Department of Cell Biology, School of Medicine, Johns Hopkins University, Baltimore, MD 21205, USA.
Mol Biol Cell. 2011 Feb 15;22(4):437-47. doi: 10.1091/mbc.E10-06-0522. Epub 2010 Dec 17.
By limiting phosphotidylinositol 3,4,5-triphosphate (PIP(3)) levels, tumor suppressor PTEN not only controls cell growth but also maintains cell polarity required for cytokinesis and chemotaxis. To identify the critical targets of PIP(3) that link it to the cytoskeleton, we deleted secondary genes to reverse the deficiencies of pten- cells in Dictyostelium. The polarity defects in pten- cells correlate with elevated phosphorylations of PKB substrates. Deletion of AKT orthologue, PkbA, or a subunit of its activator TORC2, reduced the phosphorylations and suppressed the cytokinesis and chemotaxis defects in pten- cells. In these double mutants, the excessive PIP(3) levels and, presumably, activation of other PIP(3)-binding proteins had little or no effect on the cytoskeleton. In bands with increased phosphorylation in pten- cells, we found PKB substrates, PI5K, GefS, GacG, and PakA. Disruption of PakA in pten- cells restored a large fraction of the cells to normal behavior. Consistently, expression of phosphomimetic PakA in pten- cells exacerbated the defects but nonphosphorylatable PakA had no effect. Thus, among many putative PTEN- and PIP(3)-dependent events, phosphorylation of PKB substrates is the key downstream regulator of cell polarity.
通过限制磷酸肌醇 3,4,5-三磷酸（PIP(3)）水平，肿瘤抑制因子 PTEN 不仅控制细胞生长，还维持细胞有丝分裂和趋化所必需的细胞极性。为了确定将 PIP(3)与其细胞骨架联系起来的关键靶标，我们删除了次要基因以逆转 Dictyostelium 中 pten-细胞的缺陷。pten-细胞的极性缺陷与 PKB 底物的磷酸化升高相关。AKT 同源物 PkbA 或其激活剂 TORC2 的一个亚基的缺失，降低了磷酸化水平，并抑制了 pten-细胞的有丝分裂和趋化缺陷。在这些双突变体中，过多的 PIP(3)水平和，推测，其他 PIP(3)结合蛋白的激活对细胞骨架几乎没有或没有影响。在 pten-细胞中磷酸化增加的条带中，我们发现了 PKB 底物、PI5K、GefS、GacG 和 PakA。在 pten-细胞中破坏 PakA 可使大量细胞恢复正常行为。一致地，在 pten-细胞中表达磷酸化模拟的 PakA 会加剧缺陷，但非磷酸化的 PakA 没有影响。因此，在许多假定的 PTEN 和 PIP(3)依赖性事件中，PKB 底物的磷酸化是细胞极性的关键下游调节剂。
