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Lyn expression in macrophages promotes TLR activation and restricts proliferation in an isoform-independent manner

ABSTRACT: Toll-like receptor (TLR) signaling is vital to macrophage function, and dysregulation is associated with many disease states. The Src-family kinase (SFK) Lyn serves activating and inhibitory roles downstream of TLRs, yet distinct functions of the isoforms LynA and LynB in TLR signaling have not been investigated. We used isoform-specific knockout mice (LynAKO and LynBKO) to interrogate the contribution of each isoform to TLR signaling in bone marrow-derived macrophages (BMDMs). Bulk RNA sequencing and cytokine analyses revealed that complete Lyn deficiency (LynKO) dampens TLR4- and TLR7-induced inflammatory gene expression and TNF production, but enhances extracellular matrix (ECM) gene expression and promotes cell-cycle progression and macrophage proliferation. In contrast, expression of either LynA or LynB was sufficient to preserve wild-type (WT) transcriptional responses and TNF production, despite moderately reduced total Lyn levels. These data suggest that Lyn promotes macrophage activation downstream of TLRs and restrains aberrant proliferation and matrix deposition in a largely dose-dependent rather than isoform-specific manner. Our findings provide new insight into the roles of LynA and LynB in TLR signaling and can inform future studies of macrophage-driven pa-thology in autoimmune disease, fibrosis, and cancer.

ORGANISM(S): Mus musculus

PROVIDER: GSE300925 | GEO | 2025/11/18

REPOSITORIES: GEO

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Project description:Rationale: Monocytes are key effectors of the mononuclear phagocyte system (MPS), playing critical roles in regulating tissue homeostasis and coordinating inflammatory reactions, including those involved in chronic inflammatory diseases such as atherosclerosis. Monocytes have traditionally been divided into two major subsets termed conventional (cMo) and patrolling (pMo) monocytes but recent systems immunology approaches have identified marked heterogeneity within these cells, and much of what regulates monocyte population homeostasis remains unknown. We and others have previously identified LYN tyrosine kinase as a key negative regulator of myeloid cell biology, however LYN’s role in regulating specific monocyte subset homeostasis has not been investigated. Objective: We sought to comprehensively profile monocytes in order to elucidate the underlying heterogeneity within monocytes and dissect how Lyn deficiency affects monocyte subset composition, signaling, and gene expression. We further tested the biological significance of these findings in a model of atherosclerosis. Methods and Results: CyTOF analysis of monocyte subsets and signaling pathway activation patterns in cMos and pMos revealed distinct baseline signaling profiles and far greater heterogeneity than previously described. Lyn deficiency led to a selective expansion of pMos and alterations in specific signaling pathways within these cells, revealing a critical role for LYN in pMo physiology. LYN’s role in regulating pMos was cell-intrinsic and correlated with an increased circulating half-life of Lyn-deficient pMos. Furthermore, single cell RNA sequencing revealed marked perturbations in the gene expression profiles of Lyn-/- monocytes with upregulation of genes involved in pMo development, survival, and function. Lyn deficiency also led to a significant increase in aorta-associated pMos and protected Ldlr-/- mice from high-fat diet induced atherosclerosis. Conclusions: Together our data identify LYN as a key regulator of pMo development and a potential therapeutic target in inflammatory diseases regulated by pMos.

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Lyn expression in macrophages promotes TLR activation and restricts proliferation in an isoform-independent manner

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