Kidney fibrosis is a common process that leads to the progression of various types of kidney disease. We used an integrated computational and experimental systems biology approach to identify protein kinases that regulate gene expression changes in the kidneys of human immunodeficiency virus (HIV) transgenic mice (Tg26 mice), which have both tubulointerstitial fibrosis and glomerulosclerosis. We identified homeo-domain interacting protein kinase 2 (HIPK2) as a key regulator of kidney fibrosis. HIPK2 was upregulated in the kidneys of Tg26 mice and in those of patients with various kidney diseases. HIV infection increased the protein concentrations of HIPK2 by promoting oxidative stress, which inhibited the seven in absentia homolog 1 (SIAH1)-mediated proteasomal degradation of HIPK2. HIPK2 induced apoptosis and the expression of epithelial-to-mesenchymal transition markers in kidney epithelial cells by activating the p53, transforming growth factor β (TGF-β)–SMAD family member 3 (Smad3) and Wnt-Notch pathways. Knockout of HIPK2 improved renal function and attenuated proteinuria and kidney fibrosis in Tg26 mice, as well as in other murine models of kidney fibrosis. We therefore conclude that HIPK2 is a potential target for anti-fibrosis therapy.