The objective of the research detailed in this application is to gain an understanding of one of the basic processes of eucaryotic gene expression. Considering the linear arrangement of genetic information, the formation of the 3 end of mRNA is an important feature of selective gene expression. The 3 ends of the histone mRNAs are generated by processing precursor RNAs.
The specific aim of the proposed research is to characterize the histone mRNA processing activity purified from nuclear extracts of Drosophila Kc cells. Because of the ability to observe the processing of specific precursor RNAs by a highly purified activity in vitro, details of the molecular mechanism of action will be studied. The sequence requirements for the activity will be determined by using precursor RNAs generated from various naturally occurring histone genes as well as genes modified by in vitro mutagenesis. The involvement of a small RNA species which seems to be associated with the activity will be examined. Precursor RNAs will be synthesized either in the presence of the Kc cell nuclear extract using dC-tailed templates or with no extract present using the bacterial SP6 system to see if modification of the precursor RNA by the Kc cell extract is necessary for accurate processing. Using antisera produced to the purified processing factor and/or human lupus antisera a recombinant Drosophila expression library will be screened and the gene for the processing factor will be cloned. If a small RNA species is required for activity its gene will also be obtained by screening a recombinant Drosophila genomic library with cDNA obtained from the RNA from the highly purified factor. A Kc cell transfection system will be used to study the expression of these genes in vivo. Defects in gene expression including defects in RNA processing are the cause of many of the most serious health problems. Gaining an understanding of this basic cellular process will undoubtedly contribute to our knowledge of gene expression and eventually will aid in the treatment of disease.
| Price, David H (2018) Transient pausing by RNA polymerase II. Proc Natl Acad Sci U S A 115:4810-4812 |
| Mullen, Nicholas J; Price, David H (2017) Hydrogen peroxide yields mechanistic insights into human mRNA capping enzyme function. PLoS One 12:e0186423 |
| Brogie, John E; Price, David H (2017) Reconstitution of a functional 7SK snRNP. Nucleic Acids Res 45:6864-6880 |
| Nilson, Kyle A; Lawson, Christine K; Mullen, Nicholas J et al. (2017) Oxidative stress rapidly stabilizes promoter-proximal paused Pol II across the human genome. Nucleic Acids Res 45:11088-11105 |
| Bosque, Alberto; Nilson, Kyle A; Macedo, Amanda B et al. (2017) Benzotriazoles Reactivate Latent HIV-1 through Inactivation of STAT5 SUMOylation. Cell Rep 18:1324-1334 |
| Tan, Justin L; Fogley, Rachel D; Flynn, Ryan A et al. (2016) Stress from Nucleotide Depletion Activates the Transcriptional Regulator HEXIM1 to Suppress Melanoma. Mol Cell 62:34-46 |
| Nilson, Kyle A; Guo, Jiannan; Turek, Michael E et al. (2015) THZ1 Reveals Roles for Cdk7 in Co-transcriptional Capping and Pausing. Mol Cell 59:576-87 |
| Guo, Jiannan; Turek, Michael E; Price, David H (2014) Regulation of RNA polymerase II termination by phosphorylation of Gdown1. J Biol Chem 289:12657-65 |
| Gu, Jianyou; Babayeva, Nigar D; Suwa, Yoshiaki et al. (2014) Crystal structure of HIV-1 Tat complexed with human P-TEFb and AFF4. Cell Cycle 13:1788-97 |
| Mullen Davis, Melissa A; Guo, Jiannan; Price, David H et al. (2014) Functional interactions of the RNA polymerase II-interacting proteins Gdown1 and TFIIF. J Biol Chem 289:11143-52 |
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