The enzyme UCH-L1 is a neuro-endocrine and germinal center B-cell marker that contributes to the development and aggressive behavior of mature B-cell malignancies. While mutations in this enzyme have been associated with Parkinson's disease, relatively little is known about the molecular features associated with the biochemical activities of UCH-L1. Here we use a survival-based complementation assay and site-directed mutagenesis and identify a novel role for the C-terminus of UCH-L1 in supporting cell survival. The C220 residue is required for UCH-L1 to promote the assembly of mTOR complex 2 and phosphorylation of the pro-survival kinase AKT. While this residue was previously described as a potential farnesylation site, destruction of the putative CAAX motif by adding a C-terminal epitope tag did not interfere with cell survival, indicating an alternate mechanism. We used proximity-based proteomics comparing the proteomes of wild-type and C220S UCH-L1 and identified a selective loss of association with RNA-binding proteins including components of the translation initiation machinery. As a consequence, the C220S mutant did not promote the assembly of the eIF4F complex. These data identify a novel role for the C-terminus of UCH-L1 in supporting pro-survival and metabolic activities in malignant B-cells. This finding may lead to the development of therapeutics with selective activity towards malignancy that potentially avoid neuronal toxicities.

中文翻译：

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UCH-L1 C末端附近的半胱氨酸对于催化活性是必不可少的，但对于促进AKT磷酸化，eIF4F装配和恶性B细胞存活是必需的。

UCH-L1酶是神经内分泌和生发中心B细胞标志物，有助于成熟B细胞恶性肿瘤的发生和侵袭行为。虽然这种酶的突变与帕金森氏病有关，但与UCH-L1的生化活性有关的分子特征知之甚少。在这里，我们使用基于生存的互补测定和定点诱变，并确定UCH-L1的C末端在支持细胞存活中的新作用。UCH-L1需要C220残基来促进mTOR复合物2的组装和促生存激酶AKT的磷酸化。虽然此残基先前曾被描述为潜在的法尼基化位点，但通过添加C端表位标签破坏推定的CAAX基序不会干扰细胞存活，指示备用机制。我们使用基于接近度的蛋白质组学，比较了野生型和C220S UCH-L1的蛋白质组，并确定了与RNA结合蛋白（包括翻译起始机制的组成部分）的结合的选择性丧失。结果，C220S突变体不促进eIF4F复合物的装配。这些数据确定了UCH-L1 C端在支持恶性B细胞的生存和代谢活动中的新作用。这一发现可能导致具有针对恶性肿瘤的选择性活性的治疗剂的开发，从而可能避免神经元毒性。我们使用基于接近度的蛋白质组学，比较了野生型和C220S UCH-L1的蛋白质组，并确定了与RNA结合蛋白（包括翻译起始机制的组成部分）的结合的选择性丧失。结果，C220S突变体不促进eIF4F复合物的装配。这些数据确定了UCH-L1 C端在支持恶性B细胞的生存和代谢活动中的新作用。这一发现可能导致具有针对恶性肿瘤的选择性活性的治疗剂的开发，从而有可能避免神经元毒性。我们使用基于接近度的蛋白质组学，比较了野生型和C220S UCH-L1的蛋白质组，并确定了与RNA结合蛋白（包括翻译起始机制的组成部分）的结合的选择性丧失。结果，C220S突变体不促进eIF4F复合物的装配。这些数据确定了UCH-L1 C端在支持恶性B细胞的生存和代谢活动中的新作用。这一发现可能导致具有针对恶性肿瘤的选择性活性的治疗剂的开发，从而可能避免神经元毒性。