Many forms of heart disease are associated with the loss of cardiomyocytes (CMs) and their replacement by fibrotic tissue. Cardiac fibrosis is central to the pathology of heart failure, a deadly syndrome and a leading cause of mortality in the US. Unfortunately, there are no effective therapies to prevent the progression of cardiac fibrosis. Recent findings suggest that the way the heart responds to injury, including the extent of fibrosis, is a variable trait influenced by several genes. One of the potential regulators of this outcome is the CM-specific kinase Tnni3k.  Mutations that abolish Tnni3k expression confer resistance to injury, and high Tnni3k levels are associated with rapid functional decline and pathological remodeling. Elevated Tnni3k has also been associated with increased ploidy and reduced proliferation in the zebrafish heart after injury. Despite its multiple links to disease, Tnni3k is an understudied kinase, and its downstream targets and specific mechanisms by which it defines injury outcomes are unknown.

To identify the molecular mechanisms by which Tnni3k induces cardiac diseases, we have established several new genetic models in zebrafish, an animal model with the unparalleled ability to regenerate its heart after cryoinjury through myocardial regeneration and progressive fibrosis regression. We found that elevated expression of Tnni3k in cardiomyocytes is sufficient to induce cardiomyopathy-like phenotypes in zebrafish, including fibrosis and leukocyte infiltration at baseline. While CM ploidy and proliferation after injury were unaffected, these animals developed an extensive fibrotic response that failed to regress. Conversely, we found reduced fibrosis and robust regeneration in a newly generated tnni3k mutant. Transcriptomic profiling revealed that Tnni3k induces an exacerbated inflammatory response after injury and the upregulation of several genes involved in stabilizing the scar. Using new whole locus deletions lines, we discovered that some of these responses are protective and that preventing these adaptations worsens the response to cardiac injury. Finally, modulation of Tnni3k levels using a Tnni3k-Switch allele demonstrates that decreasing the expression of this gene post-injury reduces inflammation and fibrosis, improving overall myocardial recovery. Our findings suggest that Tnni3k acts as a modifier of regenerative competence not by affecting cardiomyocyte proliferation but by inducing a pro-inflammatory and pro-fibrotic environment in the heart. Our work exemplifies the power of zebrafish genetics to serve as the springboard for the rapid discovery of new targets to treat cardiac fibrosis in the injured heart.