Our previous study showed that the phosphorylation of a highly conserved serine residue, Ser16 in the human immunodeficiency virus type 1 (HIV-1) capsid (CA) protein is promoted by virion-incorporated extracellular signal-regulated kinase 2 (ERK2) and required for proper peptidyl-prolyl isomerase (Pint)-mediated uncoating. Interestingly, western blot analysis demonstrated that phosphorylated/ activated mitogen-activated protein kinase kinase 1/2 (MEK1 /2), the upstream activator of ERK2, as well as ERK2 are incorporated into virions. Here, we show that the MEK1/2 selective allosteric inhibitor Trametinib reduces HIV-1 infectivity via the decrease in virion-incorporated ERK2 phosphorylation. The treatment of chronic HIV-1-infected T-cell line, CEM/LAV-1 cells with Trametinib results in a decrease in ERK2 phosphorylation in the virions. The viruses have relatively low infectivity and impaired reverse transcription. Cell-based fate-of-capsid uncoating assay showed that the reduction in infectivity was caused by a functional impairment of the uncoating process. Furthermore, the viruses from Trametinibtreated CEM/LAV-1 cells also showed decreased reverse transcription efficiency and attenuated multiple rounds of replication in human peripheral blood mononuclear cells (PBMC5). Taken together, these findings suggest that Trametinib suppresses HIV-1 replication by abrogating the proper disassembly of CA core. (C) 2017 Elsevier Inc. All rights reserved.