title: Functional coupling of Rab3-interacting molecule 1 (RIM1) and L-type Ca2+ channels in insulin release.
abstract: Insulin release by pancreatic β-cells is regulated by diverse intracellular signals, including changes in Ca(2+) concentration resulting from Ca(2+) entry through voltage-gated (Ca(V)) channels. It has been reported that the Rab3 effector RIM1 acts as a functional link between neuronal Ca(V) channels and the machinery for exocytosis. Here, we investigated whether RIM1 regulates recombinant and native L-type Ca(V) channels (that play a key role in hormone secretion) and whether this regulation affects insulin release. Whole-cell patch clamp currents were recorded from HEK-293 and insulinoma RIN-m5F cells. RIM1 and Ca(V) channel expression was identified by RT-PCR and Western blot. RIM1-Ca(V) channel interaction was determined by co-immunoprecipitation. Knockdown of RIM1 and Ca(V) channel subunit expression were performed using small interference RNAs. Insulin release was assessed by ELISA. Co-expression of Ca(V)1.2 and Ca(V)1.3 L-type channels with RIM1 in HEK-293 cells revealed that RIM1 may not determine the availability of L-type Ca(V) channels but decreases the rate of inactivation of the whole cell currents. Co-immunoprecipitation experiments showed association of the Ca(V)β auxiliary subunit with RIM1. The lack of Ca(V)β expression suppressed channel regulation by RIM1. Similar to the heterologous system, an increase of current inactivation was observed upon knockdown of endogenous RIM1. Co-immunoprecipitation showed association of Ca(V)β and RIM1 in insulin-secreting RIN-m5F cells. Knockdown of RIM1 notably impaired high K(+)-stimulated insulin secretion in the RIN-m5F cells. These data unveil a novel functional coupling between RIM1 and the L-type Ca(V) channels via the Ca(V)β auxiliary subunit that contribute to determine insulin secretion.
authors.affiliation: Department of Cell Biology, Center for Research and Advanced Studies, National Polytechnic Institute (Cinvestav-IPN), Colonia Zacatenco, México DF, México.
authors.forename: María A
authors.initials: MA
authors.lastname: Gandini
authors.forename: Alejandro
authors.initials: A
authors.lastname: Sandoval
authors.forename: Ricardo
authors.initials: R
authors.lastname: González-Ramírez
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Mori
authors.forename: Michel
authors.initials: M
authors.lastname: de Waard
authors.forename: Ricardo
authors.initials: R
authors.lastname: Felix
mesh_terms: D020746:Calcium Channels, L-Type / Q000096:biosynthesis* / Q000235:genetics; D045744:Cell Line, Tumor; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology*; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D007328:Insulin / Q000235:genetics / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D009419:Nerve Tissue Proteins
chemical_list: D020746:Calcium Channels, L-Type; D019204:GTP-Binding Proteins; D007328:Insulin; C491492:RIMS1 protein, human; D009419:Nerve Tissue Proteins