Zika virus (ZIKV) infection during pregnancy causes severe neurological and ocular abnormalities in infants, yet no vaccine or antivirals are available. Our transcriptomic analysis of ZIKV-infected retinal pigment epithelial (RPE) cells revealed alterations in the cholesterol pathway. Thus, we investigated the functional roles of ATP binding cassette transporter G1 (ABCG1) and sterol response element binding protein 2 (SREPB-2), two key players in cholesterol metabolism, during ocular ZIKV infection. Our in vitro data showed that increased ABCG1 activity via liver X receptors (LXRs), reduced ZIKV replication, while ABCG1 knockdown increased replication with elevated intracellular cholesterol. Conversely, inhibiting SREBP-2 or its knockdown reduced ZIKV replication by lowering cholesterol levels. In vivo, LXR agonist or SREBP-2 inhibitor treatment mitigated ZIKV-induced chorioretinal lesions in mice, concomitant with decreased expression of inflammatory mediators and increased activation of antiviral response genes. In summary, our study identifies ABCG1's antiviral role and SREBP-2's proviral effects in ocular ZIKV infection, offering cholesterol metabolism as a potential target to develop antiviral therapies.