Mitogen activated protein kinase phosphoserine/threonine/tyrosine-binding protein (MK-STYX) is a dual specificity (DUSP) member of the protein tyrosine phosphatase family. It is a pseudophosphatase, which lacks the essential amino acids histidine and cysteine in the catalytic active signature motif (HCX₅R). We previously reported that MK-STYX interacts with G3BP1 [Ras-GAP (GTPase-activating protein) SH3 (Src homology 3) domain-binding-1] and reduces stress granules, stalled mRNA. To determine how MK-STYX reduces stress granules, truncated domains, CH2 (cell division cycle 25 phosphatase homology 2) and DUSP, of MK-STYX were used. Wild-type MK-STYX and the DUSP domain significantly decreased stressed granules that were induced by sodium arsenite, in which G3BP1 (a stress granule nucleator) was used as the marker. In addition, HEK/293 and HeLa cells co-expressing G3BP1-GFP and mCherry-MK-STYX, mCherry-MK-STYX-CH2, mCherry-MK-STYX-DUSP or mCherry showed that stress granules were significantly decreased in the presence of wild-type MK-STYX and the DUSP domain of MK-STYX. Further characterization of these dynamics in HeLa cells showed that the CH2 domain increased the number of stress granules within a cell, relative to wild-type and DUSP domain of MK-STYX. To further analyze the interaction of G3BP1 and the domains of MK-STYX, coimmunoprecipitation experiments were performed. Cells co-expressing G3BP1-GFP and mCherry, mCherry-MK-STYX, mCherry-MK-STYX-CH2, or mCherry-MK-STYX-DUSP demonstrated that the DUSP domain of MK-STYX interacts with both G3BP1-GFP and endogenous G3BP1, whereas the CH2 domain of MK-STYX did not coimmunoprecipitate with G3BP1. In addition, G3BP1 tyrosine phosphorylation, which is required for stress granule formation, was decreased in the presence of wild-type MK-STYX or the DUSP domain but increased in the presence of CH2. These data highlight a model for how MK-STYX decreases G3BP1-induced stress granules. The DUSP domain of MK-STYX interacts with G3BP1 and negatively alters its tyrosine phosphorylation– decreasing stress granule formation.

丝裂原激活蛋白激酶磷酸丝氨酸/苏氨酸/酪氨酸结合蛋白 (MK-STYX) 是蛋白酪氨酸磷酸酶家族的双特异性 (DUSP) 成员。它是一种假磷酸酶，缺乏催化活性特征基序 (HCX ₅ R) 中的必需氨基酸组氨酸和半胱氨酸。我们之前报道过 MK-STYX 与 G3BP1 [Ras-GAP（GTP 酶激活蛋白）SH3（Src 同源性 3）结构域结合-1] 相互作用，并减少应激颗粒、停滞 mRNA。为了确定 MK-STYX 如何减少应激颗粒，使用了 MK-STYX 的截短结构域、CH2（细胞分裂周期 25 磷酸酶同源性 2）和 DUSP。野生型MK-STYX和DUSP结构域显着减少了亚砷酸钠诱导的应激颗粒，其中G3BP1（应激颗粒成核剂）被用作标记。此外，共表达 G3BP1-GFP 和 mCherry-MK-STYX、mCherry-MK-STYX-CH2、mCherry-MK-STYX-DUSP 或 mCherry 的 HEK/293 和 HeLa 细胞显示，在存在野生型 MK-STYX 和 MK-STYX 的 DUSP 结构域。 HeLa 细胞中这些动态的进一步表征表明，相对于野生型和 MK-STYX 的 DUSP 结构域，CH2 结构域增加了细胞内应激颗粒的数量。为了进一步分析 G3BP1 和 MK-STYX 结构域的相互作用，进行了免疫共沉淀实验。共表达 G3BP1-GFP 和 mCherry、mCherry-MK-STYX、mCherry-MK-STYX-CH2 或 mCherry-MK-STYX-DUSP 的细胞证明 MK-STYX 的 DUSP 结构域与 G3BP1-GFP 和内源性 G3BP1 相互作用，而 MK-STYX 的 CH2 结构域不与 G3BP1 共免疫沉淀。 此外，应激颗粒形成所需的 G3BP1 酪氨酸磷酸化在野生型 MK-STYX 或 DUSP 结构域存在时减少，但在 CH2 存在时增加。这些数据突出了 MK-STYX 如何减少 G3BP1 诱导的应激颗粒的模型。 MK-STYX 的 DUSP 结构域与 G3BP1 相互作用，并负面改变其酪氨酸磷酸化，从而减少应激颗粒的形成。