Investigating the Role of SWI/SNF Chromatin Remodeling Component, ARID2, in Normal and Malignant B Cells
Authors: Paolini Rachel L , Souroullas George P 
Published: December 2024

Introduction:B cell lymphopoiesis is a tightly regulated, stepwise process that produces cells capable of secreting antibodies in response to infection. Epigenetic modifiers play a critical role in this system by orchestrating the timing of processes like proliferation, differentiation, and apoptosis to promote proper cell type function. Dysregulation of these pathways in B cell progenitors can give rise to B cell acute lymphoblastic leukemia (B-ALL), the most common childhood cancer worldwide. The SWI/SNF chromatin remodeling subunit, ARID2, is highly expressed in B cell progenitors and homozygous deletions in this gene are common in B-ALL patients. Moreover, ARID2 regulates differentiation of other hematopoietic lineages and is a well-documented tumor suppressor in multiple cancers. In fact, ARID2 mutations are thought to be early, initiating events in cancer. Nonetheless, little is known about the function of ARID2 in B cells or within the context of B cell malignancy. Taken together, we hypothesized that ARID2 regulates early B cell differentiation and the loss of ARID2 expression may be an early, initiating event during the progression of B cell leukemia that predisposes cells to malignant transformation.

Methods: We generated Arid2F/F Mb1-Cre and Arid2F/F CD19-Cre knockout mice to assess the effect of Arid2 loss on B cell development in comparison to Cre+ controls. The recombination efficiency within B cell progenitors is significantly higher in the Mb1-Cre system, so we expected that any resulting phenotypes would be more drastic compared to the CD19-Cre mice. To confirm the expression of Arid2 in B cell progenitors at a protein level, we isolated bone marrow (BM) from the Mb1-Cre mice, enriched for B cells using CD19 magnetic beads, and performed a western blot for Arid2. We utilized flow cytometry to analyze the frequency of progenitor B cell populations in the BM as well as more mature B cells in the spleen (SP) and peripheral blood (PB). Additionally, we immunized the Mb1-Cre mice with sheep red blood cells (SRBCs) and analyzed changes in the production of germinal center B cells in the SP. Finally, we used fluorescence-activated cell sorting (FACS) to isolate progenitor B cells from the Mb1-Cre mice and performed RNA-sequencing to compare transcriptomic changes caused by Arid2 deletion.

Results: The western blot from the BM confirmed that Arid2 is highly expressed at the protein level in CD19+ B cells and the number of CD19+ B cells is significantly reduced in the Arid2 knockout mice. The Mb1-Cre flow cytometry data supports this result as the frequency of follicular B cells in the BM, SP, and PB are significantly reduced in the Arid2 KO mice. Follicular B cells are a type of naïve B cell that is responsible for starting the germinal center reaction in response to infection. Consistently, the frequency of germinal center B cells decreases in the Arid2 KO Mb1-Cre mice after SRBC immunization. Importantly, the frequency of follicular B cells is not significantly different in the Arid2 KO CD19-Cre mice, indicating that the phenotype is driven by the loss of Arid2 in the B cell progenitors. The RNA-seq analysis revealed multiple genes related to B cell differentiation and migration that were differentially expressed in the Arid2 KO progenitors as well as the upregulation of multiple Fos and Jun genes. Fos and Jun proteins interact to form the AP-1 transcription factors which are known to regulate proliferation and induce malignant transformation in other cell types.

Conclusions and Future Directions: Altogether, we show that the loss of Arid2 expression in B cell progenitors causes a significant reduction in the number of B cells in the BM and impacts the ability of progenitor B cells to differentiate into mature B cells. The RNA-seq data is consistent with these observations and suggests that the ability of the B cells to migrate from the BM might also contribute to this reduction in B cell output. The upregulation of AP-1 transcription factor genes is consistent with our hypothesis about malignant transformation, but to test this more directly, we are overexpressing relevant B-ALL oncogenes (i.e., Dux4-IgH) in the Arid2 Mb1-Cre model to determine if loss of Arid2 accelerates leukemia development. Understanding the mechanisms by which Arid2 regulates B cell differentiation and investigating how dysregulation contributes to leukemia development could reveal new therapeutic targets for B-ALL patients.