Wheat E3 ubiquitin ligase TaGW2-6A degrades TaAGPS to affect seed size

TaGW2 has been identified as a key determinant of the grain weight in wheat (Triticum aestivum L.). In our previous study, we found that the grain size differs in Chinese Spring (CS) and its TaGW2-6A allelic variant (NIL31). In addition, the expression of the key starch biosynthesis enzyme gene TaAGPS differs significantly in the two materials. However, the underlying molecular mechanism associated with the action of TaGW2-6A has not been reported. In the present study, we found that TaGW2-6A-CS interacted with TaAGPS, whereas TaGW2-6A-NIL31 did not interact with it in vitro and in vivo. Furthermore, we found that the C-terminal LXLX domain (376-424 aa) of TaGW2-6A recognized TaAGPS. However, the TaGW2-6A allelic variant lacked this key interaction region due to premature translation termination. We also found that TaGW2-6A-CS can ubiquitinate TaAGPS and degrade it via the 26 S proteasome pathway. In addition, our analysis of the activity of ADP-glucose pyrophosphorylase (AGPase) indicated that the AGPase level in the endosperm cells was lower in CS than NIL31. Cytological observations demonstrated that the average number of starch granules and the average area of starch granules in endosperm cells were lower in CS than NIL31. The overexpression of TaAGPS positively regulated the seed size in transgenic Arabidopsis. Our findings provide novel insights into the molecular mechanism that allows TaGW2-6A-TaAGPS to regulate seed size via the starch synthesis pathway.