimodal Protocadherin-15 ectodomain can act as a stiff or soft gating spring. These results reveal the structural determinants of tip-link-mediated inner-ear sensory perception and elucidate Protocadherin-15's structural and adhesive properties relevant in disease.
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A Mechanically Weak Extracellular Membrane-Adjacent Domain Induces Dimerization of Protocadherin-15.
Biophysical journal, 2018Co-Authors: Pedro De-la-torre, Deepanshu Choudhary, Raul Araya-secchi, Yoshie NaruiAbstract:Abstract The cadherin superfamily of proteins is defined by the presence of extracellular cadherin (EC) “repeats” that engage in protein-protein interactions to mediate cell-cell adhesion, cell signaling, and mechanotransduction. The extracellular domains of nonclassical cadherins often have a large number of EC repeats along with other subdomains of various folds. Protocadherin-15 (PCDH15), a protein component of the inner-ear tip link filament essential for mechanotransduction, has 11 EC repeats and a membrane adjacent domain (MAD12) of atypical fold. Here we report the crystal structure of a pig PCDH15 fragment including EC10, EC11, and MAD12 in a parallel dimeric arrangement. MAD12 has a unique molecular architecture and folds as a ferredoxin-like domain similar to that found in the nucleoporin protein Nup54. Analytical ultracentrifugation experiments along with size-exclusion chromatography coupled to multiangle laser light scattering and small-angle x-ray scattering corroborate the crystallographic dimer and show that MAD12 induces parallel dimerization of PCDH15 near its membrane insertion point. In addition, steered molecular dynamics simulations suggest that MAD12 is mechanically weak and may unfold before tip-link rupture. Sequence analyses and structural modeling predict the existence of similar domains in cadherin-23, Protocadherin-24, and the “giant” FAT and CELSR cadherins, indicating that some of them may also exhibit MAD-induced parallel dimerization.
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A Mechanically Weak Extracellular Membrane-Adjacent Domain Induces Dimerization of Protocadherin-15
2018Co-Authors: Pedro De-la-torre, Deepanshu Choudhary, Raul Araya-secchi, Yoshie NaruiAbstract:The cadherin superfamily of proteins is defined by the presence of extracellular cadherin (EC) repeats that engage in protein-protein interactions to mediate cell-cell adhesion, cell signaling, and mechanotransduction. The extracellular domains of non-classical cadherins often have a large number of EC repeats along with other subdomains of various folds. Protocadherin-15 (PCDH15), a protein component of the inner-ear tip link filament essential for mechanotransduction, has eleven EC repeats and a membrane adjacent domain (MAD12) of atypical fold. Here we report the crystal structure of a pig PCDH15 fragment including EC10, EC11, and MAD12 in a parallel dimeric arrangement. MAD12 has a unique molecular architecture and folds as a ferredoxin-like domain similar to that found in the nucleoporin protein Nup54. Analytical ultracentrifugation experiments along with size exclusion chromatography coupled to multi-angle laser light scattering and small-angle X-ray scattering corroborate the crystallographic dimer and show that MAD12 induces parallel dimerization of PCDH15 near its membrane insertion point. In addition, steered molecular dynamics simulations suggest that MAD12 is mechanically weak and may unfold before tip-link rupture. Sequence analyses and structural modeling predict the existence of similar domains in cadherin-23, Protocadherin-24, and the "giant" FAT and CELSR cadherins, indicating that some of them may also exhibit MAD-induced parallel dimerization.
Teruyoshi Hirayama - One of the best experts on this subject based on the ideXlab platform.
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the methyltransferase setdb1 regulates a large neuron specific topological chromatin domain
Nature Genetics, 2017Co-Authors: Yan Jiang, Prashanth Rajarajan, Teruyoshi Hirayama, Will Liao, Bibi S Kassim, Behnam Javidfar, Brigham J Hartley, Lisa Kleofas, Royce Park, Benoit LabonteAbstract:Schahram Akbarian and colleagues report that mutation of the gene encoding the SETDB1 (KMT1E) histone methyltransferase in mouse neurons leads to dissolution of chromosome conformations and a topologically associated domain at the clustered Protocadherin locus. They show that SETDB1 prevents excess CTCF binding and is important for maintaining developmentally important higher-order chromatin organization.
