Splice-specificlynknockout mice reveal a dominant function of LynB in preventing autoimmunity

Abstract

The unique roles of the Src-family kinases LynA and LynB in immune activating and inhibitory signaling have eluded definition. Here we report that LynB, the shorter splice product of lyn, carries the dominant immunosuppressive function. We used CRISPR/Cas9 gene editing to constrain lyn splicing and expression to a single product: LynAKO or LynBKO mice. While activities of both isoforms regulate homeostatic Lyn expression, only LynB protects against autoimmune disease. LynBKO monocytes and dendritic cells are TLR4-hyper-responsive, and TLR4 expression increases with age in LynBKO myeloid and B cells. These changes are accompanied by the development of an inflammatory disease that resembles human systemic lupus erythematosus (SLE). The interplay between LynB and TLR4 likely underlies the autoimmunity risk associated with LYN hypomorph and TLR4 hypermorph alleles.

SH3-domain mutations selectively disrupt Csk homodimerization or PTPN22 binding

“…Cells were rested 15 min at room temperature and then transferred to 10 ml antibiotic-free medium and incubated overnight. Viable cells were quantified with a ViCell counter (Beckman) 63 , 69 . Jurkat cells from our lab (gift from A. Weiss at UCSF) have tested negative for mycoplasma and been verified by STR profiling and TCR expression.…”

SH3-domain mutations selectively disrupt Csk homodimerization or PTPN22 binding

“…Cells were rested 15 min at room temperature and then transferred to 10 ml antibiotic-free medium and incubated overnight. Viable cells were quantified with a ViCell counter (Beck-man) 65,69 . Jurkat cells from our lab (gift from A. Weiss at UCSF) have tested negative for mycoplasma and been verified by STR profiling and TCR expression.…”

Role of inhibitory signaling in peripheral B cell tolerance*

“…Lyn A has a 21 amino acid insertion in the N terminal unique region that makes it susceptible to activation‐induced degradation 61,104 . By making mice that only express either the Lyn A or Lyn B form, the Freedman group found that the Lyn A form is more associated with activating signaling, while the Lyn B form is predominantly responsible for inhibitory signaling and associated B cell tolerance 105 . While it is unclear how the Lyn B form is biased toward activating inhibitory signaling, it suggests a mechanism by which initial activating signaling by Lyn is followed by more inhibitory signaling, due to a relative skewing toward the more inhibitory form.…”