No abstract

Trehalose 6-phosphate (T6P) is a sugar signal that regulates metabolism, growth, and development and inhibits the central regulatory SNF1-related protein kinase1 (SnRK1; AKIN10/AKIN11). To better understand the mechanism in wheat (Triticum aestivum) grain, we analyze T6P content and SnRK1 activities. T6P levels changed 178-fold 1 to 45 d after anthesis (DAA), correlating with sucrose content. T6P ranged from 78 nmol g 21 fresh weight (FW) pregrain filling, around 100-fold higher than previously reported in plants, to 0.4 nmol g 21 FW during the desiccation stage. In contrast, maximum SnRK1 activity changed only 3-fold but was inhibited strongly by T6P in vitro. To assess SnRK1 activity in vivo, homologs of SnRK1 marker genes in the wheat transcriptome were identified using Wheat Estimated Transcript Server. SnRK1-induced and -repressed marker genes were expressed differently pregrain filling compared to grain filling consistent with changes in T6P. To investigate this further maternal and filial tissues were compared pre-(7 DAA) and during grain filling (17 DAA). Strikingly, in vitro SnRK1 activity was similar in all tissues in contrast to large changes in tissue distribution of T6P. At 7 DAA T6P was 49 to 119 nmol g 21 FW in filial and maternal tissues sufficient to inhibit SnRK1; at 17 DAA T6P accumulation was almost exclusively endospermal (43 nmol g 21 FW) with 0.6 to 0.8 nmol T6P g 21 FW in embryo and pericarp. The data show a correlation between T6P and sucrose overall that belies a marked effect of tissue type and developmental stage on T6P content, consistent with tissuespecific regulation of SnRK1 by T6P in wheat grain.

During phosphate starvation, Snf1-related kinase 1 (SnRK1) activity significantly decreases compared with plants growing under normal nutritional conditions. An analysis of the expression of the genes encoding for the catalytic subunits of SnRK1 showed that these subunits were not affected by phosphate starvation. Transgenic Arabidopsis (Arabidopsis thaliana) plants overexpressing the AKIN10 and AKIN11 catalytic subunits fused with green fluorescent protein (GFP) were produced, and their localizations were mainly chloroplastic with low but detectable signals in the cytoplasm. These data were corroborated with an immunocytochemistry analysis using leaf and root sections with an anti-AKIN10/AKIN11 antibody. The SnRK1 activity in transgenic plants overexpressing AKIN11-GFP was reduced by 35% to 40% in phosphate starvation, in contrast with the results observed in plants overexpressing AKIN10-GFP, which increased the activity by 100%. No differences in activity were observed in plants growing in phosphate-sufficient conditions. Biochemical analysis of the proteins indicated that AKIN11 is specifically degraded under these limited conditions and that the increase in AKIN10-GFP activity was not due to the phosphorylation of threonine-175. These results are consistent with an important role of AKIN10 in signaling during phosphate starvation. Moreover, akin10 mutant plants were deficient in starch mobilization at night during inorganic phosphate starvation, and under this condition several genes were up-regulated and down-regulated, indicating their important roles in the control of general transcription. This finding reveals novel roles for the different catalytic subunits during phosphate starvation.