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the methyltransferase setdb1 regulates a large neuron specific topological chromatin domain
Nature Genetics, 2017Co-Authors: Yan Jiang, Prashanth Rajarajan, Teruyoshi Hirayama, Will Liao, Behnam Javidfar, Brigham J Hartley, Lisa Kleofas, Yonghwee E Loh, Bibi Kassim, Royce ParkAbstract:We report locus-specific disintegration of megabase-scale chromosomal conformations in brain after neuronal ablation of Setdb1 (also known as Kmt1e; encodes a histone H3 lysine 9 methyltransferase), including a large topologically associated 1.2-Mb domain conserved in humans and mice that encompasses >70 genes at the clustered Protocadherin locus (hereafter referred to as cPcdh). The cPcdh topologically associated domain (TADcPcdh) in neurons from mutant mice showed abnormal accumulation of the transcriptional regulator and three-dimensional (3D) genome organizer CTCF at cryptic binding sites, in conjunction with DNA cytosine hypomethylation, histone hyperacetylation and upregulated expression. Genes encoding stochastically expressed Protocadherins were transcribed by increased numbers of cortical neurons, indicating relaxation of single-cell constraint. SETDB1-dependent loop formations bypassed 0.2-1 Mb of linear genome and radiated from the TADcPcdh fringes toward cis-regulatory sequences within the cPcdh locus, counterbalanced shorter-range facilitative promoter-enhancer contacts and carried loop-bound polymorphisms that were associated with genetic risk for schizophrenia. We show that the SETDB1 repressor complex, which involves multiple KRAB zinc finger proteins, shields neuronal genomes from excess CTCF binding and is critically required for structural maintenance of TADcPcdh.
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Regulation of clustered Protocadherin genes in individual neurons.
Seminars in cell & developmental biology, 2017Co-Authors: Teruyoshi Hirayama, Takeshi YagiAbstract:Individual neurons are basic functional units in the complex system of the brain. One aspect of neuronal individuality is generated by stochastic and combinatorial expression of diverse clustered Protocadherins (Pcdhs), encoded by the Pcdha, Pcdhb, and Pcdhg gene clusters, that are critical for several aspects of neural circuit formation. Each clustered Pcdh gene has its own promoter containing conserved sequences and is transcribed by a promoter choice mechanism involving interaction between the promoter and enhancers. A CTCF/Cohesin complex induces this interaction by configuration of DNA-looping in the chromatin structure. At the same time, the semi-stochastic expression of clustered Pcdh genes is regulated in individual neurons by DNA methylation: the methyltransferase Dnmt3b regulates methylation state of individual clustered Pcdh genes during early embryonic stages prior to the establishment of neural stem cells. Several other factors, including Smchd1, also contribute to the regulation of clustered Pcdh gene expression. In addition, psychiatric diseases and early life experiences of individuals can influence expression of clustered Pcdh genes in the brain, through epigenetic alterations. Clustered Pcdh gene expression is thus a significant and highly regulated step in establishing neuronal individuality and generating functional neural circuits in the brain.
