Fu Z, Chakraborti T, Morse S, Bennett G S, Shaw G
Department of Neuroscience, University of Florida, Gainesville, Florida 32610, USA.
Exp Cell Res. 2001 Oct 1;269(2):275-86. doi: 10.1006/excr.2001.5324.
The casein kinase I (CKI) family consists of at least seven vertebrate genes, some of which can be alternatively spliced. Previously, we have studied the four splice variants of the chicken CKIalpha gene. The four proteins differ only by the presence or absence of two peptides, a 28-amino-acid "L" insert in the catalytic domain and a 12-amino-acid "S" insert near the extreme C-terminus. Here cells were transfected with DNA encoding all four isoforms fused to the green fluorescent protein (GFP) and the localization of each protein was examined. We noted that the L insert includes the sequence PVGKRKR, which has the characteristics of a nuclear localization signal (NLS), and we show that the CKIalphaL and CKIalphaLS isoforms which contain this sequence are targeted to the nucleus, where a fraction becomes associated with nuclear speckles. In contrast the two isoforms lacking the L insert remain predominantly cytoplasmic. Mutation of the first lysine in the putative NLS to asparagine prevented the nuclear entry of GFP-CKIalphaL. Therefore different CKIalpha isoforms are targeted to different cellular compartments in a fashion modulated by alternate transcription and in these locations presumably phosphorylate and regulate different cellular substrates.
酪蛋白激酶I（CKI）家族由至少七个脊椎动物基因组成，其中一些基因可以进行可变剪接。此前，我们研究了鸡CKIα基因的四种剪接变体。这四种蛋白质的区别仅在于是否存在两种肽段，一种是位于催化结构域的28个氨基酸的“L”插入肽，另一种是靠近C末端的12个氨基酸的“S”插入肽。在此，我们用编码与绿色荧光蛋白（GFP）融合的所有四种同工型的DNA转染细胞，并检测每种蛋白质的定位。我们注意到L插入肽包含序列PVGKRKR，其具有核定位信号（NLS）的特征，并且我们表明含有该序列的CKIαL和CKIαLS同工型定位于细胞核，其中一部分与核斑点相关。相比之下，缺乏L插入肽的两种同工型主要保留在细胞质中。将假定的NLS中的第一个赖氨酸突变为天冬酰胺可阻止GFP-CKIαL进入细胞核。因此，不同的CKIα同工型以交替转录调节的方式定位于不同的细胞区室，并且在这些位置可能磷酸化并调节不同的细胞底物。
