Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 34141, Republic of Korea; Center for Bioanalysis, Korea Research Institute of Standards and Science, Daejeon 34113, Republic of Korea.
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 34141, Republic of Korea.
Biochim Biophys Acta Mol Basis Dis. 2018 Oct;1864(10):3145-3153. doi: 10.1016/j.bbadis.2018.07.001. Epub 2018 Jul 3.
Human ANKRD9 (ankyrin repeat domain 9) expression is altered in some cancers.
We tested genetic association of ANKRD9 with gastric cancer susceptibility and examined functional association of ANKRD9 with altered proliferation of MKN45 gastric cancer cells. We then identified ANKRD9-binding partners in HEK 293 embryonic kidney cells using quantitative proteomics, western blotting and complex reconstitution assays. We finally demonstrated ANKRD9's role of recognizing substrates for ubiquitination using in vitro ubiquitylation assay.
ANKRD9 is associated with cancer susceptibility in a comparison of single-nucleotide polymorphisms between 1092 gastric cancer patients and 1206 healthy controls. ANKRD9 depletion accelerates tumor progression by increasing cellular proliferation, piling up, and anchorage-independent growth of MKN45 cells. We discovered that ANKRD9 is a ubiquitin ligase substrate receptor subunit and has an anti-proliferative activity. ANKRD9 associates with CUL5 (not CUL2), ELOB, ELOC, and presumably RNF7 subunits, which together assemble into a cullin-RING superfamily E3 ligase complex. ANKRD9 belongs to the ASB family of proteins, which are characterized by the presence of ankyrin repeats and a SOCS box. In addition to its interactions with the other E3 ligase subunits, ANKRD9 interacts with two isoforms of inosine monophosphate dehydrogenase (IMPDH). These IMPDH isoforms are cognate substrates of the ANKRD9-containing E3 enzyme, which ubiquitinates them for proteasomal degradation. Their ubiquitination and turnover require the presence of ANKRD9.
ANKRD9, a previously unidentified E3 substrate receptor subunit, functions in tumor suppression by recognizing the oncoprotein IMPDH isoforms for E3 ubiquitination and proteasomal degradation.
人类 ANKRD9（锚蛋白重复结构域 9）的表达在某些癌症中发生改变。
我们测试了 ANKRD9 与胃癌易感性的遗传关联，并研究了 ANKRD9 与 MKN45 胃癌细胞增殖改变的功能关联。然后，我们使用定量蛋白质组学、western blot 和复合物重建测定法，在 HEK 293 胚胎肾细胞中鉴定 ANKRD9 的结合伙伴。最后，我们使用体外泛素化测定法证明了 ANKRD9 识别泛素化底物的作用。
ANKRD9 与 1092 例胃癌患者和 1206 例健康对照者之间的单核苷酸多态性比较相关，与癌症易感性有关。ANKRD9 的缺失通过增加细胞增殖、堆积和 MKN45 细胞的无锚定生长，加速肿瘤进展。我们发现 ANKRD9 是一种泛素连接酶底物受体亚基，具有抗增殖活性。ANKRD9 与 CUL5（而非 CUL2）、ELOB、ELOC 以及推测的 RNF7 亚基结合，这些亚基共同组装成一个 cullin-RING 超家族 E3 连接酶复合物。ANKRD9 属于 ASB 蛋白家族，其特征是存在锚蛋白重复和 SOCS 盒。ANKRD9 除了与其他 E3 连接酶亚基相互作用外，还与肌苷单磷酸脱氢酶 (IMPDH) 的两种同工型相互作用。这些 IMPDH 同工型是含有 ANKRD9 的 E3 酶的同源底物，该酶将其泛素化以进行蛋白酶体降解。它们的泛素化和周转需要 ANKRD9 的存在。
ANKRD9 是一种先前未被识别的 E3 底物受体亚基，通过识别癌蛋白 IMPDH 同工型进行 E3 泛素化和蛋白酶体降解，从而发挥肿瘤抑制作用。
