Koul Dimpy, Jasser Samar A, Lu Yiling, Davies Michael A, Shen Ruijun, Shi Yuexi, Mills Gordon B, Yung W K Alfred
Department of Neuro-Oncology, Box 100, The Brain Tumor Center, The University of Texas, M.D. Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, Texas, TX 77030, USA.
Oncogene. 2002 Apr 4;21(15):2357-64. doi: 10.1038/sj.onc.1205296.
The tumor suppressor gene, MMAC/PTEN, has phosphatase, C2, and PDZ-binding domains as well as potential sites of regulation by phosphorylation, including tyrosine phosphorylation, which may contribute to its ability to modulate cell growth and viability. Several obvious and significant motifs were found in MMAC/PTEN, including most notably, a catalytic domain of tyrosine phosphatase (IHCxxGxxRS/T) and several potential tyrosine phosphorylation sites. To examine the functional significance of tyrosine phosphorylation of MMAC/PTEN, retroviral constructs were generated with mutations at two putative tyrosine phosphorylation sites (Y240A/Y240F and Y315A/Y315F). Stable expression of wild-type MMAC/PTEN in U251 human glioma cells (which do not normally produce a functional MMAC/PTEN gene product) resulted in a significant reduction of tumor growth in nude mice, decreased growth rate, saturation density, and colony formation in vitro, as well as dephosphorylation of D3-phosphorylated phosphatidylinositols (PtdIns) in vitro. Mutation of Y240 or Y315 to either alanine or phenylalanine abrogated the ability of MMAC/PTEN to alter growth rate, saturation density, and colony formation in vitro. The ability of MMAC/PTEN to limit tumor growth in nude mice was markedly decreased but not abrogated by mutation of Y240 or Y315 to alanine. Thus, Y240 and Y315 are required for MMAC/PTEN to decrease tumor growth in vitro and in vivo. In contrast to wild-type MMAC/PTEN, mutant MMAC/PTEN containing Y240A or Y315A was unable to dephosphorylate D3-phosphorylated PtdIns in vitro. Thus, Y240A and Y315A are involved in the ability of MMAC/PTEN to dephosphorylate PtdIns and regulate tumor cell growth in vitro and in vivo.
肿瘤抑制基因MMAC/PTEN具有磷酸酶、C2和PDZ结合结构域，以及包括酪氨酸磷酸化在内的潜在磷酸化调控位点，这可能有助于其调节细胞生长和活力的能力。在MMAC/PTEN中发现了几个明显且重要的基序，其中最显著的是酪氨酸磷酸酶催化结构域（IHCxxGxxRS/T）和几个潜在的酪氨酸磷酸化位点。为了研究MMAC/PTEN酪氨酸磷酸化的功能意义，构建了在两个假定的酪氨酸磷酸化位点（Y240A/Y240F和Y315A/Y315F）发生突变的逆转录病毒构建体。野生型MMAC/PTEN在U251人胶质瘤细胞（通常不产生功能性MMAC/PTEN基因产物）中的稳定表达导致裸鼠肿瘤生长显著减少，体外生长速率、饱和密度和集落形成降低，以及体外D3 - 磷酸化磷脂酰肌醇（PtdIns）的去磷酸化。将Y240或Y315突变为丙氨酸或苯丙氨酸消除了MMAC/PTEN改变体外生长速率、饱和密度和集落形成的能力。MMAC/PTEN限制裸鼠肿瘤生长的能力明显降低，但将Y240或Y315突变为丙氨酸并未消除该能力。因此，Y240和Y315是MMAC/PTEN在体外和体内降低肿瘤生长所必需的。与野生型MMAC/PTEN相比，含有Y240A或Y315A的突变型MMAC/PTEN在体外无法使D3 - 磷酸化PtdIns去磷酸化。因此，Y240A和Y315A参与了MMAC/PTEN在体外和体内使PtdIns去磷酸化并调节肿瘤细胞生长的能力。
