Browsing by Subject "Plant development"
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Item Characterization of Glabra2 and Transparent Testa Glabra2, targets of the TTG1 complex(2010-08) Hatlestad, Gregory James; Lloyd, Alan M.; Roux, Stanley; Huq, Enamul; Mehdy, Mona; Juenger, ThomasStudies on epidermal cell fate determination have been important for gaining insight into the genetic and molecular mechanisms leading to the differentiation and patterning of cells. In Arabidopsis, the organization and development of many epidermal characters including trichomes, root hairs and the seed coat have been found to be controlled by a single combinatorial transcription factor complex consisting of a WD-repeat containing protein, Transparent Testa Glabra 1 (TTG1), and various MYB and bHLH proteins. The work here consists of identification of Glabra2 (GL2) and Transparent Testa Glabra2 (TTG2) as direct transcriptional targets of the TTG1 combinatorial complex, further characterization of GL2 function, and identification of transcriptional targets of GL2 and TTG2. Both GL2 and TTG2 are important in the regulation of trichomes, root hairs and seed coat development. vii GL2 has been identified as an important regulator of epidermal cell fate for over fifteen years yet there is little known about its function and only three transcriptional targets are identified, all involved in root hair patterning. Through the examination of its function a nuclear localization signal was verified and shown that GL2 homodimerizes. Through analysis of available expression databases and differential sequence analysis using SOLiD sequencing technology, several direct targets of GL2 and many more possible transcriptional targets of both GL2 and TTG2 were identified in trichomes. Some of these targets are members of the TTG1 complex, and they are all specialized in the maturation of trichomes, suggesting that GL2 switches the focus of the complex by activating the TTG1 complex members involved in maturation of the trichome through a feedback mechanism. Examination of gl2 mutants shows that they do not produce trichome accessory cells which usually surround the trichome. An additional target of GL2 is At5g65300 which when overexpressed results in the elongation and proliferation of trichome accessory cells into a tall pillar of cells. This suggests that GL2 is involved in the regulation of accessory cell development through At5g65300. The work presented here represents important advances of our knowledge of epidermal cell fate through characterization of the major downstream regulators of epidermal development.Item Knowing them by knocking them out : initial studies to characterize Arabidopsis annexin gene family function(s)(2007-05) Cantero-Garcia, Maria Araceli, 1967-; Roux, Stanley J.Annexins are a multigene family in most plant species and are suggested to play a role in wide variety of essential cellular processes. Analysis of paralogous blocks within the genome of Arabidopsis showed that AnnAt6 was a duplicated gene from AnnAt1 and AnnAt2. The motif analysis of the promoters of these three annexins indicated some putative functional redundancy but also agreed with the model that duplicated genes functionally diverge over time and undergo functional specialization that can principally be observed in their regulatory regions. This in silico analysis contributes important information which can be used to guide phenotypic analysis of mutants. The use of quantitative real time reverse transcription PCR (quantitative real time RT-PCR) to assess differential expression of annexins in different tissues from 5.5 day-old etiolated seedlings showed a trend of major expression of annexins in root and hypocotyl tissues compared to cotyledon tissue. This result supports previous data about the participation of annexins during growth and development. The light-dependent regulation of annexin gene expression was also examined using etiolated seedlings under red and far-red light conditions by quantitative real time RT-PCR. Red light treatment significantly altered transcript levels in hypocotyls and cotyledons for AnnAt5 and AnnAt6, respectively, and this early up-regulation was reversible by far-red light, implicating phytochrome as the receptor mediating this response. In order to begin testing the hypothesis that plant annexins have individual functions, over thirty annexin putative T-DNA insertional lines were obtained and screened. After a screen of these mutants, further work was focused on fourteen lines for which annexin insertion sites were confirmed. For AnnAt1 through AnnAt5 at least two allelic lines and for AnnAt6 and AnnAt7 one allelic line has been confirmed by PCR. For some of them the absence of mRNA was determined by RT-PCR which confirms that they are true knockouts. These annexin T-DNA insertion lines will allow for future phenotypic characterization and likely provide valuable information regarding individual functions of Arabidopsis annexins.