The 3'untranslated region (UTR) and the poly(A) tail of an mRNA can exert regulatory influences that affect message stability, rate of translation, and/or subcellular localization. Thus, the choice of where the cleavage reaction takes place in a pre-mRNA can have significant influence on its subsequent activity and fate. Abnormalities in the process of 3'-end formation have been linked to viral infection, tumor development, and spermatogenesis in human and animals. The activity of the Cleavage and Polyadenylation Specificity Factor (CPSF) complex is required for the cleavage and polyadenylation of most RNA polymerase II-synthesized pre-mRNA transcripts. Analysis of genome sequences from multicellular organisms shows that there are paralogues of some of the genes whose protein products comprise CPSF, but the role(s) these gene products play in mRNA 3'-end formation is very much unknown. The goal of this proposal is to understand how one of these paralogues (CPSF73-II) works to regulate 3'- end formation of mRNAs via a combination of biochemical and molecular genetic means using Arabidopsis as the model system. CPSF73-II encodes a protein with a domain similar to an endonuclease implicated in the cleavage of pre-mRNA and some histone mRNAs. Mutation analysis of the Arabidopsis atCPSF73-II gene demonstrates an essential role in female gamete and embryo development. Our hypothesis, based on extensive preliminary data, is that this group of proteins are involved in developmentally regulated mRNA 3'-end formation which is distinguishable from the constitutive one. To test this hypothesis, we propose two specific aims. The results of the research will uncover how this essential component of mRNA 3'-end formation regulates gene expression.
Aim 1 : To map the interacting domains of two atCPSF73 proteins with atCPSF100 subunit to elucidate the nature of the complex.
Aim 2 : To reveal target mRNAs which are subject to CPSF73-II activity using microarray technology. to Health Genes encoding a novel protein CPSF73-II that may play an essential role in the development are being studied. This protein is highly conserved among multicellular organisms. Built on the understanding of the gene function in the model system of Arabidopsis, the first goal of the project is to understand the proteins interaction partners, the results of which will reveal how it works in the cell. Using the state-of-the- art DNA microarray approach, the second goal is to uncover the targets that are regulated by the protein. The results of this research will be applicable to other higher eukaryotes, including humans, in understanding the gene's contributions in messenger RNA processing which is a critical step in gene expression and regulation. The latter is known to be relevant to many human health and disease control issues.
| Shen, Yingjia; Venu, R C; Nobuta, Kan et al. (2011) Transcriptome dynamics through alternative polyadenylation in developmental and environmental responses in plants revealed by deep sequencing. Genome Res 21:1478-86 |
| Xing, Denghui; Zhao, Hongwei; Li, Qingshun Quinn (2008) Arabidopsis CLP1-SIMILAR PROTEIN3, an ortholog of human polyadenylation factor CLP1, functions in gametophyte, embryo, and postembryonic development. Plant Physiol 148:2059-69 |
| Hunt, Arthur G; Xu, Ruqiang; Addepalli, Balasubrahmanyam et al. (2008) Arabidopsis mRNA polyadenylation machinery: comprehensive analysis of protein-protein interactions and gene expression profiling. BMC Genomics 9:220 |
| Xing, Denghui; Zhao, Hongwei; Xu, Ruqiang et al. (2008) Arabidopsis PCFS4, a homologue of yeast polyadenylation factor Pcf11p, regulates FCA alternative processing and promotes flowering time. Plant J 54:899-910 |
