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United States Department of Agriculture

Agricultural Research Service

2011 Annual Report

1a.Objectives (from AD-416)
The long-term goal of this research is to develop the ability to regulate pollen development and pollen-pistil interactions. Using yeast two hybrid screens, as well as genomic and proteomic tools, we have identified candidate proteins that play roles during these processes, and have developed tools (such as cell-specific promoters) that can facilitate functional studies of these proteins. The specific objectives of this project plan are:

1. To elucidate the molecular interactions that occur during pollen development, pollen-pistil interactions, and sperm-egg interactions, using both model (e.g., Arabidopsis) and crop (e.g., tomato, corn) species. [NP 301, C 4, PS 4A]

2. To define the mechanisms underlying hybridization barriers between species and determine if manipulating gene expression levels of key genes can overcome such barriers. [NP 301, C 4, PS 4A]

1b.Approach (from AD-416)
1) Use yeast 2-hybrid screens and biochemical techniques to characterize proteins expressed in female tissues and/or in pollen that are required for signaling through receptor kinases. Use similar techniques to identify sperm- and egg-expressed proteins that might mediate egg-sperm interactions. Develop sperm- and egg-specific promoter elements for gamete-specific gene manipulations. Use comparative genomics to identify protein domains important for specificity. Use gene disruptions to test interactions genetically.

2) Use pollination assays and microscopic imaging to determine where hybridization barriers occur between selected plant species pairs. Identify target genes that might act at these steps. Generate gene disruptions for selected candidates and assay for pollen tube growth/fertilization phenotypes. Introduce DNA constructs into transgenic plants. Use pollination assays to test for expanded hybridization potential in plants that overexpress or repress the expression of selected target genes.

3.Progress Report
Discovered that a protein complex already known to target proteins for degradation plays another role, by regulating the transcription of genes whose encoded proteins are important for cell divisions. Demonstrated that a gamete-expressed protein plays an important role in early embryo development. Demonstrated that a protein phosphatase regulates protein phosphoryation during pollen tube growth, and that perturbing the balance of phosphorylation leads to autophagy (self-eating).

1. The anaphase promoting complex (APC) has dual roles in regulating the cell cycle. For cell division to occur, it is important to remove proteins called cyclins, as their continued presence prevents cell cycle progression. A protein complex called the anaphase promoting complex/cyclosome (or APC/C) was known to target cyclins for degradation. A new role for the APC/C was discovered by ARS scientists in Albany, CA by analyzing the cell divisions that occur during pollen development. The APC/C also regulates the transcription of the cyclin gene, and it does this by interacting with a microRNA that targets a key regulator of pollen cell division. This is the first report that the APC also regulates gene transciption.

Review Publications
Zheng, B., Chen, X., Mccormick, S.M. 2011. The Anaphase-Promoting Complex is a dual integrator that regulates both microRNA-mediated transcriptional regulation of Cyclin B1 and degradation of Cyclin B1 during Arabidopsis male gametophyte development. The Plant Cell. tpc.111.083980.

Last Modified: 10/25/2014
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