X-linked Alport syndrome (XLAS) caused by X-linked COL4A5 gene mutation is a hereditary disease that affects mainly the kidney. XLAS patients, especially males whose single copy of the COL4A5 gene is disrupted, suffer from a life-threatening renal disease, the mechanism of which remains unclear. Renal fibrosis is a characteristic pathology observed in XLAS kidney tissue. However, the molecular path from COL4A5 loss-of-function to fibrotic pathology is largely unknown. On the basis of a previously established XLAS mouse model, our study revealed an activated CD44-TGFβ signaling known to strongly promote fibrosis, along with an increased level of low molecular weight hyaluronan (LMW-HA) instead of high molecular weight hyaluronan (HMW-HA), to activate CD44-dependent TGFβ signaling in XLAS renal tissues. Additionally, hyaluronan synthase 2 (HAS2), an enzyme primarily responsible for HA production, was found to be upregulated in XLAS. In particular, in vitro studies revealed that COL4A5 knockdown in human kidney-derived HEK-293 cells can upregulate HAS2 at both the RNA and protein levels. The novel contribution of our study is finding that COL4A5 deficiency may lead to HAS2 overexpression and HA accumulation to activate CD44-TGFβ signaling, thereby promoting fibrosis, possibly suggesting that HAS2 and CD44 are potential therapeutic targets for impeding renal fibrosis in XLAS.