Novel Regulators of CXCL13 in Human CD4+ T Cells
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CitationBenque, Isaac. 2020. Novel Regulators of CXCL13 in Human CD4+ T Cells. Master's thesis, Harvard Medical School.
AbstractWithin chronically inflamed tissues, pathogenic T cell-B cell interactions can occur outside of secondary lymphoid organs to elicit the production of antibodies. The structure of these lymphoid aggregates depends on intercellular interactions and the production of crucial chemokines, including CXCL13. CXCL13 is essential in recruiting B cells to these aggregates as it binds CXCR5 on B cells and initiates chemotaxis. Patients with rheumatoid arthritis (RA) may develop T and B cell aggregates capable of generating antibodies locally. CXCL13 levels in RA synovium are associated with T-B cell aggregates, autoantibody production, and worsened disease. The source of CXCL13 in RA synovium is an expanded subset of T cells, termed T peripheral helper (Tph) cells. In human and primates, T follicular helper (Tfh) and Tph cells appear to be the dominant source of CXCL13; however, little is known about the extrinsic signals or the transcriptional regulators that induce T cells to produce CXCL13. In this project we have used CRISPR/Cas9 gene editing of primary human CD4+ T cells to interrogate the roles of selected candidate regulators in altering CXCL13 production by human T cells.
Using this approach, deletion of BCL6, a critical transcriptional regulator of Tfh cells, inhibited the production of CXCL13 by human blood CD4+ T cells from multiple donors. In additional, deletion of the aryl hydrocarbon receptor (AHR), a ligand gated transcription factor upregulated in Tph and Tfh cells, increased T cell production of CXCL13, demonstrating that AHR functions as a negative regulator of CXCL13 production. Deletion of the transcription factor POU2F1, a factor highly upregulated in both ex vivo and in vitro differentiated Tph cells, did not alter CXCL13 production, suggesting that POU2F1 is not an essential mediator of CXCL13 production. Attempts to delete the additional candidates SOX4, a factor reported to regulate CXCL13 production, and RNF19A, an E3 ubiquitin ligase highly expressed in synovial tissue Tph cells, have not yet successfully altered target protein expression despite attempts with more than 5 guide RNAs for each target.
Together, the results present AHR as a new and robust negative regulator of CXCL13 and confirm BCL6 as a positive regulator of CXCL13 in CD4+ T cells. Understanding how these regulators affect CXCL13 production in CD4+ T cells may prove crucial in developing strategies to mitigate the formation of pathogenic T-B aggregates in autoimmune disease.
Citable link to this pagehttps://nrs.harvard.edu/URN-3:HUL.INSTREPOS:37365274