BRYAN CULLEN

..Using high-throughput deep sequencing, we examined the miRNA repertoire in Ae. albopictus cells and Cx. quinquefasciatus mosquitoes...

..to identify the cellular targets of these viral proteins and RNA elements have led to the identification of Crm1 and Tap as essential human nuclear RNA-export factors and continue to provide insights into how mRNAs are selected for export..

..Research into viral miRNA function might also suggest new approaches for treating some virally induced diseases...

..While each of these entry points offers distinct advantages and disadvantages, they all have the potential to induce the effective knock-down of specific genes either in cell culture or in experimental animals...

..Together, these data argue that adenovirus infection can result in inhibition of RNAi and identify VA1 RNA as the first viral gene product able to inhibit RNAi in human cells...

..This review describes recent insights into the mechanisms governing microRNA transcription and processing in vertebrates and their implications for understanding the regulation of microRNA biogenesis...

..Here I briefly review the potential pitfalls associated with RNAi, and then suggest experimental approaches that can be used to maximize the specificity of RNAi or to confirm that the data obtained are indeed valid...

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..Gene Therapy (2006) 13, 503-508. doi:10.1038/sj.gt.3302656; published online 29 September 2005...

..Emerging data suggest that viral microRNAs are particularly important for regulating the transition from latent to lytic replication and for attenuating antiviral immune responses...

..Identification of HIV-1 variants capable of entering T cells via the CD8 receptor suggests a new mode of viral pathogenesis. But are these variants rare, aberrant viruses or a real problem?..

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..Surprisingly, RNA was also found to be essential for formation of the nucleocapsid--APOBEC3G complex in vitro, thus raising the possibility that RNA may form a bridge between these two proteins...

..Moreover, these results suggest that the biological barrier preventing the entry of additional SIV into the human population as zoonotic infections is potentially quite fragile...

..While this inhibition was significantly less than the level seen with wild-type forms of A3G or A3B, these data, nevertheless argue that the inhibition of HIV-1 by APOBEC3 proteins is at least partly independent of DNA editing...

..Together, these findings define an additional cellular cofactor required for miRNA biogenesis and function...

..Truly nucleoplasmic mRNAs or pre-mRNAs are, in contrast, resistant to RNAi...

..In contrast, both genes are quiescent in the permissive CEM derivative CEM-SS. Together, these data argue that HIV-1 Vif has evolved to suppress at least two distinct but related human antiretroviral DNA-editing enzymes...

..Insights derived from this research will facilitate a better understanding of viral pathogenesis and the host innate immune response to viral infection...

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..These data may suggest that truly nuclear NMD, if it occurs, is at least in part mechanistically distinct from cytoplasmic NMD...

..Here we discuss our current knowledge of viral miRNAs and virally influenced cellular miRNAs and their relationship to viral infection...

..It therefore appears that EIAV has evolved a novel mechanism to specifically neutralize the biological activities of the cognate eA3F1 and eA3F2 proteins at a step subsequent to virion incorporation...

..These studies therefore raise the possibility that activation of the endogenous APOBEC3B gene in primary human lymphoid cells could form a novel and effective strategy for inhibition of HIV-1 replication in vivo...

..Here, I discuss the identification and characterization of viral mRNA structures and noncoding RNAs that have led to important insights into the molecular mechanisms of eukaryotic cells...

..These data indicate that novel miRNAs can be readily produced in vivo and can be designed to specifically inactivate the expression of selected target genes in human cells...

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..These data suggest that NXT1 may act as a molecular switch that regulates the ability of Tap to mediate nuclear mRNA export by controlling the interaction of Tap with components of the nuclear pore...

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..Here, we discuss what is known about how viral and cellular miRNAs influence viral replication and pathogenic potential through their regulation of viral mRNAs or by reshaping cellular gene expression...

..Moreover, these results argue that Exp5 binding not only mediates pre-microRNA nuclear export but also prevents nuclear pre-microRNA degradation...

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..These data suggest that it should be possible to block the expression of specific viral or cellular genes in vivo by using viral vectors to stably express the appropriate siRNAs...

..This is the first report of a naturally occurring sequence polymorphism in an miRNA precursor that results in reduced processing and therefore lower levels of mature miRNA expression and function...

..e., the base state of a naïve retrovirus is susceptibility to inhibition...

..Moreover, the known aetiological role of miR-155 in B-cell transformation suggests that miR-K12-11 may contribute to the induction of KSHV-positive B-cell tumours in infected patients...

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..While the sites of Drosha cleavage are determined largely by the distance from the terminal loop, variations in stem structure and sequence around the cleavage site can fine-tune the actual cleavage sites chosen...

..Moreover, in the case of the highly intron-dependent beta-globin gene, splicing also significantly enhanced the translational utilization of cytoplasmic beta-globin mRNAs...

..The requirement for single-stranded extensions on the pri-miRNA hairpin substrate for Drosha processing is currently unique among the RNase III enzymes...

