Elite controllers represent a unique group of HIV-1-infected persons with undetectable HIV-1 replication in the absence of antiretroviral therapy. However, the mechanisms contributing to effective viral immune defense in these patients remain unclear. Here, we show that compared with HIV-1 progressors and HIV-1-negative persons, CD4+ T cells from elite controllers are less susceptible to HIV-1 infection. This partial resistance to HIV-1 infection involved less effective reverse transcription and mRNA transcription from proviral DNA and was associated with strong and selective upregulation of the cyclin-dependent kinase inhibitor p21 (also known as cip-1 and waf-1). Experimental blockade of p21 in CD4+ T cells from elite controllers resulted in a marked increase of viral reverse transcripts and mRNA production and led to higher enzymatic activities of cyclin-dependent kinase 9 (CDK9), which serves as a transcriptional coactivator of HIV-1 gene expression. This suggests that p21 acts as a barrier against HIV-1 infection in CD4+ T cells from elite controllers by inhibiting a cyclin-dependent kinase required for effective HIV-1 replication. These data demonstrate a mechanism of host resistance to HIV-1 in elite controllers and may open novel perspectives for clinical strategies to prevent or treat HIV-1 infection.