Catalyzes the formation of 25-hydroxycholesterol from cholesterol, leading to repress cholesterol biosynthetic enzymes (PubMed:11967195, PubMed:11207193). Plays a key role in cell positioning and movement in lymphoid tissues: 25-hydroxycholesterol is an intermediate in biosynthesis of 7-alpha,25-dihydroxycholesterol (7-alpha,25-OHC), an oxysterol that acts as a ligand for the G protein-coupled receptor GPR183/EBI2, a chemotactic receptor for a number of lymphoid cells. May play an important role in regulating lipid metabolism by synthesizing a corepressor that blocks sterol regulatory element binding protein (SREBP) processing. In testis, production of 25-hydroxycholesterol by macrophages plays a role in Leydig cell differentiation (PubMed:11967195, PubMed:11207193). Interferon-stimulated gene which has broad antiviral activities against a wide range of enveloped viruses (By similarity). Catalyzes the formation of 25-hydroxycholesterol from cholesterol, leading to repress cholesterol biosynthetic enzymes (By similarity). Plays a key role in cell positioning and movement in lymphoid tissues: 25-hydroxycholesterol is an intermediate in biosynthesis of 7-alpha,25-dihydroxycholesterol (7-alpha,25-OHC), an oxysterol that acts as a ligand for the G protein-coupled receptor GPR183/EBI2, a chemotactic receptor for a number of lymphoid cells (By similarity). May play an important role in regulating lipid metabolism by synthesizing a corepressor that blocks sterol regulatory element binding protein (SREBP) processing. As an interferon-stimulated gene, has broad antiviral activities against a wide range of enveloped viruses. Its product, 25-hydroxycholesterol, activates the ER-localized enzyme ACAT to induce internalization of accessible cholesterol on the plasma membrane and restricts virus-host membranes fusion which inhibits virus replication (By similarity). In testis, production of 25-hydroxycholesterol by macrophages plays a role in Leydig cell differentiation (PubMed:11967195, PubMed:11207193). AH2 + cholesterol + O2 = 25-hydroxycholesterol + A + H2O cholesterol + H(+) + NADPH + O2 = 25-hydroxycholesterol + H2O + NADP(+) Expressed in testicular macrophages at all stages, with the highest level in 10 day old animals. Down-regulated by testosterone. N-glycosylated. Belongs to the sterol desaturase family.