2009 |
Xiao, Wenyan |
R15Activity Code Description: Supports small-scale research projects at educational institutions that provide baccalaureate or advanced degrees for a significant number of the Nation’s research scientists but that have not been major recipients of NIH support. The goals of the program are to (1) support meritorious research, (2) expose students to research, and (3) strengthen the research environment of the institution. Awards provide limited Direct Costs, plus applicable F&A costs, for periods not to exceed 36 months. This activity code uses multi-year funding authority; however, OER approval is NOT needed prior to an IC using this activity code. |
Dme-Interacting Protein 1 Regulates Demethylatoin and Reproduction in Arabidopsis
DESCRIPTION (provided by applicant): Cytosine methylation is one of the major mechanisms to silence transposons and retroviruses, control genomic imprinting, and regulate reproduction in both mammals and plants. Altered DNA methylation is associated with diseases including leukemia, cancer and congenital syndromes in humans. Arabidopsis thaliana is an excellent system to investigate epigenetic mechanisms, which are evolutionarily conserved between mammals and plants. In Arabidopsis, DEMETER (DME) belongs to a family of DNA glycosylases that excise damaged or mispaired bases in the base excision DNA repair. DME demethylates and activates the maternal allele of imprinted genes. However, one key question is not answered: how do DME or DML DNA glycosylases find their targets to demethylate in the genome? Recently, we isolated DME-INTERACTING PROTEIN 1 (DIP1) in the yeast two-hybrid screen. DIP1 is a putative ubiquitin-conjugating enzyme 7-interacting protein 4 that has a RING domain. Interestingly, we found 50% seed abortion in the self-pollinated progeny of the DIP1/dip1 heterozygous plant. The seed inheriting a maternal dip1 mutant allele is aborted when the DIP1/dip1 female plant was crossed with the wild-type male. To better understand how DIP1 regulates DNA demethylation and imprinting, the following specific aims will be executed: 1) Determine if DIP1 binds to the promoter of imprinted genes and targets DME to demethylate DNA. Using the gel shift assay, we will examine if DIP1 binds to the promoter of imprinted MEDEA (MEA), a Polycomb gene. We will investigate if DIP1 regulates MEA expression, imprinting, and methylation. 2) Analyze molecular functions of DIP1. We will examine if the RING domain protein DIP1 has an E3 ubiquitin ligase activity, and if so, whether DME and DMLs are substrates for the DIP1-mediated ubiquitin proteolytic pathway. 3) Elucidate the mechanism used by DIP1 in regulating DNA demethylation and gene expression. We will explore if DIP1 regulates MEA imprinting through ubiquitination of another DME-interacting protein, histone H1, an event that might lead to DNA methylation and/or histone H3 methylation specifically in the MEA promoter. The proposed experiments carried out using the genetically tractable Arabidopsis system will elucidate mechanisms that link ubiquitination via a RING domain protein to DNA demethylation, gene imprinting, and gene expression - processes that are evolutionarily conserved and integral to the pathology of human diseases such as cancer. Mechanisms amenable to specific targeting of DNA demethylation will provide valuable insights for the development of future gene therapies for targeting specific DNA hypermethylation loci in human diseases. This grant will be used to train undergraduate and graduate students for scientific discovery in epigenetics and molecular biology at Saint Louis University. PI will participate actively in outreach efforts to foster scientific training and education programs in grades K-12 in St. Louis. PUBLIC HEALTH RELEVANCE The proposed specific aims executed using the epigenetically tractable Arabidopsis thaliana system will elucidate mechanisms that link ubiquitination via a RING domain protein to DNA demethylation, genomic imprinting, and gene expression - processes that are evolutionarily conserved and integral to the pathology of human diseases such as cancer, leukemia, and congenital syndromes. Mechanisms amenable to specific targeting of DNA demethylation will provide significant insights for the development of future gene therapies for targeting specific DNA hypermethylation loci in human diseases.
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0.954 |
2017 — 2020 |
Xiao, Wenyan Hsieh, Tzung Fu |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Elucidating the Molecular Mechanism of Demeter-Mediated Active Dna Demethylation @ North Carolina State University
The goal of this project is to advance the understanding of how genes important for nutrient accumulation in plant seeds are regulated. Seed production is vital for agriculture, biofuels, and human nutrition. Cereal grains account for over 50% of the world's dietary energy consumption and most of the grain nutrients are stored in a seed compartment called the endosperm. The project uses genetic, genomic, and computational tools to address fundamental seed biology questions, and provides interdisciplinary education and training opportunities for undergraduates, graduate students, and postdoctoral researchers. Undergraduate students will spend three summers in the laboratory and participate directly in the research activities in plant biology. This project will provide critical research experience for the undergraduates and important training for the graduate student and the postdoctoral fellow, thus developing a diverse, globally competitive workforce in this field.
Genomic imprinting influences seed yield by controlling resource allocation to the endosperm where the synthesis and storage of protein, starch, and lipid nutrients occurs. In flowering plants gene imprinting is established by DEMETER (DME) mediated active DNA demethylation. DME encodes a large polypeptide with multiple conserved domains, and except for the well-characterized glycosylase domain, very little is known about the function of the other domains. Elucidating how DME is regulated and directed to proper genomic locations is crucial to advance the knowledge on seed development, and can inspire development of novel molecular strategies for breeding or engineering desirable traits in important crop plants. This project aims to elucidate the roles of these conserved domains on DME activity and seed viability through molecular, genetic, genomic and epigenomic analyses. A novel bipartite model for structural and functional regulation for DME activity is proposed. Understanding why DME adopts modular catalytic and regulatory domain architecture and elucidating how linker histone H1 assists DME mediated active DNA demethylation will significantly increase the understanding of how epigenetic information is established and maintained in plants.
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0.963 |