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1.
Proc Natl Acad Sci U S A ; 117(43): 26985-26995, 2020 10 27.
Artigo em Inglês | MEDLINE | ID: mdl-33046653

RESUMO

Current models emphasize that membrane voltage (Vm) depolarization-induced Ca2+ influx triggers the fusion of vesicles to the plasma membrane. In sympathetic adrenal chromaffin cells, activation of a variety of G protein coupled receptors (GPCRs) can inhibit quantal size (QS) through the direct interaction of G protein Gißγ subunits with exocytosis fusion proteins. Here we report that, independently from Ca2+, Vm (action potential) per se regulates the amount of catecholamine released from each vesicle, the QS. The Vm regulation of QS was through ATP-activated GPCR-P2Y12 receptors. D76 and D127 in P2Y12 were the voltage-sensing sites. Finally, we revealed the relevance of the Vm dependence of QS for tuning autoinhibition and target cell functions. Together, membrane voltage per se increases the quantal size of dense-core vesicle release of catecholamine via Vm → P2Y12(D76/D127) → Gißγ → QS → myocyte contractility, offering a universal Vm-GPCR signaling pathway for its functions in the nervous system and other systems containing GPCRs.


Assuntos
Catecolaminas/metabolismo , Células Cromafins/fisiologia , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Vesículas Secretórias/fisiologia , Potenciais de Ação , Animais , Feminino , Células HEK293 , Humanos , Masculino , Camundongos Knockout , Cultura Primária de Células , Ratos Sprague-Dawley , Receptores Purinérgicos P2Y12/metabolismo , Transdução de Sinais
2.
J Gene Med ; 24(4): e3412, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35075722

RESUMO

BACKGROUND: Limb-girdle muscular dystrophy (LGMD) comprises a heterogeneous group of diseases, affecting different muscles, predominantly skeletal muscles and cardiac muscles of the body. LGMD is classified into two main subtypes A and B, which are further subclassified into eight dominant and thirty recessive subtypes. Three genes, namely POPDC1, POPDC2 and POPDC3, encode popeye domain-containing protein (POPDC), and the variants of POPDC1 and POPDC3 genes have been associated with LGMD. METHODS: In the present study, we performed whole-exome sequencing (WES) analysis on a single-family to investigate the hallmark features of LGMD. The results of WES were further confirmed by Sanger sequencing and 3D protein modeling was also conducted. RESULTS: WES data analysis and Sanger sequencing revealed a homozygous missense variant (c.460A>G; p.Lys154Glu) at a highly conserved amino acid position in the POPDC3. Mutations in the POPDC3 gene have been previously associated with recessive limb-girdle muscular dystrophy type 26. 3D protein modeling further suggested that the identified variant might affect the POPDC3 structure and proper function. CONCLUSIONS: The present study confirms the role of POPDC3 in LGMD, and will facilitate genetic counseling of the family to mitigate the risks of the carrier or affects on future pregnancies.


Assuntos
Moléculas de Adesão Celular , Proteínas Musculares , Distrofia Muscular do Cíngulo dos Membros , Moléculas de Adesão Celular/genética , Homozigoto , Humanos , Proteínas Musculares/genética , Músculo Esquelético , Distrofia Muscular do Cíngulo dos Membros/genética , Mutação , Mutação de Sentido Incorreto
3.
Mol Syndromol ; 14(1): 11-20, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36777705

RESUMO

Introduction: Intellectual disability (ID) is a lifelong disability that affects an individual‧s learning capacity and adaptive behavior. Such individuals depend on their families for day-to-day survival and pose a significant challenge to the healthcare system, especially in developing countries. ID is a heterogeneous condition, and genetic studies are essential to unravel the underlying cellular pathway for brain development and functioning. Methods: Here we studied a female index patient, born to a consanguineous Pakistani couple, showing clinical symptoms of ID, ataxia, hypotonia, developmental delay, seizures, speech abnormality, and aggressive behavior. Whole exome sequencing (WES) coupled with Sanger sequencing was performed for molecular diagnosis. Further, 3D protein modeling was performed to see the effect of variant on protein structure. Results: WES identified a novel homozygous missense variant (c.178T>C; p.Tyr60His) in the ANK3 gene. In silico analysis and 3-dimensional (3D) protein modeling supports the deleterious impact of this variant on the encoding protein, which compromises the protein‧s overall structure and function. Conclusion: Our finding supports the clinical and genetic diversity of the ANK3 gene as a plausible candidate gene for ID syndrome. Intelligence is a complex polygenic human trait, and understanding molecular and biological pathways involved in learning and memory can solve the complex puzzle of how cognition develops. Intellectual disability (ID) is defined as a deficit in an individual‧s learning and adaptive behavior at an early age of onset [American Psychiatric Association, 2013]. It is one of the major medical, and cognitive disorders with a prevalence of 1-3% in the population worldwide [Leonard and Wen, 2002]. ID often exists with other disabling mental conditions such as autism, attention deficit hyperactivity disorder, epilepsy, schizophrenia, bipolar disorder, or depression. Almost half of the cases appear to have a genetic explanation that ranges from cytogenetically visible abnormalities to monogenic defects [Flint, 2001; Ropers, 2010; Tucker-Drob et al., 2013]. Intellectual disability is a genetically heterogeneous condition, and more than 700 genes have been identified to cause ID alone or as a part of the syndrome. Research in X-linked ID has identified more than 100 disease-causing genes on the X chromosome that play a role in cognition; however, research into autosomal causes of ID is still ongoing [Vissers et al., 2016].

