DHCR24 knockdown-induced cellular cholesterol deficiency triggers tau hyperphosphorylation at Thr181, Ser199 and Ser202/Thr205 via p38 MAPK/JNK signaling.

Cholesterol deficiency is implicated in Alzheimer's disease (AD) pathogenesis, yet its regulatory role in tau phosphorylation remains unclear. We investigated whether the loss of 24-dehydrocholesterol reductase (DHCR24), a key enzyme in cholesterol biosynthesis, drives tau hyperphosphorylation via p38 MAPK and JNK signaling pathways in SH-SY5Y neuroblastoma cells. Using lentiviral vectors encoding DHCR24-cDNA or DHCR24-shRNA, we established stable DHCR24-overexpression and DHCR24-knockdown SH-SY5Y cell models. Filipin III staining and UPLC-MS/MS demonstrated that DHCR24 knockdown significantly reduced cellular cholesterol levels, whereas overexpression raised cholesterol levels. Immunoblotting showed selective increases in tau phosphorylation at Thr181, Ser199, and Ser202/Thr205 via DHCR24 deficiency; total tau levels remained unchanged. Concomitantly, phospho-p38 and phospho-JNK levels increased by 1.5- to 2.5-fold without alterations in total kinase levels, indicating pathway activation. To test causality, DHCR24-silenced cells were treated with escalating doses of the p38 inhibitor SB203580 (0-40 μM) or the JNK inhibitor SP600125 (0-40 μM). Both compounds restored tau phosphorylation to baseline in a concentration-dependent manner, with maximal suppression at 40 μM (SB203580) and 20-40 μM (SP600125). Collectively, these data establish DHCR24-controlled cholesterol homeostasis as a molecular rheostat of tau pathology through p38/JNK signaling, and nominate this axis for therapeutic intervention in AD.