Chen Jing, Williams Ifor R, Kutok Jeffery L, Duclos Nicole, Anastasiadou Ema, Masters Shane C, Fu Haian, Gilliland D Gary
Division of Hematology, Brigham and Women's Hospital, 75 Francis St, Boston, MA 02115, USA.
Blood. 2004 Jul 15;104(2):535-42. doi: 10.1182/blood-2004-01-0169. Epub 2004 Mar 30.
TEL-platelet-derived growth factor-beta receptor (TEL-PDGFbetaR) is expressed in chronic myelomonocytic leukemias associated with t(5;12)(q33;p13), and the fusion tyrosine kinase retains a conserved WW-like domain in the PDGFbetaR autoinhibitory juxtamembrane region. Here we report that mutation of the 2 conserved tryptophan residues of the WW-like domain has opposing effects on TELPDGFbetaR kinase activation. Alanine substitution of W593, essential for protein-protein interaction in the context of other WW domains, impaired TEL-PDGFbetaR-mediated transformation of hematopoietic cells due to inhibition of TEL-PDGFbetaR kinase activity. In contrast, alanine substitution of W566, essential for structural integrity of WW domain in other contexts, had no effect on TEL-PDGFbetaR activation and oncogenic activity. Surprisingly, however, the W566A mutation suppressed the W593A phenotype. Double mutant W566A/W593A was indistinguishable from the wild-type fusion protein with regard to kinase activity, ability to confer factor-independent growth to Ba/F3 cells, or ability to induce a myeloproliferative disease in mice. Additional mutational analysis identified other substitutions within the WW-like domain in addition to W566A that could also suppress the W593A phenotype, including mutations predicted to diminish the autoinhibitory function of the juxtamembrane region. Therefore, the WW-like domain in the context of TELPDGFbetaR may have both positive and negative regulatory roles in kinase activation.
TEL-血小板衍生生长因子β受体（TEL-PDGFβR）在与t(5;12)(q33;p13)相关的慢性粒单核细胞白血病中表达，并且该融合酪氨酸激酶在PDGFβR自身抑制性近膜区域保留了一个保守的类WW结构域。在此我们报告，类WW结构域中2个保守色氨酸残基的突变对TEL-PDGFβR激酶激活具有相反的作用。W593的丙氨酸替代，在其他WW结构域的背景下对蛋白质-蛋白质相互作用至关重要，由于抑制了TEL-PDGFβR激酶活性，损害了TEL-PDGFβR介导的造血细胞转化。相比之下，W566的丙氨酸替代，在其他背景下对WW结构域的结构完整性至关重要，对TEL-PDGFβR激活和致癌活性没有影响。然而，令人惊讶的是，W566A突变抑制了W593A表型。双突变体W566A/W593A在激酶活性、赋予Ba/F3细胞因子非依赖性生长的能力或在小鼠中诱导骨髓增殖性疾病的能力方面与野生型融合蛋白没有区别。额外的突变分析确定了除W566A之外类WW结构域内的其他替代也可以抑制W593A表型，包括预测会减弱近膜区域自身抑制功能的突变。因此，TEL-PDGFβR背景下的类WW结构域在激酶激活中可能具有正向和负向调节作用。
