Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis

Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis

Bibliographic Details
Main Author: Ajayi, Oyeyemi Olugbenga
Language:English
Published: Ohio University / OhioLINK 2021
Subjects:
Biology
Biochemistry
Plant Biology
Plant Sciences
Beta-Glucuronosyltransferase
Genes
Biosynthesis
Arabidopsis
Online Access:http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1625497309408518
id ndltd-OhioLink-oai-etd.ohiolink.edu-ohiou1625497309408518
record_format oai_dc
collection NDLTD
language English
sources NDLTD
topic Biology
Biochemistry
Plant Biology
Plant Sciences
Beta-Glucuronosyltransferase
Genes
Biosynthesis
Arabidopsis
spellingShingle Biology
Biochemistry
Plant Biology
Plant Sciences
Beta-Glucuronosyltransferase
Genes
Biosynthesis
Arabidopsis
Ajayi, Oyeyemi Olugbenga
Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis
author Ajayi, Oyeyemi Olugbenga
author_facet Ajayi, Oyeyemi Olugbenga
author_sort Ajayi, Oyeyemi Olugbenga
title Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis
title_short Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis
title_full Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis
title_fullStr Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis
title_full_unstemmed Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis
title_sort three beta-glucuronosyltransferase genes involved in arabinogalactan-protein biosynthesis and their roles in growth and development of arabidopsis
publisher Ohio University / OhioLINK
publishDate 2021
url http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1625497309408518
work_keys_str_mv AT ajayioyeyemiolugbenga threebetaglucuronosyltransferasegenesinvolvedinarabinogalactanproteinbiosynthesisandtheirrolesingrowthanddevelopmentofarabidopsis
_version_ 1719480783051161600
spelling ndltd-OhioLink-oai-etd.ohiolink.edu-ohiou16254973094085182021-09-16T05:10:20Z Three Beta-Glucuronosyltransferase Genes Involved in Arabinogalactan-Protein Biosynthesis and Their Roles in Growth and Development of Arabidopsis Ajayi, Oyeyemi Olugbenga Biology Biochemistry Plant Biology Plant Sciences Beta-Glucuronosyltransferase Genes Biosynthesis Arabidopsis Arabinogalactan-proteins (AGPs) are glycoproteins that function in plant growth and developmental processes, mainly through the actions of the heterogenous glycan chain attached to their protein cores. Glucuronic acid, which is transferred to the AGP glycan by β-glucuronosyltransferases (GLCATs), is the only acidic sugar in AGPs that binds calcium. Despite the importance of type II arabinogalactans (AGs), our understanding of the underlying biological role of such glycans and their sugar residues in plant growth is incomplete. To better understand the GLCATs, I carried out a comprehensive genomewide analysis of putative GLCAT gene family members belonging to the GlycosylTransferase14 (GT14) family in the Carbohydrate-Active enZYmes (CAZy) database by examining their sequence diversity, genetic architecture, phylogenetic and motif characteristics, selection pressure and gene expression in plants. I found 161 putative GLCAT genes distributed across 14 plant genomes with a widely conserved GLCAT catalytic domain. I discovered a phylogenetic clade shared between bryophytes and higher land plants of monocot grass and dicot lineages and identified positively selected sites that do not result in functional divergence of GLCATs. Also, RNA-seq and microarray data analyses of the putative GLCAT genes revealed gene expressionsignatures that likely influence the assembly of plant cell wall polymers. Among the eleven GLCAT genes found in Arabidopsis, I focused on identifying the biochemical and physiological roles of three of them, namely GLCAT14A, GLCAT14B and GLCAT14C. Based on in-silico analyses, I discovered that the GLCAT14A and GLCAT14C genes are highly expressed in both the seed coat and micropylar endosperm. Using a reverse genetics approach, I observed that glcat14a-1 mutants displayed subtle alterations in mucilage pectin homogalacturonan (HG) compared to WT, while glcat14a-1 glcat14c-1 double mutants displayed much more severe mucilage phenotypes, including loss of adherent mucilage and significant alterations in cellulose ray formation and seed coat morphology. Monosaccharide composition analysis showed significant alterations in the sugar amounts of glcat14a-1 glcat14c-1 mutants relative to WT with respect to both the adherent and non-adherent seed mucilage. A reduction in total mucilage content was also observed in glcat14a-1 glcat14c-1 mutants relative to WT. Moreover, glcat14a-1 glcat14c-1 mutants showed defects in pectin formation, calcium content and the degree of pectin methylesterification (DM) as well as reductions in crystalline cellulose content and seed size. I further determined that GLCAT14A, GLCAT14B and GLCAT14C were localized to the Golgi apparatus when transiently expressed in Nicotiana benthamiana. Sugar analysis of AGP extracts from Arabidopsis stem, leaf and siliques showed reductions in glucuronic acid in glcat14 mutants relative to WT, with concomitant effects resulting in tissue-specific alterations, especially with respect to arabinose and galactose sugars. Although I observed defects in trichome branching in glca14a/b and glcat14a/b/c mutants, scanning electron microscope/energy dispersive microanalysis (SEM/EDX) analyses showed no difference in the calcium content of the trichomes in these mutants relative to WT. Immunoblot analyses of stem and leaf showed a reduction in AGPs asdetected with the LM2 antibody in glcat14a/b and glcat14a/b/c mutants relative to WT. The biological activities of GLCATs are not limited to seed mucilage formation and plant vegetative growth, but are also important in Arabidopsis reproductive development, specifically in pollen development, polytubey block, and normal embryo development. Using biochemical and immunolabelling techniques, I found that the loss of function of GLCAT14A, GLCAT14B and GLCAT14C resulted in the disorganization of the reticulate structure of the pollen exine wall, with abnormal development of the intine layer, resulting in the collapse of pollen grains in glcat14a/b and glcat14a/b/c mutants relative to WT. Synchronous development between locules within the same anther was also lost in some glcat14a/b/c stamens, while some anther locules lacked pollen grains. Furthermore, I observed the excessive attraction of pollen tubes targeting glcat14a/b/c ovules, implying that the polytubey block mechanism was compromised. In contrast to WT, monosaccharide composition analysis revealed significant reductions in all sugars in glcat14a/b and glcat14a/b/c mutants with the exception of arabinose and galactose. Additionally, immunolabeling profiling using JIM13 and LM2 showed reduced amounts of AGP polysaccharides in glcat14a/b and glcat14a/b/c mutants relative to WT. The current work illustrates the benefits in conducting structure-function assessment of cell wall biosynthetic genes and their resulting activities (i.e., AG glucuronidation in this case) in plant growth, sexual reproduction and reproductive development of Arabidopsis. 2021-09-10 English text Ohio University / OhioLINK http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1625497309408518 http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1625497309408518 unrestricted This thesis or dissertation is protected by copyright: some rights reserved. It is licensed for use under a Creative Commons license. Specific terms and permissions are available from this document's record in the OhioLINK ETD Center.

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