Prié D, Ronco P M, Baudouin B, Géniteau-Legendre M, Antoine M, Piedagnel R, Estrade S, Lelongt B, Verroust P J, Cassingéna R
Institut National de la Santé et de la Recherche Médicale (INSERM) U.64, Hôpital Tenon, Paris, France.
J Cell Biol. 1991 May;113(4):951-62. doi: 10.1083/jcb.113.4.951.
To analyze the role of SV40 genome in the phenotypic alterations previously observed in SV40-transformed cell lines, we infected rabbit renal cortical cells with a temperature-sensitive SV40 mutant strain (tsA58) and compared the cell phenotypes at temperatures permissive (33 degrees C) and restrictive (39.5 degrees C) for SV40 genome expression. At both temperatures, the resulting cell line (RC.SVtsA58) expresses cytokeratin and uvomorulin, but epithelial differentiation is more elaborate at 39.5 degrees C as shown by the formation of a well-organized cuboidal monolayer with numerous tight junctions and desmosomes. Functional characteristics are also markedly influenced by the culture temperature: cells grown at 33 degrees C respond only to isoproterenol (ISO, 10(-6) M) by a sevenfold increase in cAMP cell content above basal values; in contrast, when transferred to 39.5 degrees C, they exhibit increased sensitivity to ISO (ISO/basal: 19.1) and a dramatic response to 10(-7) M dDarginine vasopressin (dDAVP/basal: 18.2, apparent Ka: 5 X 10(-9) M) which peaks 48 h after the temperature shift. The latter is associated with membrane expression of V2-type AVP receptors (approximately 50 fmol/10(6) cells) which are undetectable when SV40 genome is activated (33 degrees C). Clonal analysis, additivity studies, and desensitization experiments argue for the presence of a single cell type responsive to both AVP and ISO. The characteristics of the RC. SVtsA58 cell line at 39.5 degrees C (effector-stimulated cAMP profile, lack of expression of brush-border hydrolases and Tamm-Horsfall protein) suggest that it originates from the cortical collecting tubule, and probably from principal cells.
为了分析SV40基因组在先前在SV40转化细胞系中观察到的表型改变中的作用，我们用温度敏感型SV40突变株（tsA58）感染兔肾皮质细胞，并比较了在允许温度（33℃）和限制温度（39.5℃）下SV40基因组表达时的细胞表型。在这两个温度下，所得细胞系（RC.SVtsA58）均表达细胞角蛋白和桥粒芯糖蛋白，但在39.5℃时上皮分化更精细，表现为形成具有大量紧密连接和桥粒的组织良好的立方单层。功能特性也明显受培养温度的影响：在33℃下生长的细胞仅对异丙肾上腺素（ISO，10⁻⁶ M）有反应，细胞内cAMP含量比基础值增加7倍；相反，当转移到39.5℃时，它们对ISO的敏感性增加（ISO/基础值：19.1），并且对10⁻⁷ M去精氨酸血管加压素（dDAVP/基础值：18.2，表观Ka：5×10⁻⁹ M）有显著反应，在温度转变后48小时达到峰值。后者与V2型AVP受体的膜表达相关（约50 fmol/10⁶个细胞），当SV40基因组被激活时（33℃）则无法检测到。克隆分析、加和性研究和脱敏实验表明存在对AVP和ISO均有反应的单一细胞类型。RC.SVtsA58细胞系在39.5℃时的特性（效应物刺激的cAMP谱、刷状缘水解酶和Tamm-Horsfall蛋白表达缺失）表明它起源于皮质集合管，可能来自主细胞。
