DETERMINING THE FUNCTION OF THE INTERLEUKIN-1 RECEPTOR ASSOCIATED KINASE PATHWAY IN PRIMARY EFFUSION LYMPHOMA

Kaposi’s sarcoma-associated herpesvirus (KSHV) is necessary but not sufficient for the development of Primary effusion lymphoma (PEL). Alterations in cellular signaling pathways are also a characteristic of PEL development. Other B cell lymphomas have acquired an oncogenic mutation in the myeloid differentiation primary response-88 (MYD88) gene. The MYD88 L265P mutant results in the activation of the Interleukin-1 Receptor Associated Kinase (IRAK) pathway and a pro-inflammatory environment. To probe IRAK/MYD88 signaling in PEL, we employed CRISPR/Cas9 technology to generate stable deletion clones in BCBL-1Cas9 cells. To look for off-target effects, we determined the complete exome of the BCBL-1Cas9 cell line. Deletion of either MYD88, IRAK4, or IRAK1 abolished IL-1β signaling; however, we could grow stable sub-clones from each population. RNA-seq analysis of IRAK4 knockouts showed that the IRAK pathway induced cellular signals constitutively, independent of IL-1β stimulation, which was abrogated by deletion of IRAK4. Transient complementation with IRAK1 increased NF-kB activity in MYD88KO, IRAK1KO, and IRAK4KO cells even in the absence of IL-1β. We also saw that IL-10, a hallmark of PEL, is engaged with the IRAK pathway as IRAK4 knockouts reduce IL-10 levels. We surmise that, unlike B-cell receptor (BCR) signaling, MYD88/IRAK signaling is constitutively active in PEL, but that under cell culture conditions, PEL rapidly becomes independent of this pathway.Doctor of Philosoph