Keryer G, Rios R M, Landmark B F, Skalhegg B, Lohmann S M, Bornens M
Centre de Génétique Moléculaire, Centre National de la Recherche Scientifique, Gif/Yvette, France.
Exp Cell Res. 1993 Feb;204(2):230-40. doi: 10.1006/excr.1993.1029.
In the human lymphoblastic cell line KE 37, Northern blot analysis with cDNA probes for human regulatory subunits RII alpha RII beta of the cAMP-dependent protein kinase (A-kinase) type II and immunoblotting or immunoprecipitation studies with several antibodies directed against RII alpha and RII beta show that these two isoforms are expressed. The major isoform alpha is mostly cytosolic, whereas the beta isoform appears concentrated in the Golgi-centrosomal area, as judged by immunofluorescence and cell fractionation. Using a 32P-labelled RII overlay on Western blots, a 350-kDa RII-binding protein (AKAP 350) was specifically identified in centrosomes isolated from this cell line, whereas a Golgi fraction has previously been demonstrated to contain an 85-kDa RII-binding protein (AKAP 85). AKAP 350 is highly insoluble and can partially be extracted from centrosomes as a complex of AKAP 350 and RII subunit. AKAP 350 was identified as a specific centrosomal protein previously demonstrated in the pericentriolar material. The potential significance of a specific subcellular distribution for different RII-binding proteins in nonneuronal cells is discussed.
在人淋巴母细胞系KE 37中，用针对II型环磷酸腺苷依赖性蛋白激酶（A激酶）的人调节亚基RIIα和RIIβ的cDNA探针进行Northern印迹分析，以及用几种针对RIIα和RIIβ的抗体进行免疫印迹或免疫沉淀研究表明，这两种同工型均有表达。通过免疫荧光和细胞分级分离判断，主要的同工型α大多存在于胞质中，而β同工型似乎集中在高尔基体 - 中心体区域。在从该细胞系分离的中心体中，利用Western印迹上的32P标记的RII覆盖物，特异性鉴定出一种350 kDa的RII结合蛋白（A激酶锚定蛋白350，AKAP 350），而先前已证明高尔基体部分含有一种85 kDa的RII结合蛋白（A激酶锚定蛋白85，AKAP 85）。AKAP 350高度不溶性，并且可以作为AKAP 350和RII亚基的复合物从中心体中部分提取出来。AKAP 350被鉴定为先前在中心粒周围物质中发现的一种特异性中心体蛋白。讨论了不同RII结合蛋白在非神经元细胞中的特定亚细胞分布的潜在意义。
