Knockout of a starch synthase gene OsSSIIIa/Flo5 causes white-core floury endosperm in rice (Oryza sativa L.)

Abstract

To elucidate the role of SSIIIa during starch synthesis in rice (Oryza sativa L.) endosperm, we characterized null mutants of this gene, generated by T-DNA insertions. Scanning electron microscope (SEM) analysis revealed that the starch granules in these mutants are smaller and rounder compared with the wild type controls, and that the mutant endosperm is characterized by a loosely packed central portion exhibiting a floury-like phenotype. Hence, the OsSSIIIa (Oryza sativa SSIIIa) mutations are referred to as white-core floury endosperm 5-1 (flo5-1) and flo5-2. Based upon their X-ray diffraction patterns, the crystallinity of the starch in the flo5 mutant endosperm is decreased compared with wild type. Through determination of the chain-length distribution of the mutant endosperm starch, we found that flo5-1 and flo5-2 mutants have reduced the content of long chains with degree of polymerization (DP) 30 or greater compared with the controls. This suggests that OsSSIIIa/Flo5 plays an important role in generating relatively long chains in rice endosperm. In addition, DP 6 to 8 and DP 16 to 20 appeared to be reduced in endosperm starch of flo5-1 and flo5-2, whereas DP 9 to 15 and DP 22 to 29 were increased in these mutants. By the use of differential scanning calorimetry (DSC), the gelatinization temperatures of endosperm starch were found to be 1-5 degrees C lower than those of the control. We propose a distinct role for OsSSIIIa/Flo5 and the coordinated action of other SS isoforms during starch synthesis in the seed endosperm of rice.

Gradually Decreasing Starch Branching Enzyme Expression Is Responsible for the Formation of Heterogeneous Starch Granules

“…For example, once the soluble starch synthase OsSSIIIa/Flo5, which is responsible for the extension of relatively long chains of both amylopectin B2 and B3 to B4 (Gao et al, 1998;Jane et al, 1999;Fujita et al, 2006), is eliminated from the genome, a phenotype of loosely packed central portions exhibiting a floury white-core endosperm is detected. The granules located in this region are smaller than those of the parental control and exhibit round morphology with no sharp edges, while the peripheral region in ssIIIa/flo5 contains normal compound starch (Ryoo et al, 2007). A similar phenotype was identified in previously reported knockouts of BT1, RSR1, and FLO4, which encode an ADP-Glc transporter, APETALA2/ethylene-responsive element-binding protein family transcription factor, and pyruvate orthophosphate dikinase, respectively (Kang et al, 2005;Fu and Xue, 2010;Li et al, 2017).…”

Studies on rice seed quality through analysis of a large-scale T-DNA insertion population

“…This may be due to the fact that amylose constitutes only up to ∼20% of the total starch, and mutants with altered amylose may not necessarily result in significant changes in total starch. Some mutations are found in genes involved in the individual metabolism pathway, suggesting that seed quality can be regulated by directly modifying the genes involved in the synthesis or metabolism pathways of starch, fat and so on [41][42][43]. Additionally, other genes fall into many other diverse molecular function categories and some genes are associated with more than one changed constituents of seeds, indicating the close relationship between the regulation of synthesis and metabolism of these constituents.…”

“…These resources helped to identify and study genes involved in rice development [40,41]. Some mutants with marked changes of seed quality were also identified [42,43].…”

“…These resources helped to identify and study genes involved in rice development [40,41]. Some mutants with marked changes of seed quality were also identified [42,43].To facilitate the coordination of rice functional genomic studies, especially that on seed quality regulation, we have developed the Shanghai T-DNA Insertion Population (SHIP, http://ship.plantsignal.cn) of rice by large-scale Agrobacterium-mediated transformation. The flanking sequences of T-DNAs were identified by thermal asymmetric interlaced-PCR (TAIL-PCR) [44,45] and mapped onto the rice genome.…”

Recent progress toward understanding the role of starch biosynthetic enzymes in the cereal endosperm

“…SSIII has two genes responsible for its expression in the endosperms and leaves of rice. Mutants of SSIII in maize and rice display altered granule morphology and crystallinity, and a reduction in long-branch chains extending between clusters [59][60]. Loss of SSIII has also been associated with pleiotropic effects affecting other SSs, where SSs have an increase in activity, while SBEIIa activity is decreased.…”