p57Kip2蛋白是CIP/Kip家族的成员,主要位于细胞核中,发挥其Cyclin/CDKs抑制功能。此外,蛋白质在胚胎发育中起关键作用,分化,和致癌作用取决于其细胞定位和相互作用。CDKN1C突变,编码人类p57Kip2的基因,导致不同遗传疾病的发展,包括Beckwith-Wiedemann,图像和银罗素综合征。我们研究了特定的Beckwith-Wiedemann相关的CDKN1C变化(c.946C>T),该变化导致C端氨基酸(精氨酸316)被色氨酸(R316W-p57Kip2)取代。我们发现R316W-p57Kip2有明显的再分布,而野生型p57Kip2主要发生在细胞核中,突变形式也分布在细胞质中。转染编码p57Kip2N-和C-末端结构域的两个表达构建体,分别,允许在残基220-316之间映射核定位信号(NLS)。此外,通过去除蛋白质C末端的基本RKRLR序列(从312到316个残基),p57Kip2被限制在细胞质中,暗示这个序列是核进入绝对必需的。总之,我们发现了一个未报告的p57Kip2NLS,并提示其缺失或突变可能与CDKN1C相关人类疾病相关,决定了p57Kip2定位/调控作用的显著变化.
p57Kip2 protein is a member of the CIP/Kip family, mainly localized in the nucleus where it exerts its Cyclin/CDKs inhibitory function. In addition, the protein plays key roles in embryogenesis, differentiation, and carcinogenesis depending on its cellular localization and interactors. Mutations of CDKN1C, the gene encoding human
p57Kip2, result in the development of different genetic diseases, including Beckwith-Wiedemann, IMAGe and Silver-Russell syndromes. We investigated a specific Beckwith-Wiedemann associated CDKN1C change (c.946 C>T) that results in the substitution of the C-terminal amino acid (arginine 316) with a tryptophan (R316W-p57Kip2). We found a clear redistribution of R316W-
p57Kip2, in that while the wild-type
p57Kip2 mostly occurs in the nucleus, the mutant form is also distributed in the cytoplasm. Transfection of two expression constructs encoding the
p57Kip2 N- and C-terminal domain, respectively, allows the mapping of the nuclear localization signal(s) (NLSs) between residues 220-316. Moreover, by removing the basic RKRLR sequence at the protein C-terminus (from 312 to 316 residue), p57Kip2 was confined in the cytosol, implying that this sequence is absolutely required for nuclear entry. In conclusion, we identified an unreported
p57Kip2 NLS and suggest that its absence or mutation might be of relevance in CDKN1C-associated human diseases determining significant changes of p57Kip2 localization/regulatory roles.