Journal Article

Calpain-7 binds to CHMP1B at its second α-helical region and forms a ternary complex with IST1

Yuki Maemoto, Yohei Osako, Emi Goto, Eri Nozawa, Hideki Shibata and Masatoshi Maki

in The Journal of Biochemistry

Published on behalf of The Japanese Biochemical Society

Volume 150, issue 4, pages 411-421
Published in print October 2011 | ISSN: 0021-924X
Published online May 2011 | e-ISSN: 1756-2651 | DOI: https://dx.doi.org/10.1093/jb/mvr071
Calpain-7 binds to CHMP1B at its second α-helical region and forms a ternary complex with IST1

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Some intracellular proteins involved in the endosomal sorting complex required for transport (ESCRT) system have microtubule interacting and transport (MIT) domains and bind to ESCRT-III protein family members named charged multivesicular body proteins (CHMPs) at their C-terminal regions containing MIT-interacting motifs (MIMs). While two types of MIMs (MIM1 and MIM2) have been reported, CHMP1B has MIM1 and IST1 has both MIM1 and MIM2. Previously, we demonstrated that CHMP1B and IST1 directly interacted with a tandem repeat of MIT domains of calpain-7 (CL7MIT) and that autolytic activity of calpain-7 was enhanced by IST1 in vitro but not by overexpression of IST1 in HEK293T cells. In this study, we detected enhancement of autolysis of mGFP-fused calpain-7 by coexpression with CHMP1B and observed further activation by additional coexpression of IST1 in HEK293T cells. We found that CL7MIT interacted with the second α-helical region of CHMP1B but not with the canonical C-terminal region containing MIM1 in vitro. Co-immunoprecipitation assays demonstrated that the interaction between CL7MIT and CHMP1B and between CL7MIT and IST1 became stronger when IST1 or CHMP1B was additionally coexpressed, suggesting formation of ternary complex of calpain-7, IST1 and CHMP1B. Moreover, subcellular fractionation analyses revealed increase of calpain-7 in membrane/organelle fractions by concomitant overexpression of these ESCRT-III family member proteins.

Keywords: calpain; CHMP; ESCRT; IST1; MIT domain

Journal Article.  6781 words.  Illustrated.

Subjects: Biochemistry

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