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.

We describe a statistical analysis methodology designed to minimize the impact of off-target activities upon large-scale RNA interference (RNAi) screens in mammalian cells. Application of this approach enhances reconfirmation rates and facilitates the experimental validation of new gene activities through the probability-based identification of multiple distinct and active small interfering RNAs (siRNAs) targeting the same gene. We further extend this approach to establish that the optimal redundancy for efficacious RNAi collections is between 4-6 siRNAs per gene.

Mitogen Activated Protein Kinase (MAPK) pathways form the backbone of signal transduction within the mammalian cell. Here, we apply a systematic experimental and computational approach to map 2,269 interactions between human MAPK-related proteins and other cellular machinery and to assemble these data into functional modules. A core network of 641 interactions is supported by multiple lines of evidence including conservation with yeast. Using siRNA knockdowns, we reveal that a significant number of novel interactors can modulate MAPK mediated signaling. We uncover the Na-H exchanger NHE1 as a scaffold for a novel set of MAPKs, link HSP90 chaperones to MAPK pathways, and identify MUC12 as the human analogue to the yeast signaling mucin Msb2. This study makes available a large resource of MAPK interactions along with the accompanying clone libraries. It illustrates a methodology for probing signaling networks based on functional refinement of experimentally-derived protein interaction maps.

A major goal of phytoremediation is to transform fast-growing plants with genes from plant species that hyperaccumulate toxic trace elements. We overexpressed the gene encoding selenocysteine methyltransferase (SMT) from the selenium (Se) hyperaccumulator Astragalus bisulcatus in Arabidopsis and Indian mustard (Brassica juncea). SMT detoxifies selenocysteine by methylating it to methylselenocysteine, a nonprotein amino acid, thereby diminishing the toxic misincorporation of Se into protein. Our Indian mustard transgenic plants accumulated more Se in the form of methylselenocysteine than the wild type. SMT transgenic seedlings tolerated Se, particularly selenite, significantly better than the wild type, producing 3-to 7-fold greater biomass and 3-fold longer root lengths. Moreover, SMT plants had significantly increased Se accumulation and volatilization. This is the first study, to our knowledge, in which a fast-growing plant was genetically engineered to overexpress a gene from a hyperaccumulator in order to increase phytoremediation potential.Although selenium (Se) is a necessary micronutrient for humans and animals at very low doses, it is extremely toxic at higher doses (Wilber, 1983). Excess Se has been implicated in birth defects, sterility, and disease in animals, fish, and wildlife and loss of hair, teeth, and nails, fatigue, and even death in humans (Moxon, 1937;Eisler, 1985;Lemly and Smith, 1987;Sorenson, 1991). Environmental Se pollution is a worldwide problem. Anthropogenic Se pollution arises from many sources, such as aqueous discharges from electric power plants, coal ash leachates, refinery effluents, and industrial wastewater (American Medical Association, 1989). Selenium also occurs naturally in soils formed from Se-bearing shales. This leads to Se-contaminated irrigation drainage water, one of the most serious agricultural problems in the western United States and other areas with similar environments and geological conditions (Presser and Ohlendorf, 1987).Cleaning up Se-contaminated soil and water is a major concern. Phytoremediation, using plants to remove, stabilize, or detoxify pollutants, is a promising technology for remediating Se-contaminated soil and water (Terry et al., 2000). In a process called phytoextraction, plants extract Se from soils and water into their tissues, which can be harvested and removed. Selenium is unusual among trace elements because it can also be removed from the ground ecosystem by phytovolatilization, i.e. plants metabolize inorganic Se to relatively nontoxic, volatile forms (dimethyl selenide [DMSe] and dimethyl diselenide [DMDSe]), which escape to the atmosphere (Lewis et al., 1966;Terry et al., 2000). Selenium phytoremediation has been achieved under field conditions using fast-growing plant species, such as Indian mustard (Brassica juncea; Bañ uelos et al., 1997), which accumulates Se to hundreds of parts per million (Bañ uelos and Schrale, 1989). Hyperaccumulating plant species, such as Astragalus bisulcatus, have adapted to seleniferous soils and accum...