Identification of Homeodomain Proteins, PBX1 and PREP1, Involved in the Transcription of Murine Leukemia Virus

Abstract

Cyclin-dependent kinase inhibitors (CDKIs) have been shown to block human immunodeficiency virus and herpes simplex virus. It is hypothesized that CDKIs block viral replication by inhibiting transcription of specific cellular genes. Here we find that three CDKIs, flavopiridol, purvalanol A, and methoxy-roscovitine, block Moloney murine leukemia virus (MLV) transcription events. Using gene expression microarray technology to examine the inhibitory effects of CDKIs, we observed a cellular gene, the pre-B-cell leukemia transcription factor 1 (Pbx1) gene, down-regulated by CDKI treatment. The PBX consensus element (PCE), TGATTGAC, is conserved in the long terminal repeats of several murine retroviruses, including Moloney MLV. Mutations in the PCE completely inhibited viral transcription whereas overexpression of PBX1 and a PBX1-associated protein, PREP1, enhanced viral transcription. The interaction between the PCE and PBX1-PREP1 proteins was confirmed by gel shift experiments. Blocking PBX1 protein synthesis resulted in a significant decrease in viral transcription. Collectively, our results represent the first work demonstrating that the homeodomain proteins PBX1 and PREP1 are cellular factors involved in Moloney MLV transcription regulation.Cyclin-dependent kinases (CDKs) are key regulators of cell cycle control and transcription. Nine CDKs have been identified thus far. CDK1 to CDK7 are required for cell cycle regulation, and CDK7 to CDK9 are involved in RNA polymerase II-dependent transcription (33). In fact, CDK2, CDK4, and CDK6 also regulate transcription by activating E2F via specific phosphorylation of the retinoblastoma gene product (Rb protein) in complex with E2F (13, 49). It has been shown elsewhere that approximately 90% of all neoplasias are associated with CDK hyperactivation and subsequent Rb pathway inactivation (21). Therefore, CDKs represent attractive targets for cancer therapy, and several CDK inhibitors (CDKIs) have been developed (4,16,18,48). Two CDKIs, flavopiridol (Flavo) and UCN-1, are already in clinical trials as potential anticancer therapeutics (44,49).The antiviral activities of CDKIs were first observed in studies describing their interaction with positive transcription elongation factor b (P-TEFb), a factor required for human immunodeficiency virus (HIV) replication (32, 54). P-TEFb, composed of CDK9 and a cyclin subunit derived from one of three different genes (cyclin T1, T2, or K), controls RNA polymerase II-dependent transcription elongation by phosphorylating the carboxyl-terminal domain of the large subunit of RNA polymerase II (41).

Roscovitine Inhibits Activation of Promoters in Herpes Simplex Virus Type 1 Genomes Independently of Promoter-Specific Factors

“…However, it has been documented previously that Rosco (39) and a methylated derivative (15) only very limited effects on cellular transcription in kidneys of mice. In fact, only 3 (0.055%) of 5,400 transcripts were downregulated more than twofold by Rosco whereas 18 others (0.33%) were actually upregulated by the drug.…”

“…Flavo inhibits retroviral transcription (13), probably as a result of its effects on CDK9 (45), whereas Rosco inhibits herpesvirus DNA replication (9,74), probably as a result of its effect on CDK2 (37). Surprisingly, Rosco and other purine PCIs also inhibit the accumulation of transcripts driven by promoters of HSV-1 (33), Epstein-Barr virus (38), adenovirus type 5 (23), human immunodeficiency virus type 1 (84), mouse mammary tumor virus (6), and murine leukemia virus (15).…”

“…Purine-type PCIs inhibit the expression of Pbx1 and Prep1, two transcription factors that are required for activation of the murine leukemia virus promoter (15). Rosco and another purine-type PCI, CVT-313, inhibit the remodeling of chromatin that is required for binding of the glucocorticoid receptor to its cognate sequences in the mouse mammary tumor virus promoter (6).…”

Novel Neuroprotective GSK-3β Inhibitor Restricts Tat-Mediated HIV-1 Replication

“…The results in Fig. 6A show RT-PCR amplifications of CAD, CIITA, cyclin D1, and PBX-1, a set of genes that require Cdk9/cyclin T1for their transcription (108)(109)(110). None of these genes showed a decrease in transcription in various cell types, including 293T cells, CHME5 human microglial cells, and Jurkat T cells.…”

Prep1/Pbx2 complexes regulate CCL2 expression through the −2578 guanine polymorphism

“…Interestingly, Prep1/Pbx2 complexes have been reported to repress promoters 31,32 as well as activate promoters 29,33 containing the TGACAG motif in a cell-type-dependent manner, 32 whereas Prep1/Pbx1 complexes have only been reported to activate promoters. 28,34 This study has shown that Prep1/Pbx2 nuclear complexes can bind the À2578 G oligonucleotides under unstimulated and IL-1b-stimulated conditions where promoter activity from the G allele is repressed and equivalent, respectively, as compared to the A allele. These data suggest that the À2578 site does not influence IL-1b-stimulated CCL2 expression in astrocytes.…”