..These results suggest that miRNAs, and the closely similar small interfering RNAs, cannot totally discriminate between RNA targets differing by a single nucleotide...

..Together, these data show that human pri-miRNAs are not only structurally similar to mRNAs but can, in fact, function both as pri-miRNAs and mRNAs...

..These data have implications for the future clinical utilization of shRNAs and also provide a simple method to enhance RNA interference by shRNAs in culture...

..Together, these data argue that HIV-1 and HTLV-1 neither induce the production of viral small interfering RNAs or microRNAs nor repress the cellular RNA interference machinery in infected cells...

..Thus, HSV-1 expresses at least two primary miRNA precursors in latently infected neurons that may facilitate the establishment and maintenance of viral latency by post-transcriptionally regulating viral gene expression...

..Overexpression or inappropriate expression of authentic microRNAs may facilitate the study of their normal functions and expression of artificial microRNAs may permit effective, regulated RNA interference in vivo...

..These data set the stage for the mutational ablation and phenotypic analysis of RRV mutants lacking one or more viral microRNAs...

..This site can be a natural, high affinity RNA target for the nuclear export factor or alternately the export factor can be tethered to the unspliced cat mRNA by fusion to a heterologous RNA binding domain...

..These data demonstrate that H19 can function as a primary microRNA precursor and suggest that H19 expression results in the post-transcriptional downregulation of specific mRNAs during vertebrate development...

..We hypothesize that these viral miRNAs play a critical role in the establishment and/or maintenance of KSHV latent infection in vivo and, hence, in KSHV-induced oncogenesis...

..These data suggest that miRNAs and siRNAs can use similar mechanisms to repress mRNA expression and that the choice of mechanism may be largely or entirely determined by the degree of complementary of the RNA target...

..Although over 500 cellular microRNAs were cloned and identified, not a single HPV31-specific microRNA was obtained. We therefore concluded that HPV31, and possibly human papillomaviruses in general, does not express viral microRNAs...

..Moreover, these results suggest that APOBEC3 proteins may help prevent the zoonotic infection of humans by simple retroviruses and provide a mechanism for how simple retroviruses can avoid inhibition by APOBEC3 family members...

..Moreover, the differential regulation of EBV miRNA expression implies distinct roles during infection of different human tissues...

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..Techniques to achieve and validate the expression of viral miRNAs are described in this review...

..Several other RRV-derived moRNAs were obtained, including one recovered >600 times. Together, this research provides a comprehensive list of the miRNAs and moRNAs encoded by RRV...

..The second critical CTE-interacting surface on Tap is defined by three previously identified residues on the surface of the ribonucleoprotein domain. The structural and mutational data define a novel RNA-binding site on the Tap protein...

..While these data identify the foamy virus Bet protein as a functional ortholog of the HIV-1 Vif auxiliary protein, they also indicate that Vif and Bet block APOBEC3 protein function by distinct mechanisms...

..Our findings indicate that APOBEC3G restricts Ty1 and HIV-1 by similar mechanisms and suggest that the APOBEC3 proteins target a substantially broader spectrum of retroelements than previously appreciated...

..This chapter describes methods that allow the overexpression of human miRNAs and also discusses how primary miRNAs (pri-miRNAs), the much longer precursors of mature miRNAs, are processed in human cells, as well as in vitro...

..Thus, human topoisomerase I plays an important role in HIV-1 replication and infectivity, and differences in the species specificity of HIV-1 infection can at least in part be attributed to differences in topoisomerase I activities...

..These data demonstrate a differential restriction of L1 and Alu retrotransposition by APOBEC3G, and suggest that the Alu ribonucleoprotein complex may be targeted by APOBEC3G...

..CONCLUSIONS: This work is the first to demonstrate that delivery of RNAi by viral vectors has therapeutic potential for chronic HBV infection and establishes the ground work for the use of such vectors in vivo...

..Together, these studies should identify human mRNA targets for several KSHV and EBV miRNAs and shed light on the role of these miRNAs in the replication cycle of Gamma herpesviruses in general. ..

..Together, this wide-ranging series of experiments should lead to considerable insight into how HIV-1 Vif functions and potentially suggest approaches to inhibit Vif function. ..

..Finally, in specific aim 5, we seek to extend our exciting initial data demonstrating inhibition of miRNA function by the adenovirus VA1 non-coding RNA by trying to define the underlying mechanism. ..

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..Together, these studies should identify human mRNA targets for several KSHV and EBV miRNAs and shed light on the role of these miRNAs in the replication cycle of Gamma herpesviruses in general. ..

..The final specific aim focusses on the promoter regions of CCR5, the identification of transcription factors that may interact with the promoter, and the use of antisense strategies to block CCR5 expression. ..

..This research has the potential to not only shed new light on HIV-1 pathogenesis but also suggest new approaches to inhibit virus replication. ..