MS26/CYP704B is required for anther and pollen wall development in bread wheat (Triticum aestivum L.) and combining mutations in all three homeologs causes male sterility

Abstract

Development of anthers and pollen represents an important aspect of the life cycle in flowering plants. Genes contributing to anther and pollen development have been widely studied in many plant species. Ms26/CYP704B genes play an important role in pollen development through biosynthesis of sporopollenin for pollen exine formation. To investigate the role of Ms26/CYP704B genes in anther and pollen development of bread wheat, mutations in the A-, B-, and D-homeologs of the putative Ms26/CYP704B gene were analyzed. Single and double homozygous mutants in any of the homeologs did not affect pollen development and male fertility. Triple homozygous mutants resulted in completely male sterile plants that were defective in pollen and anther development. Additionally, double homozygous-single heterozygous mutants were also male sterile although with varying levels of residual fertility. The fertility of these triple mutants was dependent upon the homeolog contributing the wild-type allele. Two heterologous Ms26/CYP704B genes, when transformed into a triple homozygous mutant background, completely restored male fertility, whereas a single gene was unable to restore fertility. Functional analysis of Ms26/CYP704B furthers the understanding of male fertility genes which can be utilized for the development of novel hybrid seed production systems in wheat.

Identification of genes involved in male sterility in wheat (Triticum aestivum L.) which could be used in a genic hybrid breeding system

“…Using CRISPR/Cas9 editing, we were fortunate in our ability to show a direct phenotype in the T 0 gene‐edited plants; more work needs to be done to determine which homoeologue is the best candidate for a genic hybrid system with these two genes. Our data suggest that in contrast to Ms26 in wheat, one functional allele appears sufficient to allow normal seed set (Singh et al, ). We also show the power of combining multiple guides in one gene‐editing cassette as we achieved an overall success rate of knocking out all six alleles in 14% of the T 0 plants in the case of TaRPG1 ‐targeted homoeologues.…”

“…Our data suggest that in contrast to Ms26 in wheat, one functional allele appears sufficient to allow normal seed set (Singh et al, 2017). We also show the power of combining multiple guides in one gene-editing cassette as we achieved an overall success rate of knocking out all six alleles in 14% of the T 0 plants in the case of TaRPG1-targeted homoeologues.…”

“…Currently, only one dominant gene, Ms2 , and one recessive mutant have been cloned in wheat (Ni et al, ; Tucker, Baumann, & Kouidri, ; Wang et al, ; Xia et al, ), with Ms2 mutants being widely used for wheat breeding and potentially for hybrid wheat breeding (Ni et al, ). Other genes involved in male sterility in wheat include the genes Ms26 and Ms45 which are involved in the formation of the pollen cell wall similar to ms1 which is a transcription factor and regulates the post‐meiotic development of the anther (Dong et al, ; Singh, Kumar, Thilges, Cho, & Cigan, ; Tucker et al, ; Wang et al, ). Here, we set out to identify other genes involved in pollen viability in wheat and show their possible use as genes involved in a genic male sterility system.…”

CRISPR/Cas9‐mediated knockout of Ms1 enables the rapid generation of male‐sterile hexaploid wheat lines for use in hybrid seed production

“…Initial attempts to use (meganuclease‐based) genome editing for the generation of male‐sterile wheat lines (Cigan et al ., ) were met with limited success, as editing of only one of the three Ms26 homoeologs (A, B or D‐genome) did not confer male sterility due to functional redundancy of the gene. However, conventional crossing of single‐genome biallelic Ms26 mutants was successfully carried out to produce triple biallelic Ms26 mutants exhibiting male sterility (Singh et al ., ). More recently, the CRISPR/Cas9 system was used to edit the wheat male fertility gene Ms45 , and by selfing a triple monoallelic mutant T 1 plant, male‐sterile triple biallelic mutant T 2 plants were recovered (Singh et al ., ).…”

Wheat ms5 male‐sterility is induced by recessive homoeologous A and D genome non‐specific lipid transfer proteins

“…In an example of redundancy, triple homoeotic mutants of the sporopollenin biosynthesis gene Ms26/CYP704B were required by Singh et al . () to produce sterile wheat plants. In tetraploid durum wheat, Scarascia‐Mugnozza et al .…”