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identification of the cluster control region for the Protocadherin β genes located beyond the Protocadherin γ cluster
Journal of Biological Chemistry, 2011Co-Authors: Shinnichi Yokota, Teruyoshi Hirayama, Keizo Hirano, Ryosuke Kaneko, Shunsuke Toyoda, Yoshimi Kawamura, Masumi Hirabayashi, Takahiro Hirabayashi, Takeshi YagiAbstract:The clustered Protocadherins (Pcdhs), Pcdh-α, -β, and -γ, are transmembrane proteins constituting a subgroup of the cadherin superfamily. Each Pcdh cluster is arranged in tandem on the same chromosome. Each of the three Pcdh clusters shows stochastic and combinatorial expression in individual neurons, thus generating a hugely diverse set of possible cell surface molecules. Therefore, the clustered Pcdhs are candidates for determining neuronal molecular diversity. Here, we showed that the targeted deletion of DNase I hypersensitive (HS) site HS5-1, previously identified as a Pcdh-α regulatory element in vitro, affects especially the expression of specific Pcdh-α isoforms in vivo. We also identified a Pcdh-β cluster control region (CCR) containing six HS sites (HS16, 17, 17′, 18, 19, and 20) downstream of the Pcdh-γ cluster. This CCR comprehensively activates the expression of the Pcdh-β gene cluster in cis, and its deletion dramatically decreases their expression levels. Deleting the CCR nonuniformly down-regulates some Pcdh-γ isoforms and does not affect Pcdh-α expression. Thus, the CCR effect extends beyond the 320-kb region containing the Pcdh-γ cluster to activate the upstream Pcdh-β genes. Thus, we concluded that the CCR is a highly specific regulatory unit for Pcdh-β expression on the clustered Pcdh genomic locus. These findings suggest that each Pcdh cluster is controlled by distinct regulatory elements that activate their expression and that the stochastic gene regulation of the clustered Pcdhs is controlled by the complex chromatin architecture of the clustered Pcdh locus.
Takeshi Yagi - One of the best experts on this subject based on the ideXlab platform.
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Regulation of clustered Protocadherin genes in individual neurons.
Seminars in cell & developmental biology, 2017Co-Authors: Teruyoshi Hirayama, Takeshi YagiAbstract:Individual neurons are basic functional units in the complex system of the brain. One aspect of neuronal individuality is generated by stochastic and combinatorial expression of diverse clustered Protocadherins (Pcdhs), encoded by the Pcdha, Pcdhb, and Pcdhg gene clusters, that are critical for several aspects of neural circuit formation. Each clustered Pcdh gene has its own promoter containing conserved sequences and is transcribed by a promoter choice mechanism involving interaction between the promoter and enhancers. A CTCF/Cohesin complex induces this interaction by configuration of DNA-looping in the chromatin structure. At the same time, the semi-stochastic expression of clustered Pcdh genes is regulated in individual neurons by DNA methylation: the methyltransferase Dnmt3b regulates methylation state of individual clustered Pcdh genes during early embryonic stages prior to the establishment of neural stem cells. Several other factors, including Smchd1, also contribute to the regulation of clustered Pcdh gene expression. In addition, psychiatric diseases and early life experiences of individuals can influence expression of clustered Pcdh genes in the brain, through epigenetic alterations. Clustered Pcdh gene expression is thus a significant and highly regulated step in establishing neuronal individuality and generating functional neural circuits in the brain.
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identification of the cluster control region for the Protocadherin β genes located beyond the Protocadherin γ cluster
Journal of Biological Chemistry, 2011Co-Authors: Shinnichi Yokota, Teruyoshi Hirayama, Keizo Hirano, Ryosuke Kaneko, Shunsuke Toyoda, Yoshimi Kawamura, Masumi Hirabayashi, Takahiro Hirabayashi, Takeshi YagiAbstract:The clustered Protocadherins (Pcdhs), Pcdh-α, -β, and -γ, are transmembrane proteins constituting a subgroup of the cadherin superfamily. Each Pcdh cluster is arranged in tandem on the same chromosome. Each of the three Pcdh clusters shows stochastic and combinatorial expression in individual neurons, thus generating a hugely diverse set of possible cell surface molecules. Therefore, the clustered Pcdhs are candidates for determining neuronal molecular diversity. Here, we showed that the targeted deletion of DNase I hypersensitive (HS) site HS5-1, previously identified as a Pcdh-α regulatory element in vitro, affects especially the expression of specific Pcdh-α isoforms in vivo. We also identified a Pcdh-β cluster control region (CCR) containing six HS sites (HS16, 17, 17′, 18, 19, and 20) downstream of the Pcdh-γ cluster. This CCR comprehensively activates the expression of the Pcdh-β gene cluster in cis, and its deletion dramatically decreases their expression levels. Deleting the CCR nonuniformly down-regulates some Pcdh-γ isoforms and does not affect Pcdh-α expression. Thus, the CCR effect extends beyond the 320-kb region containing the Pcdh-γ cluster to activate the upstream Pcdh-β genes. Thus, we concluded that the CCR is a highly specific regulatory unit for Pcdh-β expression on the clustered Pcdh genomic locus. These findings suggest that each Pcdh cluster is controlled by distinct regulatory elements that activate their expression and that the stochastic gene regulation of the clustered Pcdhs is controlled by the complex chromatin architecture of the clustered Pcdh locus.
