The analysis of the different functions of starch‐phosphorylating enzymes during the development of Arabidopsis thaliana plants discloses an unexpected role for the cytosolic isoform GWD2

Abstract

The genome of Arabidopsis thaliana encodes three glucan, water dikinases. Glucan, water dikinase 1 (GWD1; EC 2.7.9.4) and phosphoglucan, water dikinase (PWD; EC 2.7.9.5) are chloroplastic enzymes, while glucan, water dikinase 2 (GWD2) is cytosolic. Both GWDs and PWD catalyze the addition of phosphate groups to amylopectin chains at the surface of starch granules, changing its physicochemical properties. As a result, GWD1 and PWD have a positive effect on transitory starch degradation at night. Because of its cytosolic localization, GWD2 does not have the same effect. Single T-DNA mutants of either GWD1 or PWD or GWD2 have been analyzed during the entire life cycle of A. thaliana. We report that the three dikinases are all important for proper seed development. Seeds from gwd2 mutants are shrunken, with the epidermal cells of the seed coat irregularly shaped. Moreover, gwd2 seeds contain a lower lipid to protein ratio and are impaired in germination. Similar seed phenotypes were observed in pwd and gwd1 mutants, except for the normal morphology of epidermal cells in gwd1 seed coats. The gwd1, pwd and gwd2 mutants were also very similar in growth and flowering time when grown under continuous light and all three behaved differently from wild-type plants. Besides pinpointing a novel role of GWD2 and PWD in seed development, this analysis suggests that the phenotypic features of the dikinase mutants in A. thaliana cannot be explained solely in terms of defects in leaf starch degradation at night.

Redox Regulation of Starch Metabolism

“…Plants accumulate starch as both a transient and long-term carbohydrate reserve. As a result, starch metabolism must be adjusted to provide ample carbon supply in response to many physiological demands mainly related to nocturnal, stress, and germination events ( Stitt and Zeeman, 2012 ; Lloyd and Kossmann, 2015 ; Hedhly et al, 2016 ; Zanella et al, 2016 ; Pirone et al, 2017 ; Thalmann and Santelia, 2017 ). Coordinated metabolic flux among starch biosynthetic enzymes can also permit correct structuring of the starch granule irrespectively of carbon flow ( Blennow and Svensson, 2010 ; Glaring et al, 2012 ; Pfister and Zeeman, 2016 ; Botticella et al, 2018 ).…”

Differential Evolution of α-Glucan Water Dikinase (GWD) in Plants

“…This location at the N-terminus in MSA was also the least conserved region, giving an indication of the intense selection pressure in the evolution of the current function of transit peptides. The absence of transit peptides in GWD2 in the plant species in which it is present supports the fact that they are localized in the cytosol [ 8 ]. Finally, recent studies have established that cTP and mTP contain specific motifs [ 56 ] and amino acid residues [ 57 , 58 ], with which they either interact with cytosolic complexes containing Hsp70 and Hsp90 (molecular chaperones that target transit peptides and guide the preprotein to the outer membranes of the organelles) or TOC–TIC (chloroplast)/TOM–TIM (mitochondria) protein import machinery [ 59 , 60 ].…”

“…A different functional evolution may exist regarding PWD, another paralog of GWD, as described [ 2 ], but not the focus of current study. In M. esculenta , GWD2 and the other GWD isoforms were implicated by Zhou et al [ 45 ] in storage starch degradation, due to increased expressions during post-harvest physiological deterioration where its activity was linked to seed development in A. thaliana and not the degradation of transitory starch at night [ 8 ]. However, gain of GWD2 in several plant families points to another unique event during GWD evolution.…”

“…While less is known about this phenomenon, the model for plastidial starch degradation starts with its phosphorylation, orchestrated by the activities of GWD1 and PWD [ 2 ]. In addition to the pervasive GWD isoforms, another isoform, GWD2, uniquely localized in the cytosol, has been identified [ 8 ]. Mainly because of its cytosolic localization, GWD2 has not been associated with transitory starch degradation [ 8 ].…”

“…In addition to the pervasive GWD isoforms, another isoform, GWD2, uniquely localized in the cytosol, has been identified [ 8 ]. Mainly because of its cytosolic localization, GWD2 has not been associated with transitory starch degradation [ 8 ].…”