Oligomeric state, hydrodynamic properties and target recognition of human Calcium and Integrin Binding protein 2 (CIB2)

Calcium- and Integrin-Binding protein 2 (CIB2) is a small and ubiquitously expressed protein with largely unknown biological function but ascertained role in hearing physiology and disease. Recent studies found that CIB2 binds Ca2+ with moderate affinity and dimerizes under conditions mimicking the physiological ones. Here we provided new lines of evidence on CIB2 oligomeric state and the mechanism of interaction with the alpha 7B integrin target. Based on a combination of native mass spectrometry, chemical cross-linking/mass spectrometry, analytical gel filtration, dynamic light scattering and molecular dynamics simulations we conclude that CIB2 is monomeric under all tested conditions and presents uncommon hydrodynamic properties, most likely due to the high content of hydrophobic solvent accessible surface. Surface plasmon resonance shows that the interaction with alpha 7B occurs with relatively low affinity and is limited to the cytosolic region proximal to the membrane, being kinetically favored in the presence of physiological Mg2+ and in the absence of Ca2+. Although CIB2 binds to an alpha 7B peptide in a 1:1 stoichiometry, the formation of the complex might induce binding of another CIB2 molecule

A Novel C-Terminal CIB2 (Calcium and Integrin Binding Protein 2) Mutation Associated with Non-Syndromic Hearing Loss in a Hispanic Family

Hearing loss is a complex disorder caused by both genetic and environmental factors. Previously, mutations in CIB2 have been identified as a common cause of genetic hearing loss in Pakistani and Turkish populations. Here we report a novel (c.556C\u3eT; p.(Arg186Trp)) transition mutation in the CIB2 gene identified through whole exome sequencing (WES) in a Caribbean Hispanic family with non-syndromic hearing loss. CIB2 belongs to the family of calcium-and integrin-binding (CIB) proteins. The carboxy-termini of CIB proteins are associated with calcium binding and intracellular signaling. The p.(Arg186Trp) mutation is localized within predicted type II PDZ binding ligand at the carboxy terminus. Our ex vivo studies revealed that the mutation did not alter the interactions of CIB2 with Whirlin, nor its targeting to the tips of hair cell stereocilia. However, we found that the mutation disrupts inhibition of ATP-induced Ca2+ responses by CIB2 in a heterologous expression system. Our findings support p.(Arg186Trp) mutation as a cause for hearing loss in this Hispanic family. In addition, it further highlights the necessity of the calcium binding property of CIB2 for normal hearing

Genetic linkage analysis of DFNB40 and DFNB48 loci in families with autosomal recessive non-syndromic hearing loss (ARNSHL) from western provinces of Iran

Background: Sensorineural hearing loss (SNHL) is the most common sensory disorder and 1 in every 500-1000 newborns is affected. Non-syndromic SNHL accounts for 70% of hereditary hearing loss and 80% of SNHL cases have an autosomal recessive mode of inheritance (ARNSHL). The Purpose of the recent study is genetic linkage analysis to determine the prevalence of DFNB40 and DFNB48 loci in studying families with ARNSHL from the western provinces of Iran. Methods: In this study, 60 families from 3 provinces of Iran involving Hamedan, Kohgiluyeh and Boyer-Ahmad and Chaharmahal and Bakhtiari with autosomal recessive non syndromic hearing loss were examined. The selected families in this study were consanguineous and had at least two patients. They also were negative for GJB2 mutations. Linkage analysis was performed by using 6 markers short tandem repeat (STR) for the DFNB40 locus and 7 markers STR for the DFNB48 locus. Findings: After examining different families, it was revealed that none of them showed linkage to the DFNB40 and DFNB48 loci. Conclusion: The recent study suggests that DFNB40 and DFNB48 loci might not play an important role in causing hearing loss in the mentioned provinces. However, further studies are necessary to determine more precisely the role of these loci in the Iranian population. © 2016, Isfahan University of Medical Sciences(IUMS). All rights reserved

Calcium- and Integrin-Binding Protein 2 (CIB2) in physiology and disease: bright and dark sides

