Koch Daniela, Sakurai Michiharu, Hummitzsch Katja, Hermsdorf Thomas, Erdmann Sabine, Schwalbe Steffen, Stolzenburg Jens-Uwe, Spanel-Borowski Katharina, Ricken Albert M
Institute of Anatomy, University of Leipzig, Leipzig, Germany.
Growth Factors. 2009 Apr;27(2):100-13. doi: 10.1080/08977190802707571.
We report the presence of KIT variants in granulosa and thecal cells of the follicle and endothelial and steroidogenic cells of the corpus luteum. Transcripts of both full-length splice variants, KIT and KITA, were ubiquitously detected in all cell types, in contrast to transcripts for truncated KIT. RT-PCR with exon-intron-specific primers suggested that KIT transcripts retained intron sequences. We used domain-specific KIT antibodies to identify truncated KIT proteins in cell conditioned media and lysates. These proteins represented soluble KIT and a so far disregarded intracellular KIT fragment, and were ubiquitously present. In contrast, glycosylated variants of full-length KIT were predominantly detected in thecal and endothelial cells. All KIT variants were encountered again in COS-7 cells transfected with a vector containing KITA. Phorbol 12-myristate-13-acetate treatment induced levels of truncated KITs, and this effect was repressed by the metalloproteinase inhibitor TAPI-1. Our findings show that ectodomain cleavage of full-length KIT generates an intracellular KIT. Our experiments suggest that replenishing full-length KIT differs among various ovarian cell types.
我们报告了在卵泡的颗粒细胞和膜细胞以及黄体的内皮细胞和类固醇生成细胞中存在KIT变体。与截短型KIT的转录本不同,全长剪接变体KIT和KITA的转录本在所有细胞类型中均普遍检测到。使用外显子-内含子特异性引物进行的RT-PCR表明,KIT转录本保留了内含子序列。我们使用结构域特异性KIT抗体来鉴定细胞条件培养基和裂解物中的截短型KIT蛋白。这些蛋白代表可溶性KIT和一个迄今被忽视的细胞内KIT片段,并且普遍存在。相比之下,全长KIT的糖基化变体主要在膜细胞和内皮细胞中检测到。在用含有KITA的载体转染的COS-7细胞中再次遇到了所有KIT变体。佛波醇12-肉豆蔻酸酯-13-乙酸酯处理诱导了截短型KIT的水平,并且这种效应被金属蛋白酶抑制剂TAPI-1所抑制。我们的研究结果表明,全长KIT的胞外域裂解会产生一种细胞内KIT。我们的实验表明,全长KIT的补充在不同的卵巢细胞类型中有所不同。