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Protocadherin α family is required for serotonergic projections to appropriately innervate target brain areas
The Journal of Neuroscience, 2009Co-Authors: Shota Katori, Shun Hamada, Yukiko Noguchi, Emi Fukuda, Toshifumi Yamamoto, Hideko Yamamoto, Sonoko Hasegawa, Takeshi YagiAbstract:Serotonergic axons from the raphe nuclei in the brainstem project to every region of the brain, where they make connections through their extensive terminal arborizations. This serotonergic innervation contributes to various normal behaviors and psychiatric disorders. The Protocadherin-alpha (Pcdha) family of clustered Protocadherins consists of 14 cadherin-related molecules generated from a single gene cluster. We found that the Pcdhas were strongly expressed in the serotonergic neurons. To elucidate their roles, we examined serotonergic fibers in a mouse mutant (Pcdha(Delta CR/Delta CR)) lacking the Pcdha cytoplasmic region-encoding exons, which are common to the gene cluster. In the first week after birth, the distribution pattern of serotonergic fibers in Pcdha(Delta CR/Delta CR) mice was similar to wild-type, but by 3 weeks of age, when the serotonergic axonal termini complete their arborizations, the distribution of the projections was abnormal. In some target regions, notably the globus pallidus and substantia nigra, the normally even distribution of serotonin axonal terminals was, in the mutants, dense at the periphery of each region, but sparse in the center. In the stratum lacunosum-molecular of the hippocampus, the mutants showed denser serotonergic innervation than in wild-type, and in the dentate gyrus of the hippocampus and the caudate-putamen, the innervation was sparser. Together, the abnormalities suggested that Pcdha proteins are important in the late-stage maturation of serotonergic projections. Further examination of alternatively spliced exons encoding the cytoplasmic tail showed that the A-type (but not the B-type) cytoplasmic tail was essential for the normal development of serotonergic projections.
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clustered Protocadherin family
Development Growth & Differentiation, 2008Co-Authors: Takeshi YagiAbstract:The brain is a complex system composed of enormous numbers of differentiated neurons, and brain structure and function differs among vertebrates. To examine the molecular mechanisms underlying brain structure and function, it is important to identify the molecules involved in generating neural diversity and organization. The clustered Protocadherin (Pcdh) family is the largest subgroup of the diverse cadherin superfamily. The clustered Pcdh proteins are predominantly expressed in the brain and their gene structures in vertebrates are diversified. In mammals, the clustered Pcdh family consists of three gene clusters: Pcdh-alpha, Pcdh-beta, and Pcdh-gamma. During brain development, this family is upregulated by neuronal differentiation, and Pcdh-alpha is then dramatically downregulated by myelination. Clustered Pcdh expression continues in the olfactory bulb, hippocampus, and cerebellum until adulthood. Structural analysis of the first cadherin domain of the Pcdh-alpha protein revealed it lacks the features that classical cadherins require for homophilic adhesiveness, but it contains Pcdh-specific loop structures. In Pcdh-alpha, an RGD motif on a specific loop structure binds beta1-integrin. For gene expression, the gene clusters are regulated by multiple promoters and alternative cis splicing. At the single-cell level, several dozen Pcdh-alpha and -gamma mRNA are regulated monoallelically, resulting in the combinatorial expression of distinct variable exons. The Pcdh-alpha and Pcdh-gamma proteins also form oligomers, further increasing the molecular diversity at the cell surface. Thus, the unique features of the clustered Pcdh family may provide the molecular basis for generating individual cellular diversity and the complex neural circuitry of the brain.
