CRISPRi screens identify the lncRNA, LOUP , as a multifunctional locus regulating macrophage differentiation and inflammatory signaling

• 成果类型: Article
• 署名作者: Halasz, Haley; Malekos, Eric; Covarrubias, Sergio; Yitiz, Samira; Montano, Christy; Sudek, Lisa; Katzman, Sol; Liu, S. John; Horlbeck, Max A.; Namvar, Leila; Weissman, Jonathan S.; Carpenter, Susan
• 署名单位: University of California System; University of California Santa Cruz; University of California System; University of California Santa Cruz; University of California System; University of California San Francisco; University of California System; University of California San Francisco; Harvard University; Harvard University Medical Affiliates; Boston Children's Hospital; Harvard University; Massachusetts Institute of Technology (MIT); Whitehead Institute; Howard Hughes Medical Institute; Massachusetts Institute of Technology (MIT); Massachusetts Institute of Technology (MIT)
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-14424
• DOI: 10.1073/pnas.2322524121
• 发表日期: 2024-05-28
• 关键词: nf-kappa-b; long noncoding rna; super-enhancers; pu.1; activation; repression; program; cancer

摘要

Long noncoding RNAs (lncRNAs) account for the largest portion of RNA from the transcriptome, yet most of their functions remain unknown. Here, we performed two independent high - throughput CRISPRi screens to understand the role of lncRNAs monocyte function and differentiation. The first was a reporter - based screen to identify lncRNAs that regulate TLR4 - NFkB signaling in human monocytes and the second screen identified lncRNAs involved in monocyte to macrophage differentiation. We successfully identified numerous noncoding and protein - coding genes that can positively or negatively regulate inflammation and differentiation. To understand the functional roles of lncRNAs in both processes, we chose to further study the lncRNA LOUP [lncRNA originating from upstream regulatory element of SPI1 (also known as PU.1)], as it emerged as a top hit in both screens. Not only does LOUP regulate its neighboring gene, the myeloid fate-determining factor SPI1 , thereby affecting monocyte macrophage differentiation, but knockdown of LOUP leads to a broad upregulation NFkB - targeted genes at baseline and upon TLR4 - NFkB activation. LOUP also harbors three small open reading frames capable of being translated and are responsible for LOUP 's ability to negatively regulate TLR4/NFkB signaling. This work emphasizes the value of high - throughput screening to rapidly identify functional lncRNAs in the innate immune system.