Genetic mapping and functional analysis of a classical tassel branch number mutant Tp2 in maize

Abstract

Tassel branch number is a key trait that contributes greatly to grain yield in maize (Zea mays). We obtained a classical mutant from maize genetics cooperation stock center, Teopod2 (Tp2), which exhibits severely decreased tassel branch. We conducted a comprehensive study, including phenotypic investigation, genetic mapping, transcriptome analysis, overexpression and CRISPR knock-out, and tsCUT&Tag of Tp2 gene for the molecular dissection of Tp2 mutant. Phenotypic investigation showed that it is a pleiotropic dominant mutant, which is mapped to an interval of approximately 139-kb on Chromosome 10 harboring two genes Zm00001d025786 and zma-miR156h. Transcriptome analysis showed that the relative expression level of zma-miR156h was significantly increased in mutants. Meanwhile, overexpression of zma-miR156h and knockout materials of ZmSBP13 exhibited significantly decreased tassel branch number, a similar phenotype with Tp2 mutant, suggesting that zma-miR156h is the causal gene of Tp2 and targets ZmSBP13 gene. Besides, the potential downstream genes of ZmSBP13 were uncovered and showed that it may target multiple proteins to regulate inflorescence structure. Overall, we characterized and cloned Tp2 mutant, and proposed a zma-miR156h-ZmSBP13 model functioning in regulating tassel branch development in maize, which is an essential measure to satisfy the increasing demands of cereals.

Identification of the Teopod1, Teopod2, and Early Phase Change genes in maize

“…This gene was repressed by about 50% in Tp1 /+ and by more than 80% in Tp2 /+. These results are consistent with a recent study of Tp2 ( Li et al . 2023 ) and strongly suggest that Tp1 and Tp2 are cis -acting mutations that increase the expression of, respectively, Zma-miR156j and Zma-miR156h .…”

“…This experiment revealed that both mutations were gain-of-function but produced different predictions for the specific nature of the gain-of-function allele in each case. We and others ( Li et al . 2023 ) have now shown that both mutations cause overexpression of genes encoding miR156, which raises the question of why this experiment produced different results for Tp1 and Tp2 .…”

Temporal regulation of vegetative phase change in plants

“…Loss-of-function mutations in tsh4 cause the development of bracts in the inflorescence 144 , while plants triply mutant for tsh4, ub2 and ub3 have numerous tillers, an unbranched leafy inflorescence, and flower at the same time as wild type but produce more leaves than normal because they have an elevated rate of leaf initiation 145 . This phenotype is very similar to that of Corngrass, Teopod1, and Teopod2 30,146,147 , all of which cause the overexpression of miR156 67,69,70 , but whether tsh4, ub2 and ub3 regulate leaf traits associated with vegetative phase change is unknown because these traits were not examined in these studies. The rice gene, OsSPL14-also known as Wealthy Farmer's Panicle (WFP) 148 , and Ideal Plant Architecture (IPA) 149 -is closely related to tsh4, ub2 and ub3, and has a similar function.…”

“…This result revealed that maize possesses juvenile and adult vegetative phases, and that the transition between these phases is under genetic control. It opened the way to subsequent genetic analyses of vegetative phase change in both maize 32,[54][55][56][57][58][59][60][61] and Arabidopsis 37,[62][63][64] and to the discovery that the microRNA miR156 is the master regulator of this transition [65][66][67][68][69][70] .…”

“…Consequently, leaf number-which is commonly used as a proxy for flowering time-is a poor predictor of flowering time in plants with elevated or reduced levels of miR156 or SPL proteins. The Tp2 mutant of maize, for example, has an elevated level of miR156 due to the over-expression of Zma-miR156h 69,70 . This mutant has an increased number of leaves and a small unbranched tassel with only a few flowers, but does not exhibit a major delay in its sensitivity to a floral inductive photoperiodic stimulus, and terminates shoot development at same time as wild type plants 56 ; the "leafy" phenotype of this mutant is attributable to a delay in the floral meristem identity transition, not to a delay in the acquisition of reproductive competence.…”