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Protocadherin family diversity structure and function
Current Opinion in Cell Biology, 2007Co-Authors: Hirofumi Morishita, Takeshi YagiAbstract:Protocadherins are predominantly expressed in the nervous system, and constitute the largest subgroup within the cadherin superfamily. The recent structural elucidation of the amino-terminal cadherin domain in an archetypal Protocadherin revealed unique and remarkable features: the lack of an interface for homophilic adhesiveness found in classical cadherins, and the presence of loop structures specific to the Protocadherin family. The unique features of Protocadherins extend to their genomic organization. Recent findings have revealed unexpected allelic and combinatorial gene regulation for clustered Protocadherins, a major subgroup in the Protocadherin family. The unique structural repertoire and unusual gene regulation of the Protocadherin family may provide the molecular basis for the extraordinary diversity of the nervous system.
Raul Araya-secchi - One of the best experts on this subject based on the ideXlab platform.
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Structural determinants of Protocadherin-15 mechanics and function in hearing and balance perception.
Proceedings of the National Academy of Sciences of the United States of America, 2020Co-Authors: Deepanshu Choudhary, Yoshie Narui, Brandon L. Neel, Lahiru N. Wimalasena, Carissa F. Klanseck, Pedro De-la-torre, Conghui Chen, Raul Araya-secchi, Elakkiya Tamilselvan, Marcos SotomayorAbstract:The vertebrate inner ear, responsible for hearing and balance, is able to sense minute mechanical stimuli originating from an extraordinarily broad range of sound frequencies and intensities or from head movements. Integral to these processes is the tip-link protein complex, which conveys force to open the inner-ear transduction channels that mediate sensory perception. Protocadherin-15 and cadherin-23, two atypically large cadherins with 11 and 27 extracellular cadherin (EC) repeats, are involved in deafness and balance disorders and assemble as parallel homodimers that interact to form the tip link. Here we report the X-ray crystal structure of a Protocadherin-15 + cadherin-23 heterotetrameric complex at 2.9-A resolution, depicting a parallel homodimer of Protocadherin-15 EC1-3 molecules forming an antiparallel complex with two cadherin-23 EC1-2 molecules. In addition, we report structures for 10 Protocadherin-15 fragments used to build complete high-resolution models of the monomeric Protocadherin-15 ectodomain. Molecular dynamics simulations and validated crystal contacts are used to propose models for the complete extracellular Protocadherin-15 parallel homodimer and the tip-link bond. Steered molecular dynamics simulations of these models suggest conditions in which a structurally diverse and multimodal Protocadherin-15 ectodomain can act as a stiff or soft gating spring. These results reveal the structural determinants of tip-link-mediated inner-ear sensory perception and elucidate Protocadherin-15's structural and adhesive properties relevant in disease.
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A Mechanically Weak Extracellular Membrane-Adjacent Domain Induces Dimerization of Protocadherin-15.
