Role of actin-regulatory gelsolin in control of cell growth

肌动蛋白调节凝溶胶蛋白在控制细胞生长中的作用

• 批准号: 06044009
• 负责人: KUZUMAKI Noboru
• 金额: $ 7.17万
• 依托单位: Hokkaido University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for international Scientific Research
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06044009/
• 关键词: Actin; Gelsolin; Cell growth; Tumor suppression; Mutants; Bladder cancer; Colon cancer; ラス癌遺伝子; フォスホリパーゼ; PIP_2; NIH; 3T3; DNA合成; 胃癌

1. We cloned a mutant gelsolin cDNA His321 from the flat revertant R1 of human activated Ha-ras oncogene-transformed NIH3T3 fibroblasts (EJ-NIH3TS). His321 suppressed tumorigenicity of EJ-NIH3T3. We expressed the His321 and wild-type gelsolin in Escherichia coli, purified them, and analyzed their effects on actin, polyphosphoinositol lipids and phospholipase C.His321 decreased actin-filament-severing activity and increased nucleating activity compared with wild-type gelsolin in vitro. Furthermore, compared to wild-type gelsolin both nucleation and severing by His321 were inhibited more strongly by the phosphoinositol lipids phosphatidylinositol 4-phosphate (PtdInsP) and phosphatidylinositol 4,5-bisphosphate (PtdInsP2). In addition, His321 inhibited PtdInsP2 hydrolysis by phospholipase C gamma 1 more strongly than wild-type gelsolin in vitro because of its higher binding capacity for phosphoinositol Lipid. Our results suggest that the segment G3 of gelsolin which contains the mutation i … More s functionally relevant for regulation of gelsolin's activities and that the region around the residue 321 may contain a phosphoinositol-lipid-binding site. Altered functions of His321 gelsolin might be important for the loss of tumorigenicity of the ras-transformed cells.2. We examined the expression of gelsolin in a number of human bladder and colon cancer cell lines and tissues. In all 6 bladder and 6 of 7 colon cancer cell lines and in 14 of the 18 bladder(77.8%)and 9 of 18 colon(50%)tumor tissues, gelsolin expression was undetectable or extremely low in comparison with its expression in normal bladder or colon epithelial cells. Furthermore, upon the introduction of the exogenous human or mouse authentic gelsolin cDNA into a human bladder cancer cell line UMUC-2 or a human colon cancer cell line LoVo, gelsolin transfectants of UMUC-2 and LoVo greatly reduced the colony-forming ability and the tumorigenicity in vivo. These results suggest that gelsolin plays a key role as a tumor suppressor in human urinary bladder and colon carcinogenesis. Less

1.从人活化Ha-ras癌基因转化的NIH3T3成纤维细胞(EJ-NIH3TS)平板回复突变体R1中克隆了突变的明胶蛋白cDNAHis321。His321抑制EJ-NIH3T3的致瘤性。我们在大肠杆菌中表达并纯化了His321和野生型明胶蛋白，分析了它们对肌动蛋白、肌醇多聚脂和磷脂酶C的影响。与野生型明胶蛋白相比，His321在体外降低了肌动蛋白细丝的切割活性，提高了成核活性。此外，与野生型明胶蛋白相比，His321的成核和切割都受到磷脂酰肌醇4-磷酸(PtdInsP)和磷脂酰肌醇4，5-二磷酸(PtdInsP2)的更强的抑制。此外，His321在体外对磷脂酶C-γ1对PtdInsP2的抑制作用比野生型明胶更强，这是因为His321对磷脂酰肌醇的结合能力更强。我们的结果表明，含有突变I…的明胶蛋白片段G3更多的是S在功能上与调节明胶蛋白的活性有关，并且第321位残基周围的区域可能包含一个肌醇-磷脂结合部位。His321明胶蛋白功能的改变可能是ras转化细胞丧失致瘤性的重要原因。我们研究了明胶蛋白在一些人膀胱癌和结肠癌细胞系和组织中的表达。与正常膀胱或结肠上皮细胞相比，6株膀胱癌和6株结肠癌细胞株、18例膀胱癌组织中14例(77.8%)和18例结肠癌组织中9例(50%)明胶蛋白表达阴性或极低。此外，将外源性人或小鼠真核表达的明胶蛋白基因导入人膀胱癌细胞株UMUC-2或人结肠癌细胞株LoVo后，UMUC-2和LoVo的明胶蛋白基因表达载体显著降低了细胞的集落形成能力和体内致瘤性。这些结果表明，明胶蛋白在人膀胱癌和结肠癌的发生过程中发挥了重要的肿瘤抑制作用。较少

项目成果

Furuuchi, K.: "Growth inhibition of human colon carcinoma cells by gelsolin cDNA transfer." Tumor. Targeting.(in press).

Furuuchi, K.：“通过凝溶胶蛋白 cDNA 转移抑制人结肠癌细胞的生长。”

Ishizaki, A.: "Growth inhibitory functions of a mutated gelsolin (His321) in NIH/3T3 mouse fibroblasts." Exp. Cell Res.217. 448-452 (1995)

Ishizaki, A.：“突变凝溶胶蛋白 (His321) 在 NIH/3T3 小鼠成纤维细胞中的生长抑制功能。”

Fujita, H. ほか: "Enhanced inhibitory effect of the mutated gelsolin His321 on phospolipase Cγ_1." Cell Structure and Function. 18. 591 (1994)

Fujita, H. 等人：“突变凝溶胶蛋白 His321 对磷脂酶 Cγ_1 的增强抑制作用。细胞结构和功能”。

Ishizaki,A.ほか: "Growth inhibitory functions of a mutated gelsolin (His321) in NIH/3T3 mouse fibroblasts." Exp.Cell Res.217. 448-452 (1995)

Ishizaki, A. 等人：“突变凝溶胶蛋白 (His321) 在 NIH/3T3 小鼠成纤维细胞中的生长抑制功能。”Exp.Cell Res.217 (1995)。

Tanaka, M.: "Gelsolin : a candidate for suppressor of fuman bladder cancer" Cancer Res.55. 3228-3232 (1995)

Tanaka, M.：“凝溶胶蛋白：fuman 膀胱癌抑制剂的候选者”Cancer Res.55。