高通量筛选鉴定哺乳动物细胞中抗增殖肽，揭示关键转录因子家族

A High-Throughput Screen for Antiproliferative Peptides in Mammalian Cells Identifies Key Transcription Factor Families

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ACS Synthetic Biology

影响因子:3.9

分区:生物学2区 / 生化研究方法1区

发表日期:2024 Nov 15

作者: Shane M Liila-Fogarty, Grace E Boyum, Claire L Schwabe, Gaelen T Hess

DOI: 10.1021/acssynbio.4c00337

摘要

转录因子（TFs）是多种疾病的有潜力的治疗靶点。TFs通过参与多种特异性蛋白-蛋白相互作用实现其细胞功能。例如，一些TFs的同源或异源二聚化控制DNA结合，而TFs与基础转录机械或染色质修饰因子的相互作用也可能至关重要。理论上，小分子可以用来破坏这些特定的蛋白-蛋白界面以干扰TF的功能，但实际上，鉴定具有必要特异性和效力的小分子存在困难，可能是由于TF功能所依赖的广泛蛋白-蛋白界面。然而，与小分子相比，肽具有提供所需特异性和效力以破坏此类界面的潜力。本研究利用一项高通量联合筛选，从源自人类核定位蛋白的80个氨基酸片段（肽）库中筛选出约15个能抑制白血病细胞增殖的肽。这些抗增殖肽富集于已知参与特定转录因子二聚化的区域，包括碱性亮氨酸拉链（bZIP）家族。来自转录因子JDP2的bZIP_1肽，是抗增殖效果最突出的肽，将K562细胞的增殖抑制了2倍。JDP2;bZIP_1抑制AP-1的转录活性，表现出与JDP2过表达相似的表型，提示该肽通过内源性JDP2机制影响细胞增殖。令人意外的是，尽管bZIP域高度保守，非二聚化域的残基对肽的抗增殖作用至关重要。该肽通过诱导红细胞分化并在多个细胞模型中增加G0/G1期细胞数量。同时发现，许多此类抗增殖肽（包括JDP2;bZIP_1）不依赖核定位信号即可发挥作用，这对其在治疗中的递送具有潜在益处。

Abstract

Transcription factors (TFs) are a promising therapeutic target for a multitude of diseases. TFs perform their cellular roles by participating in multiple specific protein-protein interactions. For example, homo- or heterodimerization of some TFs controls DNA binding, while interactions between TFs and components of basal transcriptional machinery or chromatin modifiers can also be critical. While, in theory, small molecules could be used to disrupt specific protein-protein interfaces required for TF function, in practice, it is difficult to identify small molecules with the necessary specificity and efficacy, likely due to the extensive protein-protein interfaces that often underlie TF function. However, in contrast to small molecules, peptides have the potential to provide both the specificity and efficacy required to disrupt such interfaces. Here, we identified ∼15 peptides that inhibit the proliferation of leukemia cells using a high-throughput pooled screen of a library of 80-mer protein regions (peptides) derived from human nuclear-localized proteins. The antiproliferative peptides were enriched for regions known to be involved in specific TF dimerization, including the basic leucine zipper (bZIP) domain family. One of these bZIP domains, JDP2;bZIP_1, from the TF JDP2, was the top antiproliferative peptide, reducing the proliferation of K562 cells by 2-fold. JDP2;bZIP_1 inhibited AP-1 transcriptional activity and phenocopied JDP2 overexpression, suggesting that the peptide affected proliferation through a native JDP2 mechanism. Unexpectedly, given the strong conservation of the bZIP domain, residues outside of the annotated dimerization domain were critical for the peptide's antiproliferative potency. The peptide-mediated antiproliferative effect initiated erythrocyte differentiation in K562 cells and increased G0/G1 cells across multiple cell line models. We also found that many of the antiproliferative peptides identified in this study, including JDP2;bZIP_1, did not require a nuclear localization signal to function, a potential benefit for delivering these peptides in therapeutic applications.