Cancer Research Group, University of Saskatchewan, 107 Wiggins Road, Saskatoon, Saskatchewan, S7N 5E5, Canada.
Department of Biochemistry, University of Saskatchewan, 107 Wiggins Road, Saskatoon, Saskatchewan, S7N 5E5, Canada.
Sci Rep. 2018 May 8;8(1):7108. doi: 10.1038/s41598-018-25487-5.
The p85α protein regulates flux through the PI3K/PTEN signaling pathway, and also controls receptor trafficking via regulation of Rab-family GTPases. In this report, we determined the impact of several cancer patient-derived p85α mutations located within the N-terminal domains of p85α previously shown to bind PTEN and Rab5, and regulate their respective functions. One p85α mutation, L30F, significantly reduced the steady state binding to PTEN, yet enhanced the stimulation of PTEN lipid phosphatase activity. Three other p85α mutations (E137K, K288Q, E297K) also altered the regulation of PTEN catalytic activity. In contrast, many p85α mutations reduced the binding to Rab5 (L30F, I69L, I82F, I177N, E217K), and several impacted the GAP activity of p85α towards Rab5 (E137K, I177N, E217K, E297K). We determined the crystal structure of several of these p85α BH domain mutants (E137K, E217K, R262T E297K) for bovine p85α BH and found that the mutations did not alter the overall domain structure. Thus, several p85α mutations found in human cancers may deregulate PTEN and/or Rab5 regulated pathways to contribute to oncogenesis. We also engineered several experimental mutations within the p85α BH domain and identified L191 and V263 as important for both binding and regulation of Rab5 activity.
p85α 蛋白调节 PI3K/PTEN 信号通路的通量，还通过调节 Rab 家族 GTPases 控制受体运输。在本报告中，我们测定了先前显示与 PTEN 和 Rab5 结合并调节其各自功能的 p85α N 端结构域中的几种癌症患者衍生的 p85α 突变对的影响。p85α 突变体 L30F 显著降低与 PTEN 的稳态结合，但增强了对 PTEN 脂质磷酸酶活性的刺激。另外 3 种 p85α 突变（E137K、K288Q、E297K）也改变了对 PTEN 催化活性的调节。相比之下，许多 p85α 突变降低了与 Rab5 的结合（L30F、I69L、I82F、I177N、E217K），并且几种突变影响 p85α 对 Rab5 的 GAP 活性（E137K、I177N、E217K、E297K）。我们为牛 p85α BH 测定了这些 p85α BH 结构域突变体（E137K、E217K、R262T、E297K）中的几个的晶体结构，发现突变并未改变整体结构域结构。因此，在人类癌症中发现的几种 p85α 突变可能使 PTEN 和/或 Rab5 调节途径失活，从而促进肿瘤发生。我们还在 p85α BH 结构域内设计了几种实验性突变，并确定 L191 和 V263 对于结合和调节 Rab5 活性都很重要。
