Characterization and functional analysis of arabinogalactan protein 31 in Arabidopsis
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Arabinogalactan proteins (AGPs) are highly glycosylated cell wall proteins specific to plants. AGPs have been implicated in almost all aspects of plant development and defense responses, nevertheless, most of such studies are correlative. To define the specific functions of individual AGPs, direct evidence from analyses of genetic knockout mutants of individual AGPs is required. Up to now, only a few AGPs have been demonstrated to have defined functions by mutant analyses. This dissertation identified a non-classical AGP (AGP31), described its expression and characterized the null mutant of AGP31 in Arabidopsis. In agp31 mutant, microarray analyses revealed that the expressions of genes encoding a subset of seed storage proteins (SSP): CRU3, CRA1 and OLEOSIN2 were induced. Further analysis showed that induction by agp31 knockout was specific to these three SSP genes, indicating a novel pathway to regulate the SSP gene expression. Comprehensive characterizations of AGP31 were carried out. Yariv reagent staining and monosaccharide analysis of purified AGP31 showed that AGP31 was a bona fide galactose-rich AGP. The cell wall localization of AGP31 was confirmed by expression of an AGP31::eGFP fusion protein. AGP31 promoter-GUS reporter gene analysis showed that AGP31 was expressed in the vascular bundle throughout the plant, except in the flower. In the flower, it was expressed throughout the pistil except in the stigma. Detailed analysis showed that GUS staining occurred in all cell types in the vascular bundle of roots, while GUS staining was restricted to phloem cells in the inflorescence stem. AGP31 mRNA was down-regulated by several stress treatments, including wounding, methyl jasmonic acid (MeJA) and abscisic acid (ABA). In response to MeJA treatment of whole seedlings, AGP31 mRNA level decreased to about 30% of its original level within 8 hr and almost returned to its original level after 24 hr. Nuclei run-on assay showed that the down-regulation of AGP31 mRNA upon MeJA treatment was due to reduced transcription. The strong preferential expression in vascular tissues and negative regulations by MeJA and ABA suggest that AGP31 may be involved in vascular tissue function both during development and the defense response.