Gilbert W, Siebel C W, Guthrie C
Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0448, USA.
RNA. 2001 Feb;7(2):302-13. doi: 10.1017/s1355838201002369.
Mammalian SR proteins are currently thought to function in mRNA export as well as splicing. They contain multiple phosphorylated serine/arginine (RS/SR) dipeptides. Although SR domains can be phosphorylated by many kinases in vitro, the physiologically relevant kinase(s), and the role(s) of these modifications in vivo have remained unclear. Npl3 is a shuttling protein in budding yeast that we showed previously to be a substrate for the mammalian SR protein kinase, SRPK1, as well as the related yeast kinase, Sky1. Here we demonstrate that Sky1p phosphorylates only one of Npl3p's eight SR/RS dipeptides. Mutation of the C-terminal RS to RA, or deletion of SKY1, results in the cytoplasmic accumulation of Npl3p. The redistribution of Npl3p is accompanied by its increased association with poly(A)+ RNA and decreased association with its import receptor, Mtr10p, in vivo. We propose that phosphorylation of Npl3p by the cytoplasmically localized Sky1p is required for efficient release of mRNA upon termination of export.
目前认为哺乳动物的SR蛋白在mRNA输出以及剪接过程中发挥作用。它们含有多个磷酸化的丝氨酸/精氨酸（RS/SR）二肽。虽然SR结构域在体外可被多种激酶磷酸化，但生理相关的激酶以及这些修饰在体内的作用仍不清楚。Npl3是芽殖酵母中的一种穿梭蛋白，我们之前已证明它是哺乳动物SR蛋白激酶SRPK1以及相关酵母激酶Sky1的底物。在此我们证明，Sky1p仅磷酸化Npl3p的八个SR/RS二肽中的一个。将C末端的RS突变为RA，或缺失SKY1，会导致Npl3p在细胞质中积累。在体内，Npl3p的重新分布伴随着其与多聚腺苷酸（poly(A)+）RNA的结合增加以及与其输入受体Mtr10p的结合减少。我们提出，由定位于细胞质的Sky1p对Npl3p进行磷酸化是在输出终止时有效释放mRNA所必需的。