Biophysical journal, 2018Co-Authors: Pedro De-la-torre, Deepanshu Choudhary, Raul Araya-secchi, Yoshie NaruiAbstract:Abstract The cadherin superfamily of proteins is defined by the presence of extracellular cadherin (EC) “repeats” that engage in protein-protein interactions to mediate cell-cell adhesion, cell signaling, and mechanotransduction. The extracellular domains of nonclassical cadherins often have a large number of EC repeats along with other subdomains of various folds. Protocadherin-15 (PCDH15), a protein component of the inner-ear tip link filament essential for mechanotransduction, has 11 EC repeats and a membrane adjacent domain (MAD12) of atypical fold. Here we report the crystal structure of a pig PCDH15 fragment including EC10, EC11, and MAD12 in a parallel dimeric arrangement. MAD12 has a unique molecular architecture and folds as a ferredoxin-like domain similar to that found in the nucleoporin protein Nup54. Analytical ultracentrifugation experiments along with size-exclusion chromatography coupled to multiangle laser light scattering and small-angle x-ray scattering corroborate the crystallographic dimer and show that MAD12 induces parallel dimerization of PCDH15 near its membrane insertion point. In addition, steered molecular dynamics simulations suggest that MAD12 is mechanically weak and may unfold before tip-link rupture. Sequence analyses and structural modeling predict the existence of similar domains in cadherin-23, Protocadherin-24, and the “giant” FAT and CELSR cadherins, indicating that some of them may also exhibit MAD-induced parallel dimerization.
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A Mechanically Weak Extracellular Membrane-Adjacent Domain Induces Dimerization of Protocadherin-15
2018Co-Authors: Pedro De-la-torre, Deepanshu Choudhary, Raul Araya-secchi, Yoshie NaruiAbstract:The cadherin superfamily of proteins is defined by the presence of extracellular cadherin (EC) repeats that engage in protein-protein interactions to mediate cell-cell adhesion, cell signaling, and mechanotransduction. The extracellular domains of non-classical cadherins often have a large number of EC repeats along with other subdomains of various folds. Protocadherin-15 (PCDH15), a protein component of the inner-ear tip link filament essential for mechanotransduction, has eleven EC repeats and a membrane adjacent domain (MAD12) of atypical fold. Here we report the crystal structure of a pig PCDH15 fragment including EC10, EC11, and MAD12 in a parallel dimeric arrangement. MAD12 has a unique molecular architecture and folds as a ferredoxin-like domain similar to that found in the nucleoporin protein Nup54. Analytical ultracentrifugation experiments along with size exclusion chromatography coupled to multi-angle laser light scattering and small-angle X-ray scattering corroborate the crystallographic dimer and show that MAD12 induces parallel dimerization of PCDH15 near its membrane insertion point. In addition, steered molecular dynamics simulations suggest that MAD12 is mechanically weak and may unfold before tip-link rupture. Sequence analyses and structural modeling predict the existence of similar domains in cadherin-23, Protocadherin-24, and the "giant" FAT and CELSR cadherins, indicating that some of them may also exhibit MAD-induced parallel dimerization.
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An elastic element in the Protocadherin-15 tip link of the inner ear
Nature communications, 2016Co-Authors: Raul Araya-secchi, Brandon L. Neel, Marcos SotomayorAbstract:Tip link filaments convey force and gate inner-ear hair-cell transduction channels to mediate perception of sound and head movements. Cadherin-23 and Protocadherin-15 form tip links through a calcium-dependent interaction of their extracellular domains made of multiple extracellular cadherin (EC) repeats. These repeats are structurally similar, but not identical in sequence, often featuring linkers with conserved calcium-binding sites that confer mechanical strength to them. Here we present the X-ray crystal structures of human Protocadherin-15 EC8-EC10 and mouse EC9-EC10, which show an EC8-9 canonical-like calcium-binding linker, and an EC9-10 calcium-free linker that alters the linear arrangement of EC repeats. Molecular dynamics simulations and small-angle X-ray scattering experiments support this non-linear conformation. Simulations also suggest that unbending of EC9-10 confers some elasticity to otherwise rigid tip links. The new structure provides a first view of Protocadherin-15's non-canonical EC linkers and suggests how they may function in inner-ear mechanotransduction, with implications for other cadherins.
