Human Immunodeficiency Viruses (HIV-1 and HIV-2) rely upon host-encoded proteins to facilitate their replication. Here we combined genome-wide siRNA analyses with interrogation of human interactome databases to assemble a host-pathogen biochemical network containing 213 confirmed host cellular factors and 11 HIV-1-encoded proteins. Protein complexes that regulate ubiquitin conjugation, proteolysis, DNA damage response and RNA splicing were identified as important modulators of early stage HIV-1 infection. Additionally, over 40 new factors were shown to specifically influence initiation and/or kinetics of HIV-1 DNA synthesis, including cytoskeletal regulatory proteins, modulators of post-translational modification, and nucleic acid binding proteins. Finally, fifteen proteins with diverse functional roles, including nuclear transport, prostaglandin synthesis, ubiquitination, and transcription, were found to influence nuclear import or viral DNA integration. Taken together, the multi-scale approach described here has uncovered multiprotein virus-host interactions that likely act in concert to facilitate early steps of HIV-1 infection.

The molecular mechanisms involved in transgene‐induced gene silencing (‘quelling’) in Neurospora crassa were investigated using the carotenoid biosynthetic gene albino‐1 (al‐1) as a visual marker. Deletion derivatives of the al‐1 gene showed that a transgene must contain at least approximately 132 bp of sequences homologous to the transcribed region of the native gene in order to induce quelling. Transgenes containing only al‐1 promoter sequences do not cause quelling. Specific sequences are not required for gene silencing, as different regions of the al‐1 gene produced quelling. A mutant defective in cytosine methylation (dim‐2) exhibited normal frequencies and degrees of silencing, indicating that cytosine methylation is not responsible for quelling, despite the fact that methylation of transgene sequences frequently is correlated with silencing. Silencing was shown to be a dominant trait, operative in heterokaryotic strains containing a mixture of transgenic and non‐transgenic nuclei. This result indicates that a diffusable, trans‐acting molecule is involved in quelling. A transgene‐derived, sense RNA was detected in quelled strains and was found to be absent in their revertants. These data are consistent with a model in which an RNA‐DNA or RNA‐RNA interaction is involved in transgene‐induced gene silencing in Neurospora.

IB kinase 2 (IKK2 or IKK␤) is a component of the IKK complex that coordinates the cellular response to a diverse set of extracellular stimuli, including cytokines, microbial infection, and stress. In response to an external stimulus, the complex is activated, resulting in the phosphorylation and subsequent proteasome-mediated degradation of IB proteins. This event triggers the nuclear import of the NF-B transcription factor, which activates the transcription of genes that regulate a variety of fundamental biological processes, including immune response, cell survival, and development. Here, we define an essential role for IKK2 in normal mitotic progression and the maintenance of spindle bipolarity. Chemical and genetic perturbation of IKK2 promotes the formation of multipolar spindles and chromosome missegregation. Depletion of IKK2 results in the deregulation of Aurora A protein stability and coincident hyperactivation of a putative Aurora A substrate, the mitotic motor KIF11. These data support a function for IKK2 as an antagonist of Aurora A signaling during mitosis. Additionally, our results indicate a direct role for IKK2 in the maintenance of genome stability and underscore the potential for oncogenic consequences in targeting this kinase for therapeutic intervention.Aurora A ͉ mitosis ͉ NF-B ͉ spindle polarity

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The CDKN2A locus encodes two important tumor suppressors, INK4a and ARF, which respond to oncogenic stresses by inducing cellular senescence. We conducted a genome-scale cDNA overexpression screen using a reporter containing INK4a regulatory sequences to identify novel transcriptional activators of this locus. This screen revealed 285 cDNAs that putatively regulate the transcriptional activation of INK4a. Of these, 56 are annotated as transcription factors, including two previously reported activators of the locus, ETS2 and JUNB. Fourteen genes were further validated for activity and specificity, including several homeodomain proteins. We found that the transcription of one of these, the homeodomain protein MEOX2 (GAX) is enhanced in primary cells during the induction of senescence, and forced expression of this protein results in the induction of premature senescence. We further demonstrate that MEOX2-induced senescence is dependent upon INK4a activity, and chromatin immunoprecipitation studies indicate that MEOX2 directly binds the INK4a promoter. These results support a role for this homeodomain protein as a direct regulator of INK4a transcription and senescence in human cells.