Calcium- and integrin-binding protein 2 (CIB2) is a small EF-hand protein capable of binding Mg2+ and Ca2+ ions. While its biological function remains largely unclear, an increasing number of studies have shown that CIB2 is an essential component of the mechano-transduction machinery that operates in cochlear hair cells. Mutations in the gene encoding CIB2 have been associated with non-syndromic deafness. In addition to playing an important role in the physiology of hearing, CIB2 has been implicated in a multitude of very different processes, ranging from integrin signaling in platelets and skeletal muscle to autophagy, suggesting extensive functional plasticity. In this review, we summarize the current understanding of biochemical and biophysical properties of CIB2 and the biological roles that have been proposed for the protein in a variety of processes. We also highlight the many molecular aspects that remain unclarified and deserve further investigation

Genetic linkage analysis of the DFNB48 and DFNB98 loci in families with Autosomal Recessive Non-Syndromic Hearing Loss (ARNSHL) from Khouzestan province

Background and aims: Hearing loss, a sensorineural disorder, is one of the most common congenital impairments, occurring in approximately 1 in 500 newborns. Hearing loss is a highly heterogenic disease and half of the cases of deafness are attributed to genetic causes; environmental and unknown factors account for the remainder. Non-syndromic type forms 70% of hearing loss cases. Pattern of inheritance of nearly 80% of this type of HL is recessive autosomal. Iranian population provides a valuable genetic resource to study this kind of HL because of high ratio of consanguinity. In this study, genetic linkage of DFNB48 (CIB2) and DFNB98 (TSPEAR) is investigated in families with ARNSHL impairment from Khouzestan province. Methods: In this descriptive study 300 individuals of 25 families with hearing loss were examined in order to determine type and frequency of mutation of DFNB48 and DFNB98 loci in Khouzestan province. Families' selection had some criteria. Families with healthy parents, consanguineous marriage and negative result for mutations of GJB2 gene with at least two affected individuals were selected. 3 families which were detected positive for mutations of GJB2 gene were excluded from study. Linkage analysis was done for 22 families by using six STR markers which were located in or were tightly linked to each locus. Results: None of these families inspected by linkage analysis was linked to the DFNB48 or DFNB98 loci. Conclusion: Considering these results it seems that CIB2 and TSPEAR genes mutations have not important roles in hearing loss in Khouzestan province

Integrated multiomics approach identifies calcium and integrin-binding protein-2 as a novel gene for pulse wave velocity

Background: Carotid-femoral pulse wave velocity (PWV) is an important measure of arterial stiffness, which is an independent predictor of cardiovascular morbidity and mortality. In this study, we used an integrated genetic, epigenetic and transcriptomics approach to uncover novel molecular mechanisms contributing to PWV. Methods and results: We measured PWV in 1505 healthy twins of European descendent. A genomewide association analysis was performed using standardized residual of the inverse of PWV. We identified one single-nucleotide polymorphism (rs7164338) in the calcium and integrin-binding protein-2 (CIB2) gene on chromosome 15q25.1 associated with PWV [beta = -0.359, standard error (SE) = 0.07, P = 4.8 x 10(-8)]. The same variant was also associated with increased CIB2 expression in leucocytes (beta = 0.034, SE = 0.008, P = 4.95 x 10(-5)) and skin (beta = 0.072, SE = 0.01, P = 2.35 x 10(-9)) and with hypomethylation of the gene promoter (beta = -.899, SE = 0.098, P = 3.63 x 10(-20)). Conclusion: Our data indicate that reduced methylation of the CIB2 promoter in individuals carrying rs7164338 may lead to increased CIB2 expression. Given that CIB2 is thought to regulate intracellular calcium levels, an increase in protein levels may prevent the accumulation of serum calcium and phosphate, ultimately slowing down the process of vascular calcification. This study shows the power of integrating multiple omics to discover novel cardiovascular mechanisms

Gene Expression Profiles of the Cochlea and Vestibular Endorgans: Localization and Function of Genes Causing Deafness

