Published online 1 April 2011 Nucleic Acids Research, 2011, Vol. 39, No. 14 5879–5892 doi:10.1093/nar/gkr198 Cellular heat shock factor 1 positively regulates human immunodeficiency virus-1 gene expression and replication by two distinct pathways Pratima Rawat and Debashis Mitra* National Centre for Cell Science, Ganeshkhind, Pune 411 007, India Received January 18, 2011; Revised March 2, 2011; Accepted March 17, 2011 ABSTRACT Human immunodeficiency virus-1 (HIV-1) infection leads to changes in cellular gene expression, which in turn tend to modulate viral gene expression and replication. Cellular heat shock proteins (HSPs) are induced upon heat shock, UV irradiation and mi- crobial or viral infections. We have reported earlier Nef-dependent induction of HSP40 leading to incre- ased HIV-1 gene expression; however, the mechan- ism of induction remained to be elucidated. As expression of HSPs is regulated by heat shock factors (HSFs), we have now studied the role of HSF1 not only in Nef-dependent HSP40 induction but also in HIV-1 gene expression. Our results show that HSF1 is also induced during HIV-1 infection and it positively regulates HIV-1 gene expression by two distinct pathways. First, along with Nef it activates HSP40 promoter which in turn leads to increased HIV-1 gene expression. Second, HSF1 directly inter- acts with newly identified HSF1 binding sequence on HIV-1 LTR promoter and induces viral gene ex- pression and replication. Thus, the present work not only identifies a molecular basis for HSF1-mediated enhancement of viral replication but also provides another example of how HIV-1 uses host cell machinery for its successful replication in the host. INTRODUCTION Cellular heat shock proteins (HSPs) are molecular chap- erones primarily involved in protein folding, transport and assembly. In addition, some of these proteins are speciﬁc- ally induced during stress conditions like heat shock, UV irradiation and microbial/viral infection (1,2). Recent studies have revealed that HSPs are also involved in apop- tosis and immune response (3,4). Viruses modulate expres- sion of many cellular proteins for their successful replication and induction of HSPs has been reported as one of the earliest change following viral infection (5). Human immunodeﬁciency virus-1 (HIV-1) was reported to induce HSP27 and HSP70 expression during early in- fection (6). HIV-1 Nef, a 27–30 kDa myristoylated phosphoprotein, contributes to viral pathogenesis by modulating cellular gene expression and signaling pathways (7). Nef has been also implicated in the activation of T cells, making the cells permissible to the virus (8). Although initially reported as a negative factor for HIV-1 replication in T-cell lines (9,10), Nef has been later demonstrated to be an enhancer of virus replication (11–14). However, the molecular mechanism of this positive effect remains to be clearly understood. We have earlier shown that Nef interacts with HSP40, and this interaction was necessary for Nef mediated increase in viral gene expression and replication. Furthermore, it was also shown that HSP40 expression increased in HIV-1 NL4-3 transfected cells in a Nef-dependent manner (15). However, the mechanism of HSP40 upregulation during HIV-1 infection remains to be elucidated. The inducible expression of HSPs is primarily regulated by heat shock factors (HSFs). HSF1 is the major tran- scription factor that regulates the transcription of HSP genes in response to stress. It binds to conserved regula- tory sequences in the HSP promoters known as heat shock elements (HSE), which is represented by two or three inverted repeats of the sequence nGAAn (16). Normally, HSF1 is predominantly present in a cytoplasmic mono- meric inactive form; however, upon stress it gets homo- trimerized and translocated to nucleus and acquires high afﬁnity HSE binding and transcription enhancing activity. Recent studies indicate that phosphorylation also plays a major role in regulation of HSF1 activity; speciﬁcally Ser230 and Ser326 are inducibly phosphorylated during stress resulting in increased transcriptional activity (17,18). The response of HIV-1 to various stress proteins, including HSPs, could also lead to modulation of HIV-1 long ter- minal repeat promoter (LTR)-driven gene expression (19). *To whom correspondence should be addressed. Tel: +91 20 25708151; Fax: +91 20 25692259; Email: dmitra@nccs.res.in; dmitra01@yahoo.co.in The Author(s) 2011. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.