Abstract
BackgroundThe ankyrin repeat and suppressor of cytokine signalling (SOCS) box proteins (Asbs) are a large protein family implicated in diverse biological processes including regulation of proliferation and differentiation. The SOCS box of Asb proteins is important in a ubiquitination-mediated proteolysis pathway. Here, we aimed to evaluate expression and function of human Asb-9 (ASB9).ResultsWe found that a variant of ASB9 that lacks the SOCS box (ASB9ΔSOCS) was naturally detected in human cell lines but not in peripheral blood mononuclear cells or normal hepatocytes. We also identified ubiquitous mitochondrial creatine kinase (uMtCK) as a new target of ASB9 in human embryonic kidney 293 (HEK293) cells. The ankyrin repeat domains of ASB9 can associate with the substrate binding site of uMtCK in a SOCS box-independent manner. The overexpression of ASB9, but not ASB9ΔSOCS, induces ubiquitination of uMtCK. ASB9 and ASB9ΔSOCS can interact and colocalise with uMtCK in the mitochondria. However, only expression of ASB9 induced abnormal mitochondrial structure and a decrease of mitochondrial membrane potential. Furthermore, the creatine kinase activities and cell growth were significantly reduced by ASB9 but not by ASB9ΔSOCS.ConclusionsASB9 interacts with the creatine kinase system and negatively regulates cell growth. The differential expression and function of ASB9 and ASB9ΔSOCS may be a key factor in the growth of human cell lines and primary cells.
Highlights
The ankyrin repeat and suppressor of cytokine signalling (SOCS) box proteins (Asbs) are a large protein family implicated in diverse biological processes including regulation of proliferation and differentiation
To investigate mitochondrial structure in the cells in detail, we compared the architecture of mitochondria with the aid of DSRed2-Mito and three-dimensional analysis of confocal data and found rounding and swelling of mitochondria in ASB9 expressing cells (Figure 12b and Additional files 1 and 2). These results revealed that the ASB9 interacts and colocalises with ubiquitous mitochondrial creatine kinase (uMtCK) in the mitochondria, leading to structural organisation defects, which occur in a SOCS box-dependent manner
When we measured the activity of CKB and uMtCK after immunoprecipitation with anti-CKB or anti-uMtCK, the results clearly show that the activity of CKB and uMtCK was abolished by ASB9 and decreased by ASB9ΔSOCS
Summary
The ankyrin repeat and suppressor of cytokine signalling (SOCS) box proteins (Asbs) are a large protein family implicated in diverse biological processes including regulation of proliferation and differentiation. The SOCS box of Asb proteins is important in a ubiquitination-mediated proteolysis pathway. The largest family of suppressor of cytokine signalling (SOCS) box-containing superfamily proteins are the ankyrin repeat and SOCS box proteins (Asbs; ASBs in humans). The Asbs have two functional domains, a SOCS box and a variable number of N-terminal ankyrin (ANK) repeats [1]. While SOCS family members use the SH2 domain to recruit substrates, the ANK repeat regions of Asb family members serve as specific protein-protein interaction platforms to recruit target substrates in different biological processes [1]. Asb-15 reportedly regulates protein synthesis in skeletal muscle and alters the differentiation of mouse myoblast and the phosphorylation of mitogen-activated protein kinase and Akt [8,9]
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