Starch synthases SSIIa and GBSSI control starch structure but do not determine starch granule morphology in the absence of SSIIIa and SSIVb.

key messageHigh levels of two major starch synthases, SSIIa and GBSSI, in ss3a ss4b double mutant rice alter the starch structure but fail to recover the polygonal starch granule morphology. The endosperm starch granule is polygonal in wild-type rice but spherical in double mutant japonica rice lacking genes encoding two of the five major Starch synthase (SS) isozymes expressed in endosperm, SSIIIa and SSIVb. Japonica rice naturally has low levels of SSIIa and Granule-bound SSI (GBSSI). Therefore, introduction of active SSIIa allele and/or high-expressing GBSSI allele from indica rice into the japonica rice mutant lacking SS isozymes can help elucidate the compensatory roles of SS isozymes in starch biosynthesis. In this study, we crossed the ss3a ss4a double mutant japonica rice with the indica rice to generate three new rice lines with high and/or low SSIIa and GBSSI levels, and examined their starch structure, physicochemical properties, and levels of other starch biosynthetic enzymes. Lines with high SSIIa levels showed more SSI and SSIIa bound to starch granule, reduced levels of short amylopectin chains (7 ≤ DP ≤ 12), increased levels of amylopectin chains with DP > 13, and consequently higher gelatinization temperature. Lines with high GBSSI levels showed an increase in amylose content. The ADP-glucose content of the crude extract was high in lines with low or high SSIIa and low GBSSI levels, but was low in lines with high GBSSI. Addition of high SSIIa and GBSSI altered the starch structure and physicochemical properties but did not affect the starch granule morphology, confirming that SSIIIa and SSIVb are key enzymes affecting starch granule morphology in rice. The relationship among SS isozymes and its effect on the amount of substrate (ADP-glucose) is discussed.