4.
Adv Sci (Weinh) ; 9(27): e2202263, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-35896896

RESUMO

Large dense-core vesicles (LDCVs) are larger in volume than synaptic vesicles, and are filled with multiple neuropeptides, hormones, and neurotransmitters that participate in various physiological processes. However, little is known about the mechanism determining the size of LDCVs. Here, it is reported that secretogranin II (SgII), a vesicle matrix protein, contributes to LDCV size regulation through its liquid-liquid phase separation in neuroendocrine cells. First, SgII undergoes pH-dependent polymerization and the polymerized SgII forms phase droplets with Ca2+ in vitro and in vivo. Further, the Ca2+ -induced SgII droplets recruit reconstituted bio-lipids, mimicking the LDCVs biogenesis. In addition, SgII knockdown leads to significant decrease of the quantal neurotransmitter release by affecting LDCV size, which is differently rescued by SgII truncations with different degrees of phase separation. In conclusion, it is shown that SgII is a unique intravesicular matrix protein undergoing liquid-liquid phase separation, and present novel insights into how SgII determines LDCV size and the quantal neurotransmitter release.


Assuntos
Neuropeptídeos , Secretogranina II , Vesículas de Núcleo Denso , Hormônios , Lipídeos , Neurotransmissores/metabolismo , Secretogranina II/metabolismo
6.
Nat Commun ; 9(1): 268, 2018 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-29348579

RESUMO

SALM5, a synaptic adhesion molecule implicated in autism, induces presynaptic differentiation through binding to the LAR family receptor protein tyrosine phosphatases (LAR-RPTPs) that have been highlighted as presynaptic hubs for synapse formation. The mechanisms underlying SALM5/LAR-RPTP interaction remain unsolved. Here we report crystal structures of human SALM5 LRR-Ig alone and in complex with human PTPδ Ig1-3 (MeA-). Distinct from other LAR-RPTP ligands, SALM5 mainly exists as a dimer with LRR domains from two protomers packed in an antiparallel fashion. In the 2:2 heterotetrameric SALM5/PTPδ complex, a SALM5 dimer bridges two separate PTPδ molecules. Structure-guided mutations and heterologous synapse formation assays demonstrate that dimerization of SALM5 is prerequisite for its functionality in inducing synaptic differentiation. This study presents a structural template for the SALM family and reveals a mechanism for how a synaptic adhesion molecule directly induces cis-dimerization of LAR-RPTPs into higher-order signaling assembly.


Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/metabolismo , Baculoviridae , Dimerização , Células HEK293 , Humanos , Domínios de Imunoglobulina , Estrutura Quaternária de Proteína
7.
FEBS Lett ; 592(11): 1789-1803, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29754429

RESUMO

Human periostin plays a multifaceted role in remodeling the extracellular matrix milieu by interacting with other proteins and itself in both a heterophilic and homophilic manner. However, the structural mechanism for its extensive interactions has remained elusive. Here, we report the crystal structures of human periostin (EMI-Fas1I-IV ) and its Cys60Ala mutant. In combination with multi-angle light-scattering analysis and biochemical assays, the crystal structures reveal that periostin mainly exists as a dimer in solution and its homophilic interaction is mainly mediated by the EMI domain. Furthermore, Cys60 undergoes cysteinylation as confirmed by mass spectroscopy, and this site hardly affects the homophilic interaction. Also, the structures yield insights into how periostin forms heterophilic interactions with other proteins under physiological or pathological conditions.


Assuntos
Moléculas de Adesão Celular/química , Multimerização Proteica , Substituição de Aminoácidos , Moléculas de Adesão Celular/genética , Cisteína/química , Cisteína/genética , Humanos , Mutação de Sentido Incorreto , Domínios Proteicos , Estrutura Quaternária de Proteína
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