Although current antiretroviral therapies (ART) are successful in controlling HIV-1 infection, a stable viral reservoir reactivates when ART is discontinued. Consequently, there is a major research effort to develop approaches to disrupt the latent viral reservoir and enhance the immune system's ability to clear HIV-1. A number of small molecules, termed latency reversal agents (LRAs), have been identified which can reactivate latent HIV-1 in cell lines and patients' cells ex vivo. However, clinical trials have suggested that combinations of LRAs will be required to efficiently reactivate HIV-1 in vivo, especially LRAs that act synergistically by functioning through distinct pathways. To identify novel LRAs, we used an image-based assay to screen a natural compound library for the ability to induce a low level of aggregation of resting primary CD4+ T cells from healthy donors. We identified celastrol as a novel LRA. Celastrol functions synergistically with other classes of LRA to reactivate latent HIV-1 in a Jurkat cell line, suggesting a novel mechanism in its LRA activity. Additionally, celastrol does not appear to activate resting CD4+ T cells at levels at which it can reactivate latent HIV-1. Celastrol appears to represent a novel class of LRAs and it therefore can serve as a lead compound for LRA development.