Exportin-5 Links miRNA Biogenesis to the Preservation of B Cell Receptor Surface Expression 2310271

Abstract

The B cell receptor (BCR) dictates B lymphocyte identity and function, allowing for a robust adaptive immune response through antigen recognition. Strict regulation of BCR surface density dictates proper B cell signaling, immune regulation, and the prevention of malignancy, yet the factors regulating this density remain undefined. To systematically identify these regulators, we performed a genome-wide CRISPR interference (CRISPRi) screen in Ramos B cells, a human non-Hodgkin’s lymphoma cell line that undergoes constitutive somatic hypermutation (SHM) and identified Exportin-5 (XPO5) as a central regulator of BCR surface expression. We transduced dCas9-KRAB Ramos cells with a genome-wide guide RNA library and isolated BCR-positive and -negative populations by flow cytometry. Hits were validated with individual guide RNAs. Mechanisms were investigated using bulk and small RNA sequencing, and BCR mutation frequencies were quantified via the TRUST4 pipeline. XPO5 deficient cells exhibited an accelerated loss of surface BCR with no change in transcript levels, suggesting a potential post-transcriptional regulatory mechanism. Further analysis revealed XPO5 depletion led to increased mutation rates specifically within the BCR light chain, implicating XPO5 in regulating the rate and fidelity of somatic hypermutation (SHM). Transcriptomic and small RNA sequencing revealed a global reduction in miRNA levels and enrichment of target gene sets indicative of cell cycle arrest and increased DNA damage response. We identify XPO5 as a multi-faceted regulator of B cell identity. By maintaining the global miRNA landscape, XPO5 supports cell cycle progression and regulates the fidelity of somatic hypermutation. Loss of XPO5 uncouples these processes, driving light chain biased hypermutation and the loss of functional BCR surface expression. This highlights a novel link between miRNA nuclear export machinery and the preservation of BCR integrity. n/a Immune Response Regulation: Molecular Mechanisms (IRM)