HIV-1 and BLV are insensitive to SERINC5 restriction under the cell-cell infection

<u>Ser</u>ine <u>inc</u>orporator 5 (<u>SER</u>INC5, SER5) suppresses viral cell-free infection. However, its antiviral potency under viral cell-cell infection is not examined yet. Here, we established the <i>in vitro</i> cell-cell infection systems to assess SER5's antiviral activity on HIV-1 and bovine leukemia virus (BLV). Our results showed SER5 from different mammalian species, including <i>Homo sapiens</i>, <i>Bos taurus</i>, and <i>Felis catus</i>, was capable of significantly inhibiting HIV-1 cell-free infection. However, these SER5s were unable to restrict HIV-1 cell-cell infection. Intriguingly, Ebola virus glycoprotein-mediated pseudoviral cell-free infection was greatly enhanced by <i>Homo sapiens</i> and <i>Bos taurus</i> SER5. Notably, BLV was also insensitive to SER5-mediated restriction under the cell-cell infection. In addition, BLV envelope glycoproteins, which contained one glycosylation variation, were not restricted by SER5 in the model of cell-cell infection. Overall, our results indicated SER5 was ineffective against HIV-1 and BLV under the cell-cell infection, which was utilized by them to circumvent SER5-imposed restriction <i>in vitro</i> and may aid their transmission <i>in vivo</i>.IMPORTANCESER5 potently inhibits virus infection under the cell-free mode. However, few studies check whether SER5 keeps its restriction under the virus cell-cell transmission. In this work, we uncover SER5 loses its restriction to HIV-1 and BLV under the mode of cell-cell infection, demonstrating the viruses could employ this mode to overcome SER5 restriction and thus facilitate their transmission.