CRISPR/Cas9‐based genome editing of 14 lipid metabolic genes reveals a sporopollenin metabolon ZmPKSB‐ZmTKPR1‐1/‐2 required for pollen exine formation in maize

Abstract

SummaryLipid biosynthesis and transport are essential for plant male reproduction. Compared with Arabidopsis and rice, relatively fewer maize lipid metabolic genic male‐sterility (GMS) genes have been identified, and the sporopollenin metabolon in maize anther remains unknown. Here, we identified two maize GMS genes, ZmTKPR1‐1 and ZmTKPR1‐2, by CRISPR/Cas9 mutagenesis of 14 lipid metabolic genes with anther stage‐specific expression patterns. Among them, tkpr1‐1/‐2 double mutants displayed complete male sterility with delayed tapetum degradation and abortive pollen. ZmTKPR1‐1 and ZmTKPR1‐2 encode tetraketide α‐pyrone reductases and have catalytic activities in reducing tetraketide α‐pyrone produced by ZmPKSB (polyketide synthase B). Several conserved catalytic sites (S128/130, Y164/166 and K168/170 in ZmTKPR1‐1/‐2) are essential for their enzymatic activities. Both ZmTKPR1‐1 and ZmTKPR1‐2 are directly activated by ZmMYB84, and their encoded proteins are localized in both the endoplasmic reticulum and nuclei. Based on protein structure prediction, molecular docking, site‐directed mutagenesis and biochemical assays, the sporopollenin biosynthetic metabolon ZmPKSB‐ZmTKPR1‐1/‐2 was identified to control pollen exine formation in maize anther. Although ZmTKPR1‐1/‐2 and ZmPKSB formed a protein complex, their mutants showed different, even opposite, defective phenotypes of anther cuticle and pollen exine. Our findings discover new maize GMS genes that can contribute to male‐sterility line‐assisted maize breeding and also provide new insights into the metabolon‐regulated sporopollenin biosynthesis in maize anther.

GhTKPR1_8 functions to inhibit anther dehiscence and reduce pollen viability in cotton

“…Homologous proteins are generally similar functionally, reaffirming the preserved function of GhTKPR1_8 in pollen wall formation and shedding light on its potential additional roles in anther dehiscence (Peng et al, 2023). Generally, TKPR1 s show congruous expression patterns and localization at the tetrad stage in other species, such as Arabidopsis , rice, and maize (Grienenberger et al, 2010; Xu et al, 2019; An et al, 2024). We demonstrated that GhTKPR1_8 and GhTKPR1_17 showed typical expression patterns in the anthers of 3–6 mm buds (Figure 2).…”

“…OsTKPR1‐ loss‐of‐function mutations in rice degraded the tapetum, suppressed the levels of anther cuticular lipids, and disrupted the ubisch body and exine formation, ultimately leading to total male sterility (Xu et al, 2019). A maize double mutant, zmtkpr1 _ 1 / zmtkpr1 _ 2 , exhibited absolute male sterility, accompanied by delayed tapetal degradation, a defective exine, and abnormal anther cuticles (An et al, 2024). The tkpr1 mutant of Cryptomeria japonica displayed a thinner pollen wall compared to the wild type, reduced pollen grain count, and the production of non‐viable pollen (Kakui et al, 2023).…”

Genome-Wide Characterization of the Aldehyde Dehydrogenase Gene Superfamily in Maize and Its Potential Role in Anther Development

“…RNA-seq data for ZmALDHs in 14 tissues were obtained from MaizeGDB qTeller (). RNA-seq data of three maize inbred lines W23, B73, and Oh43 developing anthers were obtained from previously published articles. − Expression data are visualized in a heat map by TBtools.…”

Maize Improvement Based on Modern Breeding Strategies: Progress and Perspective

“…It can accelerate the efficiency of breeding by performing precise and predictable modifications of the target genes. For example, CRISPR/Cas9-based gene editing was used to simultaneously mutate multiple homologous genes required for pollen development and male fertility in maize, which definitely contributes to male-sterility line-assisted maize hybrid breeding. − CRISPR/Cas9 was used to make weak promoter alleles of CLV3/ESR-RELATED ( CLE ) genes and a null allele of a newly identified partially redundant compensating CLE gene to increase multiple maize grain-yield-related traits . Meanwhile, CRISPR/Cas9 is often used to make double knockout mutants.…”