Rouffle, M. | bioRxiv Science

bioRxiv2023

TBL38 is an atypical homogalacturonan acetylesterase with a peculiar cell wall microdomain localization in Arabidopsis seed mucilage secretory cells

Plant cell walls are made of complex polysaccharidic/proteinaceous network whose biosynthesis and dynamics implicate several cell compartments and impact plant development. The synthesis and remodeling of homogalacturonan pectins is associated with multiple developmental processes ranging from growth to response to biotic/abiotic stress. It encompasses Golgi-localized methylation and acetylation and subsequent demethylation and deacetylation in the cell wall. In the last decade, our comprehension of plant polysaccharides acetylation has increased significantly thanks to the study of the TRICHOME BIREFRINGENCE-LIKE (TBL) protein family. TBLs are mostly described as Golgi-localized acetyltransferases specifically targeting diverse hemicelluloses or pectins. Various tbl mutants showed altered wall mechanical properties and dynamics. Here, we study TBL38 that is co-expressed with PECTIN METHYLESTERASE INHIBITOR6 (PMEI6) and PEROXIDASE 36 (PRX36) during the development of Arabidopsis seed mucilage secretory cells (MSCs). We demonstrate the atypical TBL38 cell wall localization restricted to the PMEI6/PRX36 MSC cell wall microdomain. A tbl38 mutant displays an intriguing homogalacturonan immunological phenotype in this cell wall microdomain and in a MSC surface-enriched abrasion powder. This fraction was further characterized by mass spectrometry oligosaccharide profiling revealing an increased homogalacturonan acetylation phenotype. Finally, a recombinant TBL38 is shown to display pectin acetylesterase activity in vitro. These results indicate that TBL38 is an atypical cell wall-localized TBL that displays a homogalacturonan acetylesterase activity rather than a Golgi-localized acetyltransferase activity as observed in previously studied TBLs. TBL38 function during seed development is discussed.

plant biology↗

bioRxiv2022

Differences in the structure of plant polygalacturonases specify enzymes dynamics and processivities to fine-tune pectins and root development

The fine-tuning of pectins by polygalacturonases (PGs) plays a key role in modulating plant cell wall chemistry and mechanics, impacting plant development. In plants, the high number of PGs encoded in the genome questions the regulation of pectin depolymerization and the roles of distinct isozymes in the control of development. Here we report the first crystal structures of two PGs from Arabidopsis, PGLR and ADPG2 whose expression overlap in roots. Albeit having overall conserved folds and active sites, PGLR and ADPG2 differed in the structure of their binding grooves and in the amino-acids of the subsites. We determined the structural features that explain the absence of inhibition of the plant PGs by endogenous PG-Inhibiting Proteins (PGIPs). By combining molecular dynamic simulations, analysis of enzymes kinetics and hydrolysis products, we showed that subtle differences in PGLR and ADPG2 structures translated into distinct enzyme-substrate dynamics and enzymes processivities. Using the plant root as a developmental model, exogenous application of purified enzymes showed that these distinct PGLR/ADPG2 processivities ultimately translated into different impacts on development. The highly processive ADPG2 had major effects on both root cell elongation and cell adhesion. Our study suggests that, in plants, gene redundancy is unlikely to reflect redundant biochemical specificities. Isozymes of distinct specificities and processivities are likely to be of major importance for the fine spatial and temporal regulation of pectin structure. Significance StatementPlant polygalacturonases (PG) are enzymes that play a key role in the regulation of cell wall pectin chemistry by controlling the degree of polymerization of the HG chains. The high number of genes encoding PG in Arabidopsis questions the rationale for such abundance. We solved the crystal structure of two PG (PGLR and ADPG2) whose expression overlap in roots and showed, using combined computational and experimental approaches, that they differ in their enzyme-substrate dynamics, leading to distinct processivities. The highly processive ADPG2 can generate digestion products of shorter degree of polymerization, and upon exogenous application on developing roots, induced drastic developmental defects. Our study suggests that gene redundancy is unlikely to reflect redundant biochemical specificities of isozymes.

biochemistry↗