Sun X G, Rotenberg S A
Department of Chemistry and Biochemistry, Queens College, City University of New York, Flushing 11367, USA.
Cell Growth Differ. 1999 May;10(5):343-52.
A nonmetastatic human mammary epithelial cell line (MCF-10A) was engineered to overproduce protein kinase Calpha (PKCalpha) so as to investigate a role for this isoform in the metastatic phenotype. PKCalpha transfectants (clone 26alpha) expressed an 8-fold higher level of PKCalpha protein without compensatory alterations in other isoforms. Clone 26alpha proliferated slowly (accumulating in G1 of the cell cycle) but exhibited pronounced increases in motility and adhesion. Elevated expression of cell cycle inhibitor p27 and focal adhesion proteins was observed, whereas E-cadherin expression decreased to undetectable levels. These observations were consistent with the morphology of PKCalpha transfectants (large, disaggregated, and flat, with lamellipodia and extensive actin fibers) and control cells (small, aggregated, and refractile). Treatment with PKC inhibitors or transfection of a dominant negative (dn) mutant of Rac1, but neither dn RhoA nor dn cdc42, reduced the motility of clone 26alpha, implicating PKCalpha catalytic activity and endogenous Rac1, respectively, in the PKCalpha-induced phenotype. Overall, PKCalpha overexpression suppresses proliferation while endowing MCF-10A cells with properties consistent with the metastatic phenotype.
构建了一种非转移性人乳腺上皮细胞系（MCF - 10A），使其过量表达蛋白激酶Cα（PKCα），以研究该亚型在转移表型中的作用。PKCα转染细胞（克隆26α）表达的PKCα蛋白水平高出8倍，而其他亚型没有代偿性改变。克隆26α增殖缓慢（在细胞周期的G1期积累），但运动性和黏附性显著增加。观察到细胞周期抑制剂p27和黏着斑蛋白的表达升高，而E - 钙黏蛋白表达降至不可检测水平。这些观察结果与PKCα转染细胞（大、分散、扁平，有片状伪足和广泛的肌动蛋白纤维）和对照细胞（小、聚集、有折光性）的形态一致。用PKC抑制剂处理或转染Rac1的显性负性（dn）突变体，但dn RhoA或dn cdc42均不能降低克隆26α的运动性，分别表明PKCα催化活性和内源性Rac1参与了PKCα诱导的表型。总体而言，PKCα过表达抑制增殖，同时赋予MCF - 10A细胞与转移表型一致的特性。
