Abstract Body (Do not enter title and authors here): Background:
Upon cardiac injury, cardiac fibroblasts (CFB) become activated and differentiate into myofibroblasts to maintain cardiac structure and function through secretion of stabilizing extracellular matrix (ECM) proteins. However, sustained CFB activation and ECM secretion leads to myocardial stiffening and heart failure (HF). The unfolded protein response (UPR) responds to increased demands for folding nascent ECM proteins (e.g. collagen) trafficked through the endoplasmic reticulum. IRE1α, the most evolutionarily conserved arm of the UPR, acts adaptively both by enhancing folding of secreted proteins and degrading neighboring mRNA transcripts thereby decreasing the protein folding burden as would be expected to occur during cardiac fibrosis.

Hypothesis:
IRE1α inhibits CFB activation to protect against pathological fibrotic remodeling and HF.

Methods: Primary CFB were cultured with transforming growth factor beta (TGFβ) to promote activation, in vitro. Selective small molecule activators or inhibitors were utilized to modulate IRE1α activity. IRE1α-floxed mice were injected with AAV9 to knockout IRE1α specifically in myofibroblasts (IRE1mfbKO) and were subjected to transverse aortic constriction (TAC) as a model of HF and CFB activation, in vivo.

Results:
While increased IRE1α activity decreased CFB activation in response to TGFβ, specific inhibition of the endonuclease domain of IRE1α increased CFB activation implicating IRE1α as a potential regulator of pro-fibrotic transcript stability and degradation. Transcriptomics of CFBs identified Tmem100, a regulator of calcium channel activity, as a putative target of IRE1α and critical for CFB activation in response to TGFβ. Mechanistically, IRE1α-mediated degradation of Tmem100 prevented CFB activation in a calcineurin-dependent manner. Finally, in response to TAC, IRE1mfbKO mice had increased cardiac dysfunction and fibrotic remodeling that was coordinate with increased Tmem100 expression.

Conclusions:
IRE1α protects against cardiac fibrosis via degradation of a novel subset of pro-fibrotic transcripts, namely, Tmem100. Thus, small molecule strategies to increase IRE1α activation represent a novel approach to mitigating cardiac fibrosis. Future directions include determining the mechanism by which Tmem100 regulates CFB calcium channel activity and subsequent activation of the calcineurin-NFAT pathway leading to sustained CFB activation and differentiation.