Objectives: We sought to elucidate the gene expression profiles of the causative genes as well as the localization of the encoded proteins involved in hereditary hearing loss. Methods: Relevant articles (as of September 2014) were searched in PubMed databases, and the gene symbols of the genes reported to be associated with deafness were located on the Hereditary Hearing Loss Honnepage using localization, expression, and distribution as keywords. Results: Our review of the literature allowed us to systematize the gene expression profiles for genetic deafness in the inner ear, clarifying the unique functions and specific expression patterns of these genes in the cochlea and vestibular endorgans. Conclusions: The coordinated actions of various encoded molecules are essential for the normal development and maintenance of auditory and vestibular function.ArticleANNALS OF OTOLOGY RHINOLOGY AND LARYNGOLOGY. 124:6S-48S (2015)journal articl

Evolutionary-Conserved Allosteric Properties of Three Neuronal Calcium Sensor Proteins

Neuronal Calcium Sensors (NCS) are highly conserved proteins specifically expressed in neurons. Calcium (Ca2+)-binding to their EF-hand motifs results in a conformational change, which is crucial for the recognition of a specific target and the downstream biological process. Here we present a comprehensive analysis of the allosteric communication between Ca2+-binding sites and the target interfaces of three NCS, namely NCS1, recoverin (Rec), and GCAP1. In particular, Rec was investigated in different Ca2+-loading states and in complex with a peptide from the Rhodopsin Kinase (GRK1) while NCS1 was studied in a Ca2+-loaded state in complex with either the same GRK1 target or a peptide from the D2 Dopamine receptor. A Protein Structure Network (PSN) accounting for persistent non-covalent interactions between amino acids was built for each protein state based on exhaustive Molecular Dynamics simulations. Structural network analysis helped unveiling the role of key amino acids in allosteric mechanisms and their evolutionary conservation among homologous proteins. Results for NCS1 highlighted allosteric inter-domain interactions between Ca2+-binding motifs and residues involved in target recognition. Robust long range, allosteric protein-target interactions were found also in Rec, in particular originating from the EF3 motif. Interestingly, Tyr 86, involved the hydrophobic packing of the N-terminal domain, was found to be a key residue for both intra- and inter-molecular communication with EF3, regardless of the presence of target or Ca2+ ions. Finally, based on a comprehensive topological PSN analysis for Rec, NCS1, and GCAP1 and multiple sequence alignments with homolog proteins, we propose that an evolution-driven correlation may exist between the amino acids mediating the highest number of persistent interactions (high-degree hubs) and their conservation. Such conservation is apparently fundamental for the specific structural dynamics required in signaling events

Evolutionary-conserved allosteric properties of three neuronal calcium sensor proteins

Neuronal Calcium Sensors (NCS) are highly conserved proteins specifically expressed in neurons. Calcium (Ca2+)-binding to their EF-hand motifs results in a conformational change, which is crucial for the recognition of a specific target and the downstream biological process. Here we present a comprehensive analysis of the allosteric communication between Ca2+-binding sites and the target interfaces of three NCS, namely NCS1, recoverin (Rec), and GCAP1. In particular, Rec was investigated in different Ca2+-loading states and in complex with a peptide from the Rhodopsin Kinase (GRK1) while NCS1 was studied in a Ca2+-loaded state in complex with either the same GRK1 target or a peptide from the D-2 Dopamine receptor. A Protein Structure Network (PSN) accounting for persistent non-covalent interactions between amino acids was built for each protein state based on exhaustive Molecular Dynamics simulations. Structural network analysis helped unveiling the role of key amino acids in allosteric mechanisms and their evolutionary conservation among homologous proteins. Results for NCS1 highlighted allosteric inter-domain interactions between Ca2+-binding motifs and residues involved in target recognition. Robust long range, allosteric protein-target interactions were found also in Rec, in particular originating from the EF3 motif. Interestingly, Tyr 86, involved the hydrophobic packing of the N-terminal domain, was found to be a key residue for both intra- and inter-molecular communication with EF3, regardless of the presence of target or Ca2+ ions. Finally, based on a comprehensive topological PSN analysis for Rec, NCS1, and GCAP1 and multiple sequence alignments with homolog proteins, we propose that an evolution-driven correlation may exist between the amino acids mediating the highest number of persistent interactions (high-degree hubs) and their conservation. Such conservation is apparently fundamental for the specific structural dynamics required in signaling events