Total, not isoform-specific, Lyn expression by macrophages promotes TLR activation and restricts proliferation

Abstract Toll-like receptor (TLR) signaling is vital to antimicrobial macrophage function, and its dysregulation is associated with many disease states, including lupus, multiple sclerosis, pulmonary fibrosis, and cancer. The Src-family kinase Lyn plays activating and inhibitory roles downstream of TLRs, yet distinct functions of the Lyn splice variants LynA and LynB in TLR signaling had not been investigated. We used isoform-specific Lyn knockout mice (LynA KO and LynB KO ) to interrogate the contribution of each isoform to TLR signaling in bone marrow-derived macrophages. Bulk RNA sequencing and cytokine analyses revealed that complete Lyn deficiency (Lyn KO ) dampens TLR4- and TLR7-induced inflammatory gene expression and TNF production, but enhances the expression of genes responsible for synthesizing the extracellular matrix and promoting proliferation. Despite a reduction in total Lyn levels, the expression of either LynA or LynB alone was sufficient to preserve wild-type transcriptional responses and TNF production in response to the TLR7 agonist R848. However, LyA KO and LynB KO macrophages did have partially impaired TNF production in response to the TLR4 agonist lipopolysaccharide. Additionally, LynA KO and LynB KO macrophages were hyperproliferative, like Lyn KO cells. These data suggest that Lyn promotes macrophage activation downstream of TLRs and restrains aberrant proliferation and matrix deposition in a dose-dependent rather than isoform-specific manner. Summary Sentence RNA sequencing and functional assays demonstrate that both LynA and LynB restrict macrophage proliferation and drive TLR-induced ECM-remodeling and inflammatory cytokine production.