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Interactome Analysis of BetaM Protein Encoded by ATP1B4 Gene Using Two‐hybrid Systems
Author(s) -
Pestov Nikolay,
Korneenko Tatyana,
Modyanov Nikolai
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.567.16
Subject(s) - interactome , integral membrane protein , transmembrane domain , two hybrid screening , regulator , biology , gene , protein subunit , transmembrane protein , microbiology and biotechnology , complementary dna , membrane protein , computational biology , chemistry , biochemistry , membrane , receptor
The BetaM, a type II integral membrane protein, became a muscle‐specific protein of inner nuclear membrane in eutherians through co‐option of the ATP1B4 gene that encodes Na,K‐ATPase β‐subunit in lower vertebrates. Earlier we determined that BetaM directly interacts with transcriptional co‐regulator SKIP and functions as a regulator of gene expression. Here we present the expanded search for BetaM interactors by screening of cDNA libraries using conventional yeast two‐hybrid and split‐ubiquitin systems. The list of identified BetaM interactors includes BetaM itself, SKIP, Syne‐1, sarcoglycan β, LZIP, RCN 3, LAP1, HO 1, HO2, PHD finger 3, cadherin16, frizzled 1, SC65, tetraspanins 3 and CD63, TGN29, ERGIC 3, BCRAP‐31, C4orf3 and SftpC. Only sarcoglycan β was identified by both assays. Analysis of BetaM mutants revealed that deletion of 56 N‐terminal residues including the Arg‐rich peptide and a half of Glu‐rich stretch does not disrupt the BetaM dimerization and other identified interactions. Transmembrane and/or C‐terminal domains are required for interactions with LZIP, Syne, reticulocalbin 3 and sarcoglycan β. N‐terminal domain seems to be sufficient for interaction with SKIP, LAP‐1, PHF‐3 and heme oxygenases. Further analysis indicates that the second half of the Glu‐rich intrinsically disordered N‐terminal domain is required for the interaction with SKIP. We also show that the relatively conserved fragments of N‐terminal domain adjacent to the membrane are sufficient for BetaM dimerization. Supported by RFBR (Grant 13‐04‐01413) and by Dept of Physiology & Pharmacology, University of Toledo College of Medicine.