However, the stringency of the screen provides confi dence that the RHFs that were identified are necessary for retrotransposition regardless of the genetic back ground. Although RHFs are Imatinib not a comprehensive set of Ty1 co factors, they are broadly distributed among mo lecular function and biological process categories, sug gesting that they affect many different stages of the Ty1 replication cycle or that numerous cellular pathways in fluence a central process that is necessary for retrotran sposition. A few RHF genes, particularly those whose deletion results in extremely elevated Ty1 cDNA levels, may have been misidentified. This group includes MMS2 and CTF4, two characterized Ty1 repressors.

Moreover, we assume that POL32, a DNA replication and repair gene whose absence increased Ty1 cDNA more than 30 fold, is a Ty1 repressor, since many other genome maintenance genes function as Ty1 repressors. Other genes that may have been misidentified as RHFs are those required for efficient splicing, because the intron within the his3AI indicator gene must be removed by splicing in order to be activated. However, there are only a few RHF genes that are known to play a role in RNA splicing. Our study identified many RHF genes that are con served in eukaryotes. More than half of the RHF genes have statistically robust human homologs, and multiple examples of co factors with human orthologs were iden tified. Human orthologs of RHF genes could play a role in retroviral replication. indeed, human orthologs of the Ty1 co factor Dbr1 and a few repressors of Ty1 retrotransposition have been implicated in analo gous roles in HIV 1 replication.

The human ortholog of DBF20, a novel RHF gene that is necessary for Ty1 cDNA accumulation, encodes the serine threonine kinase, NDR2. NDR2 is incorporated into HIV 1 particles and processed by the HIV 1 prote ase . however, it has not yet been shown to influence Carfilzomib HIV 1 replication directly. Two additional RHFs that are necessary for Ty1 cDNA synthesis or sta bility have human homologs that have been identified in an RNAi screen as presumptive HIV 1 co factors Upf3 and Snf1. One example of an RHF that could provide a clue to facilitate the characterization of an HIV 1 co factor is the class E vacuolar protein sort ing factor, Bro1. Bro1, which was also identified previ ously as a Ty3 co factor, is a homolog of ALIX, which binds to HIV 1 Gag p6 and promotes HIV 1 virion bud ding. Bro1 is also a co factor for replication of Brome Mosaic Virus, a positive strand RNA virus that replicates in S. cerevisiae. BMV replication takes place in membrane bound vesicular invaginations at the perinuclear endoplasmic reticulum.