Brown Rachel A, Dangel Andrew W, Saini Ankita, Collins Patrick L, Colonna Marco, Oltz Eugene M
Department of Microbial Infection and Immunity, The Ohio State University, Columbus, OH, 43210, USA.
Pelotonia Institute for Immuno-Oncology, The Ohio State University, Columbus, OH 43210, USA.
bioRxiv. 2025 Aug 15:2025.08.15.670561. doi: 10.1101/2025.08.15.670561.
The cytokines interleukin (IL)-22 and IL-17 are secreted by innate and adaptive immune cells to drive "type III" responses that protect against extracellular pathogens, promote mucosal barrier integrity, and foster microbiota homeostasis. However, dysregulation of IL-22 and/or IL-17 contributes to autoimmunity, chronic inflammation, and malignancy. Thus, a deeper understanding of mechanisms regulating type III cytokine production could provide new therapeutic targets for a spectrum of immune-mediated diseases. Toward this goal, we performed a genome-wide CRISPR inhibition (CRISPRi) screen to identify factors that regulate IL-22/IL-17 expression in a murine type III innate lymphoid cell (ILC3) model, MNK3, following stimulation with IL-23 and IL-1b. In addition to previously known regulators of type III cytokines, including IL-23 receptor components IL23R and IL12RB1, the screen identified a large set of new factors that either potentiate or attenuate expression of IL-22 and/or IL-17. A subset of these novel factors was chosen for validation, from which two were selected for further study. The nuclear protein, SON, which binds both DNA and RNA, impaired expression of IL12RB1 at the levels of de novo transcription and RNA processing. The second, MAP4K1 (HPK1), is a serine/threonine kinase that is required for IL-22 but not IL-17 expression. Depletion of MAP4K1 in MNK3 also enhanced expression of the type I cytokine, IFNg, which was co-expressed with IL-17, a phenotype reminiscent of pathogenic Th17 cells. Together, results from the CRISPRi screen broaden our understanding of the factors involved in type III immune responses and offer new targets for modulating IL-22/17 expression.
细胞因子白细胞介素（IL）-22和IL-17由先天性和适应性免疫细胞分泌，以驱动“III型”反应，这些反应可抵御细胞外病原体、促进粘膜屏障完整性并维持微生物群稳态。然而，IL-22和/或IL-17的失调会导致自身免疫、慢性炎症和恶性肿瘤。因此，更深入地了解调节III型细胞因子产生的机制可为一系列免疫介导的疾病提供新的治疗靶点。为了实现这一目标，我们进行了全基因组CRISPR抑制（CRISPRi）筛选，以确定在用IL-23和IL-1β刺激后，在小鼠III型先天性淋巴细胞（ILC3）模型MNK3中调节IL-22/IL-17表达的因子。除了先前已知的III型细胞因子调节因子，包括IL-23受体成分IL23R和IL12RB1外，该筛选还鉴定出大量新的因子，它们要么增强要么减弱IL-22和/或IL-17的表达。从这些新因子中选择了一个子集进行验证，并从中选择了两个进行进一步研究。核蛋白SON既能结合DNA又能结合RNA，它在从头转录和RNA加工水平上损害了IL12RB1的表达。第二个是MAP4K1（HPK1），它是一种丝氨酸/苏氨酸激酶，是IL-22而非IL-17表达所必需的。在MNK3中耗尽MAP4K1也增强了与IL-17共表达的I型细胞因子IFNγ的表达，这种表型让人联想到致病性Th17细胞。总之，CRISPRi筛选的结果拓宽了我们对参与III型免疫反应的因子的理解，并为调节IL-22/17表达提供了新的靶点。
