The transactivation responsive element (TAR) plays a crucial role in the transcription of the HIV-1 genome upon specific binding of the viral protein Tat and cellular proteins. We have previously identified a RNA hairpin aptamer forming a stable and specific kissing complex with TAR RNA (Ducongé, F., and Toulmé, J. J. (1999) RNA 5, 1605-1614). We chemically modified this aptamer with hexitol nucleic acid (HNA) residues. We demonstrate that a fully HNA-modified aptamer is a poor ligand but, in contrast, mixmers containing both HNA and unmodified RNA nucleotides display interesting properties. Two HNA-RNA mixmers bind to TAR with an equilibrium dissociation constant in the low-nanomolar range and show a reduced nuclease sensitivity. In addition, they show a moderate dependence on magnesium ions for binding to TAR. These HNA-RNA mixmers are able to inhibit transactivation of transcription in an in vitro assay.

HCF-1 is a cellular transcriptional coactivator that is critical for mediating the regulated expression of the immediate-early genes of the alphaherpesviruses herpes simplex virus type 1 and varicella-zoster virus. HCF-1 functions, at least in part, by modulating the modification of nucleosomes at these viral promoters to reverse cell-mediated repressive marks and promote activating marks. Strikingly, HCF-1 is specifically sequestered in the cytoplasm of sensory neurons where these viruses establish latency and is rapidly relocalized to the nucleus upon stimuli that result in viral reactivation. However, the analysis of HCF-1 in latently infected neurons and the protein's specific subcellular location have not been determined. Therefore, in this study, the localization of HCF-1 in unstimulated and induced latently infected sensory neurons was investigated and was found to be similar to that observed in uninfected mice, with a time course of induced nuclear accumulation that correlated with viral reactivation. Using a primary neuronal cell culture system, HCF-1 was localized to the Golgi apparatus in unstimulated neurons, a unique location for a transcriptional coactivator. Upon disruption of the Golgi body, HCF-1 was rapidly relocalized to the nucleus in contrast to other Golgi apparatus-associated proteins. The location of HCF-1 is distinct from that of CREB3, an endoplasmic reticulum-resident HCF-1 interaction partner that has been proposed to sequester HCF-1. The results support the model that HCF-1 is an important component of the viral latency-reactivation cycle and that it is regulated by association with a component that is distinct from the identified HCF-1 interaction factors.HCF-1 is a cellular transcription coactivator consisting of amino-and carboxy-terminal subunits generated by site-specific proteolytic processing of a 220-kDa precursor protein (19, 43). The protein was originally identified as a required component of the herpes simplex virus type 1 (HSV-1) immediateearly (IE) gene enhancer core complex in association with the cellular POU-homeodomain protein Oct-1 and the viral IE transactivator VP16 (18,20,42). It has since been shown to be essential for the complex combinatorial regulation of the IE genes of both HSV-1 and varicella-zoster virus (31). The requirement for HCF-1 for the function of multiple transcription factors that regulate the IE genes suggests that it must mediate a common rate-limiting step in transcription initiation and that induction of the viral IE genes is therefore determined at the level of the coactivator.Biochemically, HCF-1 has been identified as a component of several chromatin modification complexes including that of the Set1/MLL1 methyltransferase, Sin3A/histone deacetylase, and the ATAC/GNC5 acetyltransferase (10,44,45). In its role in regulation of the alphaherpesvirus IE genes during lytic infection, HCF-1 mediates the Set1/MLL1-dependent trimethylation of histone H3-lysine 4 of nucleosomes occupying the IE gene promoters, thus promoting the transcriptional in...