mechanisms that govern pattern formation during plant development
	Vivian Irish, Ph.D. Associate Professor of Molecular, Cellular & Developmental Biology and Ecology & Evolutionary Biology Email: vivian.irish@yale.edu Room: OML 252A Phone: (203) 432-5572/ (203) 432-5571 Web site B.A. Wesleyan University 1980; Ph.D. Harvard University 1986

A wild type Arabidopsis flower.

AP3 expression in the Arabidopsis inflorescence as visualized by an AP3 promoter:GUS reporter gene. AP3 expression is first seen in very young flower buds as a ring encircling the meristem corresponding to the presumptive petal and stamen primordia; later in flower development, AP3 expression is seen in these organs as they mature. Lower levels of expression appear pink, higher levels of expression appear blue. We are also investigating the evolution of molecular genetic mechanisms controlling floral form in other plant species. Normally poppy flowers contain four petals, but in this poppy homeotic mutant, some stamens are converted to petals, resulting in multiple extra petals.

Flowers have a very regular architecture, yet floral form varies immensely between different species. We are interested in understanding how the stereotypical floral pattern arises, as well as how these developmental processes have been modulated in different plant species. We are utilizing Arabidopsis for many of these investigations; Arabidopsis has a short generation time, it can be transformed and genetically manipulated, and the Arabidopsis genome is the first plant genome to be completely sequenced, making it an ideal system in which to study pattern formation.

We are focussing on analyzing two homeotic genes, APETALA3 (AP3) and PISTILLATA (PI), which are required for specifying petal and stamen identities in the Arabidopsis flower. AP3 and PI both encode MADS-box containing transcription factors, and bind to DNA as an obligate heterodimer. We have shown that the AP3 and PI function is regulated at the post-transcriptional level, since these proteins need to interact to be effectively localized to the nucleus, a process which may be mediated by heterodimerization. We are also examining the regulation of AP3 expression, and have defined specific promoter elements that are required for different temporal and spatial aspects of AP3 transcription. Using molecular, genetic, and biochemical techniques, we are now in the process of identifying trans-acting factors that are required to activate AP3 expression in the petal and stamen primordia. These factors include LEAFY, a master transcriptional regulator of flowering, which acts in part by directly binding to sequences in the AP3 promoter and activating its expression. In addition, other experiments are aimed at identifying new genes that are required for initiation of flowering and floral organ development.

We are also examining the role that these floral homeotic gene products play in regulating the specification of petal and stamen cell types. We are taking several approaches, including microarray experiments, genetic screens and biochemical assays to defining the populations of genes regulated by the AP3-PI heterodimeric transcription factor. Microarray analyses have defined a relatively small group of genes that are specifically involved in petal and stamen cell type differentiation, and we are currently analyzing the roles of these genes in such processes. In addition, various genetic screens have yielded several intriguing candidate genes that appear to be required for petal and/or stamen differentiation.

In addition to our work on Arabidopsis, we are interested in understanding the underlying basis of the tremendous variability in floral form in the angiosperms. To this end, we are characterizing homologs of the AP3 and PI genes from a variety of other species. We have shown that there are variations in the expression patterns of AP3 and PI orthologs from other angiosperms. We are currently carrying out functional analyses in several other angiosperm species, which should tell us the degree to which the developmental circuitry specifying floral organ identity is similar among species. These studies are part of a larger effort to characterize the changing roles of MADS domain containing transcription factors in regulating floral development in different angiosperm species.

By combining both genetic and molecular approaches to the study of floral development, we hope to elucidate how dividing floral meristematic cells acquire information about their position and then differentiate accordingly, as well as how these processes may have been modulated during evolution.

Selected Publications

Lamb, R.S., Hill, T.A., Tan, Q.K-G., and V.F. Irish. (2002) Regulation of APETALA3 floral homeotic gene expression by meristem identity genes. Development, 129: 2079-2086.

Zik, M., and Irish, V.F. (2003). Global identification of target genes regulated by APETALA3 and PISTILLATA floral homeotic gene action. Plant Cell, 15: 207-222.

Lamb, R.S. and Irish, V.F. (2003). Functional divergence within the APETALA3/PISTILLATA floral homeotic gene lineages. Proc. Nat. Acad. Sci. in press.

			The Arabidopsis AP3 and PI proteins are both required for petal and stamen identity and function as a heterodimeric transcription factor. A. wild type Arabidopsis flower. B. apetala3 (ap3) mutant in which petals are converted to sepals and stamens are converted to carpels. C. pistillata (pi) mutant displays the same homeotic muant phenotype as ap3 mutants. D. AP3 and PI proteins must form a heterodimer to bind to DNA. In this gel-shift assay, AP3 or PI proteins cannot bind to DNA independently, but when mixed together, can bind to target DNA sequences, resulting in a band shift. Because both proteins are required for function, mutation in either AP3 or PI results in the same homeotic transformation.

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