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bioRxiv · 10.1101/2024.03.31.587501

Assessing a Role for Sarcospan in Immune Function

Valera, I. C.·Rahimi Kahmini, A.·Smith, K. A.·Crawford, R. Q.·Parvatiyar, M. S.
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Abstract

Sarcospan (SSPN) is a tetraspanin-like member of the dystrophin-glycoprotein complex (DGC) with expression reported at the transcript level in B cells and several other immune cell populations. SSPN is best known for its expression in skeletal, cardiac, and vascular smooth muscle, however it is expressed in other cell types and its roles in non-muscle cells have not been well described. Analysis of functional interactions of SSPN with effectors of hematopoietic cell lineages uncovered strong associations with several proteins with known roles in hematopoiesis and immune response. In this study, we utilized flow cytometry to uncover SSPN+ immune cell populations expressing the SSPN protein. We performed immunophenotyping of global SSPN-deficient (SSPN-/-) C57BL/6J male and female mice to assess whether SSPN plays an essential role in generation of specific immune cells. By flow cytometry we found that B cells, CD11b+ and CD11c+ (monocytes, tissue macrophages, and a subset of dendritic cells) express the highest levels of SSPN at their cell membranes. To pinpoint potential roles for SSPN in B cell development, SSPN-/- bone marrow cells were isolated and differentiated using phorbol myristate acetate (PMA). To test B cell effector functions, antibody isotype analysis was performed to assess whether SSPN influences antibody class switching. Blood serum of WT and SSPN-/- mice were tested for presence of important developmental signals for B cell maturation, development and isotype switching. Our study is the first to document SSPN protein expression on distinct classes of immune cells including CD11b+, CD11c+ and B cells (CD19+). Immunophenotyping revealed several differences in immune cell populations in the murine spleens: 1) a significant increase in % T cells in SSPN-/- males, 2) slight reduction of % B cells in SSPN-/- females, 3) a significant increase % of dendritic cells in SSPN-/- females, and 4) no significant alterations in NK cells. Under unstimulated conditions, SSPN-/- female mice had a significantly lower IgG2b titers compared to WT. Overall, the observation of SSPN expression in distinct immune cells, potential role(s) in hematopoiesis and B cell effector function is intriguing and warrants further investigation. HighlightsThe sarcospan protein has an established role in stabilization of striated muscle membranes and is an essential member of the dystrophin-glycoprotein complex. This study investigates the reported presence of Sspn in B cells and examines potential roles in immune function including: O_LIDetermining a connection with hematopoiesis and establishment of niches C_LIO_LIImpact on immune content in spleens of sarcospan-deficient mice C_LIO_LIIdentifying a role in B cell effector functions C_LI

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Valera, I. C., Rahimi Kahmini, A., Smith, K. A., Crawford, R. Q., Parvatiyar, M. S.. 2024-04-01. Assessing a Role for Sarcospan in Immune Function. https://doi.org/10.1101/2024.03.31.587501

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