Key Points B cell–specific deletion of Ship2 phosphatase reduces CLL formation in a mouse model. Ship2 inhibition reduces Akt/mTORC1/S6 signaling in murine and human CLL cells. Ship2 supports survival of CLL cells via the key antiapoptotic protein Mcl-1. Balanced activity of kinases and phosphatases downstream of the BCR is essential for B cell differentiation and function and is disturbed in chronic lymphocytic leukemia (CLL). In this study, we employed IgH.TEμ mice, which spontaneously develop CLL, and stable EMC CLL cell lines derived from these mice to explore the role of phosphatases in CLL. Genome-wide expression profiling comparing IgH.TEμ CLL cells with wild-type splenic B cells identified 96 differentially expressed phosphatase genes, including SH2-containing inositol phosphatase (Ship2). We found that B cell–specific deletion of Ship2, but not of its close homolog Ship1, significantly reduced CLL formation in IgH.TEμ mice. Treatment of EMC cell lines with Ship1/2 small molecule inhibitors resulted in the induction of caspase-dependent apoptosis. Using flow cytometry and Western blot analysis, we observed that blocking Ship1/2 abrogated EMC cell survival by exerting dual effects on the BCR signaling cascade. On one hand, specific Ship1 inhibition enhanced calcium signaling and thereby abrogated an anergic response to BCR stimulation in CLL cells. On the other hand, concomitant Ship1/Ship2 inhibition or specific Ship2 inhibition reduced constitutive activation of the mTORC1/ribosomal protein S6 pathway and downregulated constitutive expression of the antiapoptotic protein Mcl-1, in both EMC cell lines and primary IgH.TEμ CLL cells. Importantly, also in human CLL, we found overexpression of many phosphatases including SHIP2. Inhibition of SHIP1/SHIP2 reduced cellular survival and S6 phosphorylation and enhanced basal calcium levels in human CLL cells. Taken together, we provide evidence that SHIP2 contributes to CLL pathogenesis in mouse and human CLL.

中文翻译：

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含有 SH2 的肌醇磷酸酶的过度表达有助于慢性淋巴细胞白血病的存活

关键点 Ship2 磷酸酶的 B 细胞特异性缺失减少了小鼠模型中 CLL 的形成。Ship2 抑制减少了小鼠和人类 CLL 细胞中的 Akt/mTORC1/S6 信号传导。Ship2 通过关键的抗凋亡蛋白 Mcl-1 支持 CLL 细胞的存活。BCR 下游激酶和磷酸酶的平衡活性对于 B 细胞分化和功能至关重要，并且在慢性淋巴细胞白血病 (CLL) 中受到干扰。在本研究中，我们使用自发发展 CLL 的 IgH.TEμ 小鼠和源自这些小鼠的稳定 EMC CLL 细胞系来探索磷酸酶在 CLL 中的作用。比较 IgH.TEμ CLL 细胞与野生型脾 B 细胞的全基因组表达谱鉴定了 96 个差异表达的磷酸酶基因，包括含有 SH2 的肌醇磷酸酶 (Ship2)。我们发现 Ship2 的 B 细胞特异性缺失，但不是其密切同源物 Ship1，显着降低了 IgH.TEμ 小鼠中 CLL 的形成。用 Ship1/2 小分子抑制剂处理 EMC 细胞系导致诱导半胱天冬酶依赖性细胞凋亡。使用流式细胞术和蛋白质印迹分析，我们观察到阻断 Ship1/2 通过对 BCR 信号级联产生双重影响来消除 EMC 细胞存活。一方面，特定的 Ship1 抑制增强了钙信号，从而消除了 CLL 细胞对 BCR 刺激的无反应性反应。另一方面，在 EMC 细胞系和原代 IgH.TEμ CLL 细胞中，伴随的 Ship1/Ship2 抑制或特定的 Ship2 抑制减少了 mTORC1/核糖体蛋白 S6 通路的组成型激活和抗凋亡蛋白 Mcl-1 的组成型表达下调。重要的是，同样在人类 CLL 中，我们发现包括 SHIP2 在内的许多磷酸酶过度表达。抑制 SHIP1/SHIP2 会降低细胞存活率和 S6 磷酸化，并提高人类 CLL 细胞中的基础钙水平。总之，我们提供证据表明 SHIP2 有助于小鼠和人类 CLL 的 CLL 发病机制。