The TAR microRNA is expressed in all stages of the viral life cycle, can be detected in latently infected cells, and represents a mechanism wherein the virus extends the life of the infected cell for the purpose of increasing viral replication.

MicroRNAs (miRNAs) are a class of small RNA molecules that function to control gene expression and restrict viral replication in host cells. The production of miRNAs is believed to be dependent upon the DICER enzyme. Available evidence suggests that in T lymphocytes, HIV-1 can both suppress and co-opt the host's miRNA pathway for its own benefit. In this study, we examined the state of miRNA production in monocytes and macrophages as well as the consequences of viral infection upon the production of miRNA. Monocytes in general express low amounts of miRNA-related proteins, and DICER in particular could not be detected until after monocytes were differentiated into macrophages. In the case where HIV-1 was present prior to differentiation, the expression of DICER was suppressed. MicroRNA chip results for RNA isolated from transfected and treated cells indicated that a drop in miRNA production coincided with DICER protein suppression in macrophages. We found that the expression of DICER in monocytes is restricted by miR-106a, but HIV-1 suppressed DICER expression via the viral gene Vpr. Additionally, analysis of miRNA expression in monocytes and macrophages revealed evidence that some miRNAs can be processed by both DICER and PIWIL4. Results presented here have implications for both the pathology of viral infections in macrophages and the biogenesis of miRNAs. First, HIV-1 suppresses the expression and function of DICER in macrophages via a previously unknown mechanism. Second, the presence of miRNAs in monocytes lacking DICER indicates that some miRNAs can be generated by proteins other than DICER. Human immunodeficiency virus, type 1 (HIV-1) infects multiple types of cells, including CD4ϩ T-cells, macrophages, monocytes, dendritic cells, and microglial cells (1-5). It has also been reported that latently infected hematopoietic stem cells can be detected in HIV-1-positive patients (6, 7). However, despite the range of susceptible cell types, HIV-1 is typically discussed in the context of T-cell infection and the resulting depletion of CD4 ϩ T-cells. Conversely, research into HIV-1 infection in the context of monocytes and macrophages remains underdeveloped. Monocytes and macrophages have been previously shown to harbor both productive and latent infections, serving as a part of the viral reservoir that could potentially be essential to chronic infection (8, 9). Unlike T-cells, macrophages can resist the cytotoxic effects of HIV-1 infection and persist for several months while producing new infectious viral particles (10 -12). In fact, it has been observed that latently infected macrophages are responsible for a large amount of viral resurgence after cessation of anti-retroviral therapies (13). Interestingly, we have previously observed that monocytes differ greatly from T-cells in their expression of miRNA-related enzymes. Specifically, monocytes appeared to be deficient for the DICER enzyme (14). This observation was intriguing because previous reports have indicated that HIV-1 can manipulate the host's RNA inter...

The development of novel techniques and systems to study human infectious diseases in both an in vitro and in vivo settings is always in high demand. Ideally, small animal models are the most efficient method of studying human afflictions. This is especially evident in the study of the human retroviruses, HIV-1 and HTLV-1, in that current simian animal models, though robust, are often expensive and difficult to maintain. Over the past two decades, the construction of humanized animal models through the transplantation and engraftment of human tissues or progenitor cells into immunocompromised mouse strains has allowed for the development of a reconstituted human tissue scaffold in a small animal system. The utilization of small animal models for retroviral studies required expansion of the early CB-17 scid/scid mouse resulting in animals demonstrating improved engraftment efficiency and infectivity. The implantation of uneducated human immune cells and associated tissue provided the basis for the SCID-hu Thy/Liv and hu-PBL-SCID models. Engraftment efficiency of these tissues was further improved through the integration of the nonobese diabetic (NOD) mutation leading to the creation of NODSCID, NOD/Shi-scid IL2rγ -/-, and NOD/SCID β2-microglobulin null animals. Further efforts at minimizing the response of the innate murine immune system produced the Rag2 -/-γ c -/-model which marked an important advancement in the use of human CD34+ hematopoietic stem cells. Together, these animal models have revolutionized the investigation of retroviral infections in vivo.

BackgroundApproximately half of hereditary breast cancers have mutations in either BRCA1 or BRCA2. BRCA1 is a multifaceted tumor suppressor protein that has implications in processes such as cell cycle, transcription, DNA damage response and chromatin remodeling. This multifunctional nature of BRCA1 is achieved by exerting its many effects through modulation of transcription. Many cellular events are dictated by covalent modification of proteins, an important mechanism in regulating protein and genome function; of which protein methylation is an important posttranslational modification with activating or repressive effects.Methods/Principal FindingsHere we demonstrate for the first time that BRCA1 is methylated both in breast cancer cell lines and breast cancer tumor samples at arginine and lysine residues through immunoprecipitation and western blot analysis. Arginine methylation by PRMT1 was observed in vitro and the region of BRCA1 504–802 shown to be highly methylated. PRMT1 was detected in complex with BRCA1 504–802 through in vitro binding assays and co-immunoprecipitated with BRCA1. Inhibition of methylation resulted in decreased BRCA1 methylation and alteration of BRCA1 binding to promoters in vivo as shown through chromatin immunoprecipitation assays. Knockdown of PRMT1 also resulted in increased BRCA1 binding to particular promoters in vivo. Finally, following methylation inhibition, Sp1 was found to preferentially associate with hypo-methylated BRCA1 and STAT1 was found to preferentially associate with hyper-methylated BRCA1.Conclusions/SignificanceThese results suggest that methylation may influence either the ability of BRCA1 to bind to specific promoters or protein-protein interactions which alters the recruitment of BRCA1 to these promoters. Thus, given the importance of BRCA1 to genomic stability, methylation of BRCA1 may ultimately affect the tumor suppressor ability of BRCA1.