As previously reported (18), we found that HDAC3 was recruited to the HIV-1 LTR (Fig. (Fig.3B).3B). Additionally, in combination with global HDAC enzymatic inhibition by TSA, siRNA-mediated knockdown of HDAC3 resulted in a synergistic increase in HIV-1 expression (Fig. (Fig.4B).4B). We achieved a substantial knockdown of HDAC3 mRNA (93%; data not shown), but LTR upregulation was seen only when the HDAC3 knockdown was combined with submaximal global HDAC inhibition. Persistent activity of a large cellular pool of HDAC protein, despite mRNA inhibition, could explain this finding. Alternatively, as has been reported with other HDACs, HDAC3 may function as a transcriptional repressor via a function that does not depend on its deacetylase activity (7, 17, 28). In the presence of global HDAC enzymatic inhibition, HDAC3 may continue to restrict LTR expression. This repression could be relieved by HDAC3 knockdown but would not be apparent without inhibition of other resident HDACs, e.g., HDAC1 and HDAC2.