Sapkota G P, Kieloch A, Lizcano J M, Lain S, Arthur J S, Williams M R, Morrice N, Deak M, Alessi D R
Medical Research Council Protein Phosphorylation Unit, School of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, Scotland.
J Biol Chem. 2001 Jun 1;276(22):19469-82. doi: 10.1074/jbc.M009953200. Epub 2001 Jan 31.
Peutz-Jeghers syndrome is an inherited cancer syndrome that results in a greatly increased risk of developing tumors in those affected. The causative gene is a protein kinase termed LKB1, predicted to function as a tumor suppressor. The mechanism by which LKB1 is regulated in cells is not known. Here, we demonstrate that stimulation of Rat-2 or embryonic stem cells with activators of ERK1/2 or of cAMP-dependent protein kinase induced phosphorylation of endogenously expressed LKB1 at Ser(431). We present pharmacological and genetic evidence that p90(RSK) mediated this phosphorylation in response to agonists that activate ERK1/2 and that cAMP-dependent protein kinase mediated this phosphorylation in response to agonists that activate adenylate cyclase. Ser(431) of LKB1 lies adjacent to a putative prenylation motif, and we demonstrate that full-length LKB1 expressed in 293 cells was prenylated by addition of a farnesyl group to Cys(433). Our data suggest that phosphorylation of LKB1 at Ser(431) does not affect farnesylation and that farnesylation does not affect phosphorylation at Ser(431). Phosphorylation of LKB1 at Ser(431) did not alter the activity of LKB1 to phosphorylate itself or the tumor suppressor protein p53 or alter the amount of LKB1 associated with cell membranes. The reintroduction of wild-type LKB1 into a cancer cell line that lacks LKB1 suppressed growth, but mutants of LKB1 in which Ser(431) was mutated to Ala to prevent phosphorylation of LKB1 were ineffective in inhibiting growth. In contrast, a mutant of LKB1 that cannot be prenylated was still able to suppress the growth of cells.
黑斑息肉综合征是一种遗传性癌症综合征,会使患者患肿瘤的风险大幅增加。致病基因是一种名为LKB1的蛋白激酶,预计其发挥肿瘤抑制因子的作用。LKB1在细胞中的调控机制尚不清楚。在此,我们证明用ERK1/2激活剂或cAMP依赖性蛋白激酶激活剂刺激大鼠2型细胞或胚胎干细胞,会诱导内源性表达的LKB1在Ser(431)位点发生磷酸化。我们提供了药理学和遗传学证据,表明p90(RSK)在响应激活ERK1/2的激动剂时介导了这种磷酸化,而cAMP依赖性蛋白激酶在响应激活腺苷酸环化酶的激动剂时介导了这种磷酸化。LKB1的Ser(431)紧邻一个假定的异戊二烯化基序,我们证明在293细胞中表达的全长LKB1通过在Cys(433)上添加法尼基基团而发生异戊二烯化。我们的数据表明,LKB1在Ser(431)位点的磷酸化不影响异戊二烯化,而异戊二烯化也不影响Ser(431)位点的磷酸化。LKB1在Ser(431)位点的磷酸化不会改变LKB1自身磷酸化或肿瘤抑制蛋白p53的活性,也不会改变与细胞膜相关的LKB1的量。将野生型LKB1重新导入缺乏LKB1的癌细胞系中可抑制生长,但将LKB1的Ser(431)突变为Ala以阻止LKB1磷酸化的突变体在抑制生长方面无效。相比之下,无法进行异戊二烯化的LKB1突变体仍能够抑制细胞生长。