Kinases play a critical role in the development of heart failure (HF) and, therefore, represent a potential therapeutic target. While the cardiac kinome is comprised of several hundred kinases, the vast majority of the literature is focused on only a few kinases. Indeed, the function of numerous highly expressed cardiac kinases is unexplored; these may include potential therapeutic targets for cardiovascular diseases. In order to identify novel cardiac kinase(s) potentially involved in HF development, we employed an integrated transcriptome and bioinformatics approach (Expression2Kinases (X2K)) with control and failing hearts. Indeed, in our screening results, most of the identified kinases are well-known for their roles in cardiac biology, validating the applied approach. Intriguingly, we also identified a novel kinase, Homeodomain-Interacting Protein Kinase 2 (HIPK2), as potentially being involved in the regulation of cardiac remodeling and HF. Indeed, there is no literature describing the function of HIPK2 in the heart. The overall goal of this proposal is to define the role of HIPK2 in cardiac pathophysiology. As a first step towards this goal, we have generated mouse models including fibroblast (FB)-specific HIPK2 KOs (periostinMCM and TCF21MCM) and cardiomyocyte (CM)-specific HIPK2 KOs (αMHCCre and αMHCMCM). Three specific aims are designed to critically examine the role of HIPK2 in cardiac biology. Aim 1: to define the role of HIPK2 in cardiac fibrosis and remodeling. We will test the hypothesis that HIPK2 exerts a critical break on fibrotic remodeling by inhibiting profibrotic TGF-β1/SMAD-3 and p38 pathways. In Aim 2, we will test the hypothesis that CM-HIPK2 is essential in maintaining basal cardiac homeostasis and required to protect against cardiac stress. We will utilize inducible CM-specific cre mice to delete HIPK2 only in the fully mature CM, evaluate the mechanism(s) whereby HIPK2 may exert its cardioprotective effect. Aim 3 will assess the therapeutic efficacy of AAV9 mediated HIPK2 gene therapy to mitigate the pathologies induced by cardiac stress. Expression of HIPK2 is significantly reduced in failing human hearts, we hypothesize that restoration of HIPK2 in cardiac cells will reduce and may reverse the cardiac pathologies. AAV9-HIPK2 will be used to deliver the FB-specific (under periostin promoter and TCF21 promoter) and CM-specific (troponin-t promoter) gene expression. These studies will directly test the translational potential of the findings from Aims 1 and 2. Thus, the proposed studies are of high significance, not only by contributing translationally-relevant information but also by providing novel mechanistic insights into cardiac pathophysiology.