Regulation of intracellular localization and transcriptional activity of FOXO4 by protein kinase B through phosphorylation at the motif sites conserved among the FOXO family
Regulation of intracellular localization and transcriptional activity of FOXO4 by protein kinase B through phosphorylation at the motif sites conserved among the FOXO family
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DOI：
10.1093/jb/mvi146
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发表时间：
2005-10-01
影响因子：
2.7
通讯作者：
Kikkawa, U
中科院分区：
文献类型：
--
作者：
Matsuzaki, H;Ichino, A;Kikkawa, U
FOXO4转录因子又称AFX，含有蛋白激酶B(PYB)、Thr32、Ser197和Ser262三个可能的磷酸化基序，推测磷酸化的FOXO4停留在胞浆中，通过去磷酸化进入细胞核诱导靶基因表达。体外磷酸化多肽分析和磷酸化位点特异性抗体免疫印迹实验表明，这三个位点均被PKB磷酸化。Ala取代Thr32或Ser197的突变体主要分布在细胞核中，但不存在胞浆部分，经血小板衍生生长因子处理后，它们在细胞中的分布没有改变。在这两个位点突变的FOXO4蛋白的转录活性是野生型的3-5倍。相反，Ser262的替换并没有改变定位或转录活性。这些结果表明，Thr32和Ser197处的磷酸化对于调节FOXO4的核定位和转录活性是必不可少的，而Ser262处的磷酸化不是关键的。这些特性类似于FOXO1和FOX03，因此FOXO转录因子似乎通过生长因子信号通路中PKB的共同机制来调节。
FOXO4 transcription factor, also referred to AFX, contains three putative phosphorylation motif sites for protein kinase B (PYB), Thr32, Ser197, and Ser262, and it is proposed that phosphorylated FOXO4 stays in the cytosol and is imported to the nucleus through dephosphorylation to induce target gene expression. These three sites were revealed to be phosphorylated by PKB in vitro on phosphopeptide analysis, and in cultured cells on immunoblotting with phosphorylation-site specific antibodies. The mutants with either Thr32 or Ser197 replaced by Ala were found mostly in the nuclear but not the cytosol fraction, and treatment with platelet-derived growth factor did not change their distributions in the cells. FOXO4 proteins mutated at these two sites showed 3- to 5-fold higher transcriptional activity than that of the wild type. In contrast, the replacement of Ser262 did not alter the localization or transcriptional activity. These results indicate that phosphorylation at Thr32 and Ser197 is indispensable, whereas that at Ser262 is not critical, for regulation of the nuclear localization and transcriptional activity of FOXO4. These properties are similar to those of FOXO1 and FOXO3, and thus FOXO transcription factors seem to be regulated through a common mechanism by PKB in the growth factor signaling pathway.