Biochemical and Functional Characterization of GALT8, an Arabidopsis GT31 β-(1,3)-Galactosyltransferase That Influences Seedling Development

Joan Oñate Narciso; Wei Zeng; Kris Ford; Edwin R Lampugnani; John Humphries; Ingvild Austarheim; Allison van de Meene; Antony Bacic; Monika S Doblin

doi:10.3389/fpls.2021.678564

Biochemical and Functional Characterization of GALT8, an Arabidopsis GT31 β-(1,3)-Galactosyltransferase That Influences Seedling Development

Front Plant Sci. 2021 May 25:12:678564. doi: 10.3389/fpls.2021.678564. eCollection 2021.

Authors

Joan Oñate Narciso¹, Wei Zeng^{1

2}, Kris Ford¹, Edwin R Lampugnani¹, John Humphries¹, Ingvild Austarheim¹, Allison van de Meene¹, Antony Bacic^{1

2}, Monika S Doblin^{1

2}

Affiliations

¹ ARC Centre of Excellence on Plant Cell Walls, School of BioSciences, The University of Melbourne, Melbourne, VIC, Australia.
² Sino-Australia Plant Cell Wall Research Centre, State Key Laboratory of Subtropical Silviculture, School of Forestry and Biotechnology, Zhejiang A & F University, Hangzhou, China.

Abstract

Arabinogalactan-proteins (AGPs) are members of the hydroxyproline-rich glycoprotein (HRGP) superfamily, a group of highly diverse proteoglycans that are present in the cell wall, plasma membrane as well as secretions of almost all plants, with important roles in many developmental processes. The role of GALT8 (At1g22015), a Glycosyltransferase-31 (GT31) family member of the Carbohydrate-Active Enzyme database (CAZy), was examined by biochemical characterization and phenotypic analysis of a galt8 mutant line. To characterize its catalytic function, GALT8 was heterologously expressed in tobacco leaves and its enzymatic activity tested. GALT8 was shown to be a β-(1,3)-galactosyltransferase (GalT) that catalyzes the synthesis of a β-(1,3)-galactan, similar to the in vitro activity of KNS4/UPEX1 (At1g33430), a homologous GT31 member previously shown to have this activity. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) confirmed the products were of 2-6 degree of polymerisation (DP). Previous reporter studies showed that GALT8 is expressed in the central and synergid cells, from whence the micropylar endosperm originates after the fertilization of the central cell of the ovule. Homozygous mutants have multiple seedling phenotypes including significantly shorter hypocotyls and smaller leaf area compared to wild type (WT) that are attributable to defects in female gametophyte and/or endosperm development. KNS4/UPEX1 was shown to partially complement the galt8 mutant phenotypes in genetic complementation assays suggesting a similar but not identical role compared to GALT8 in β-(1,3)-galactan biosynthesis. Taken together, these data add further evidence of the important roles GT31 β-(1,3)-GalTs play in elaborating type II AGs that decorate AGPs and pectins, thereby imparting functional consequences on plant growth and development.

Keywords: Arabidopsis thaliana; GALT8; KNS4/UPEX1; arabinogalactan-proteins; galactosyltransferase; plant cell wall; seedling development.