Single-nucleotide polymorphism rs71327024 located in the human 3p21.31 locus has been associated with an elevated risk of hospitalization upon SARS-CoV-2 infection. The 3p21.31 locus contains several genes encoding chemokine receptors potentially relevant to severe COVID-19. In particular, CXCR6, which is prominently expressed in T lymphocytes, NK, and NKT cells, has been shown to be involved in the recruitment of immune cells to non-lymphoid organs in chronic inflammatory and respiratory diseases. In COVID-19, CXCR6 expression is reduced in lung resident memory T cells from patients with severe disease as compared to the control cohort with moderate symptoms. We demonstrate here that rs71327024 is located within an active enhancer that augments the activity of the CXCR6 promoter in human CD4⁺ T lymphocytes. The common rs71327024(G) variant makes a functional binding site for the c-Myb transcription factor, while the risk rs71327024(T) variant disrupts c-Myb binding and reduces the enhancer activity. Concordantly, c-Myb knockdown in PMA-treated Jurkat cells negates rs71327024’s allele-specific effect on CXCR6 promoter activity. We conclude that a disrupted c-Myb binding site may decrease CXCR6 expression in T helper cells of individuals carrying the minor rs71327024(T) allele and thus may promote the progression of severe COVID-19 and other inflammatory pathologies.

Chromosomal rearrangements that lead to the formation of oncogenic gene fusions, such as EML4-ALK, are thought to arise from incorrect repair of double-strand breaks in DNA. However, the mechanisms and factors driving rearrangement formation remain poorly understood, and analysis of these processes is limited by detection methods that are labor-intensive, low-throughput, and not readily quantitative at single-cell resolution. Here, we developed a genetically encoded ALK reporter based on A549 lung adenocarcinoma cells, created by inserting an ALK-P2A-mCherry cassette into the endogenous ALK locus, so that induced EML4-ALK fusion activated mCherry fluorescence. Reporter activation yielded a readily quantifiable mCherry-positive subpopulation that could be measured and enriched by flow cytometry and correlated with EML4-ALK levels. Using this platform, we combined CRISPR-mediated rearrangement induction with knockdown of DNA repair factors using RNA interference. Of the factors involved in base excision repair, homologous recombination-related pathways and canonical non-homologous end joining, knockdown of the APEX1 gene encoding apurinic endonuclease 1 (APE1) selectively increased EML4-ALK levels both in the reporter cell line and in parental A549 cells. Together, this work provides a sensitive, single-cell A549-based ALK reporter platform and a framework for future studies aimed at identifying cellular and environmental factors that modulate oncogenic EML4-ALK rearrangement formation.

SLAMF1 encodes CD150, an immunoregulatory receptor involved in lymphocyte activation, T–B-cell interactions, and humoral immune responses. The SLAMF1 promoter polymorphism rs2295613(G>A) was previously associated with systemic lupus erythematosus (SLE) susceptibility in a Chinese case–control cohort. Here, we investigated the regulatory activity of rs2295613 in the transformed B-cell lines Raji and MP1 and in primary human CD19⁺ B cells. The rs2295613(A)-containing reporter showed higher promoter activity than the rs2295613(G)-containing reporter in all three cellular systems. Bioinformatic analysis predicted that the G to A substitution strengthens a pre-existing MYC-compatible motif. Substitutions disrupting the motif-containing region attenuated the rs2295613(A)-associated increase in reporter activity and reduced enrichment of the promoter fragment in anti-c-MYC DNA pull-down assays. Partial siRNA-mediated reduction in MYC mRNA also decreased the activity of the rs2295613(A)-containing reporter in Raji cells. Together, these findings identify rs2295613 as a functional SLAMF1 promoter variant in B-cell reporter systems and support a contribution of c-MYC-associated regulation to the enhanced activity of the rs2295613(A)-containing promoter.