Division of Cell and Developmental Biology, University of Dundee, Dundee DD1 5EH, Scotland, U.K.
Division of Cell Signalling and Immunology, University of Dundee, Dundee DD1 5EH, Scotland, U.K.
Biochem J. 2022 Aug 31;479(16):1709-1725. doi: 10.1042/BCJ20220184.
The protein kinases PAK4, PAK5 and PAK6 comprise a family of ohnologues. In multiple cancers including melanomas PAK5 most frequently carries non-synonymous mutations; PAK6 and PAK4 have fewer; and PAK4 is often amplified. To help interpret these genomic data, initially we compared the cellular regulation of the sister kinases and their roles in melanoma cells. In common with many ohnologue protein kinases, PAK4, PAK5 and PAK6 each have two 14-3-3-binding phosphosites of which phosphoSer99 is conserved. PAK4 localises to the leading edge of cells in response to phorbol ester-stimulated binding of 14-3-3 to phosphoSer99 and phosphoSer181, which are phosphorylated by two different PKCs or PKDs. These phosphorylations of PAK4 are essential for its phorbol ester-stimulated phosphorylation of downstream substrates. In contrast, 14-3-3 interacts with PAK5 in response to phorbol ester-stimulated phosphorylation of Ser99 and epidermal growth factor-stimulated phosphorylation of Ser288; whereas PAK6 docks onto 14-3-3 and is prevented from localising to cell-cell junctions when Ser133 is phosphorylated in response to cAMP-elevating agents via PKA and insulin-like growth factor 1 via PKB/Akt. Silencing of PAK4 impairs viability, migration and invasive behaviour of melanoma cells carrying BRAFV600E or NRASQ61K mutations. These defects are rescued by ectopic expression of PAK4, more so by a 14-3-3-binding deficient PAK4, and barely by PAK5 or PAK6. Together these genomic, biochemical and cellular data suggest that the oncogenic properties of PAK4 are regulated by PKC-PKD signalling in melanoma, while PAK5 and PAK6 are dispensable in this cancer.
蛋白激酶 PAK4、PAK5 和 PAK6 构成了同源蛋白家族。在包括黑色素瘤在内的多种癌症中，PAK5 最常携带非同义突变；PAK6 和 PAK4 则较少；而 PAK4 则经常被扩增。为了帮助解释这些基因组数据，我们首先比较了姐妹激酶的细胞调节及其在黑色素瘤细胞中的作用。与许多同源蛋白激酶一样，PAK4、PAK5 和 PAK6 每个都有两个 14-3-3 结合磷酸化位点，其中磷酸化丝氨酸 99 是保守的。PAK4 响应佛波酯刺激的 14-3-3 与磷酸化丝氨酸 99 和磷酸化丝氨酸 181 的结合，将定位于细胞的前缘，这两个磷酸化位点分别由两种不同的 PKC 或 PKD 磷酸化。PAK4 的这些磷酸化对于其佛波酯刺激的下游底物的磷酸化是必不可少的。相比之下，14-3-3 与 PAK5 相互作用，是响应佛波酯刺激的丝氨酸 99 磷酸化和表皮生长因子刺激的丝氨酸 288 磷酸化；而 PAK6 与 14-3-3 对接，并在响应 cAMP 升高剂的 PKA 和胰岛素样生长因子 1 的 PKB/Akt 介导的 Ser133 磷酸化时，阻止其定位到细胞-细胞连接。沉默 PAK4 会损害携带 BRAFV600E 或 NRASQ61K 突变的黑色素瘤细胞的活力、迁移和侵袭行为。这些缺陷可以通过 PAK4 的异位表达来挽救，更具体地说，通过 14-3-3 结合缺陷的 PAK4 来挽救，而通过 PAK5 或 PAK6 则几乎无法挽救。这些基因组、生化和细胞数据表明，PKC-PKD 信号在黑色素瘤中调节 PAK4 的致癌特性，而 PAK5 和 PAK6 在这种癌症中是可有可无的。
