Genetic Perturbation of the Starch Biosynthesis in Maize Endosperm Reveals Sugar-Responsive Gene Networks

Abstract

In maize, starch mutants have facilitated characterization of key genes involved in endosperm starch biosynthesis such as large subunit of AGPase Shrunken2 (Sh2) and isoamylase type DBE Sugary1 (Su1). While many starch biosynthesis enzymes have been characterized, the mechanisms of certain genes (including Sugary enhancer1) are yet undefined, and very little is understood about the regulation of starch biosynthesis. As a model, we utilize commercially important sweet corn mutations, sh2 and su1, to genetically perturb starch production in the endosperm. To characterize the transcriptomic response to starch mutations and identify potential regulators of this pathway, differential expression and coexpression network analysis was performed on near-isogenic lines (NILs) (wildtype, sh2, and su1) in six genetic backgrounds. Lines were grown in field conditions and kernels were sampled in consecutive developmental stages (blister stage at 14 days after pollination (DAP), milk stage at 21 DAP, and dent stage at 28 DAP). Kernels were dissected to separate embryo and pericarp from the endosperm tissue and 3′ RNA-seq libraries were prepared. Mutation of the Su1 gene led to minimal changes in the endosperm transcriptome. Responses to loss of sh2 function include increased expression of sugar (SWEET) transporters and of genes for ABA signaling. Key regulators of starch biosynthesis and grain filling were identified. Notably, this includes Class II trehalose 6-phosphate synthases, Hexokinase1, and Apetala2 transcription factor-like (AP2/ERF) transcription factors. Additionally, our results provide insight into the mechanism of Sugary enhancer1, suggesting a potential role in regulating GA signaling via GRAS transcription factor Scarecrow-like1.

Genetic variation for endosperm carbohydrates and total soluble solids in shrunken2, sugary1, waxy1, and wild‐type near‐isogenic corn lines across three harvest dates

“…The same effect was observed at 22 and 25 DAP (Table S4 -8). This finding agrees with Finegan et al (2022), who used the same set of NILs and reported C40 per se had higher total sugar levels at 21 DAP in su1 and WT NILs compared to C68. Although Finegan et al (2022) found no difference in total sugar between sh2 C40 and C68 NILs per se at 21 DAP, we observed a significant difference in total sugar content between these two sh2 NILs when used in hybrid combination at three harvest dates, 19, 22, and 25 DAP.…”

“…Experimental entries included four six-line half diallels with 15 hybrids each, or 60 total hybrids, generated by crossing four sets of six inbreds in a half-diallel mating design without parents per Griffing's Method 4, Model 1 (Griffing, 1956). The sets of inbreds include four NILs in six inbred genetic backgrounds, generated in the method described by Finegan et al (2022). The six inbreds were Connecticut 40 (C40), Connecticut 68 (C68), Iowa 453 (Ia453), Iowa 5125 (Ia5125), Illinois 101t (Il101t), and Purdue 39 (P39).…”

Genetic dissection of QTLs for starch content in four maize DH populations

“…This gene was also identified in a GWAS study and linked with natural variation in the amylose content (Li et al, 2018 ). Brittle2 ( Bt2 ) in qSC-2-1 encodes the small unit of AGPase, which catalyzes the upstream products converting into adenosine 5'diphosphate-glucose (ADPG), that is, the glucosyl donors of amylose and amylopectin (Preiss et al, 1990 ) and the first rate-limiting enzyme in the starch biosynthetic pathways (Ma et al, 2021 ; Finegan et al, 2022 ). ZMHXK3a in qSC-3-1, PHI1 in qSC-4-2 , and TPI3 in qSC-1-3 also play very important roles in glycolysis and starch metabolism.…”

Sugar loading of crop seeds – a partnership of phloem, plasmodesmal and membrane transport

“…The resulting T6P signal is transduced by an auxin‐regulated expression of starch biosynthetic genes (Weber et al ., 2005; Meitzel et al ., 2021). A reverse (feedback) mechanism has been discovered using mutants of starch biosynthesis in pea ( rugous – rrRbRb ; Zhou et al ., 2009) and maize ( AGPase Shrunken2 – Sh2 ; Finegan et al ., 2022). Elevated sucrose levels (pea cotyledons) and sucrose : hexose ratio (maize endosperm) in mutant filial storage tissues induce a titrated sugar‐induced reduction in PsSUT1 and ZmSUT1 expression.…”