Photosynthetic characteristics and genetic mapping of a new yellow leaf mutant crm1 in Brassica napus.

Chlorophyll is one of the key factors for photosynthesis and plays an important role in plant growth and development. We previously isolated an EMS mutagenized rapeseed chlorophyll-reduced mutant (crm1), which had yellow leaf, reduced chlorophyll content and fewer thylakoid stacks. Here, we found that crm1 showed attenuated utilization efficiency of both light energy and CO2 but enhanced heat dissipation efficiency and greater tolerance to high-light intensity. BSA-Seq analysis identified a single nucleotide change (C to T) and (G to A) in the third exon of the BnaA01G0094500ZS and BnaC01G0116100ZS, respectively. These two genes encode the magnesium chelatase subunit I 1 (CHLI1) that catalyzes the insertion of magnesium into protoporphyrin IX, a pivotal step in chlorophyll synthesis. The mutation sites resulted in an amino acid substitution P144S and G128E within the AAA+ domain of the CHLI1 protein. Two KASP markers were developed and co-segregated with the yellow leaf phenotype in segregating F2 population. Loss of BnaA01.CHLI1 and BnaC01.CHLI1 by CRISPR/Cas9 gene editing recapitulated the mutant phenotype. BnaA01.CHLI1 and BnaC01.CHLI1 were located in chloroplast and highly expressed in the leaves. Furthermore, RNA-seq analyses revealed the expression of chlorophyll synthesis-related genes were upregulated in the crm1 mutant. These findings provide a new insight into the regulatory mechanism of chlorophyll synthesis in rapeseed and suggest a novel target for improving the photosynthetic efficiency and tolerance to high-light intensity in crops. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-023-01429-6.

Characterization and fine mapping of a new dwarf mutant in Brassica napus.

BACKGROUND: Plant height is an important plant characteristic closely related to yield performance of many crops. Reasonable reduction of plant height of crops is beneficial for improving yield and enhancing lodging resistance. RESULTS: In the present study, we described the Brassica napus dwarf mutant bnd2 that was isolated using ethyl methanesulfonate (EMS) mutagenesis. Compared to wild type (WT), bnd2 exhibited reduced height and shorter hypocotyl and petiole leaves. By crossing the bnd2 mutant with the WT strain, we found that the ratio of the mutant to the WT in the F2 population was close to 1:3, indicating that bnd2 is a recessive mutation of a single locus. Following bulked segregant analysis (BSA) by resequencing, BND2 was found to be located in the 13.77-18.08 Mb interval of chromosome A08, with a length of 4.31 Mb. After fine mapping with single nucleotide polymorphism (SNP) and insertion/deletion (InDel) markers, the gene was narrowed to a 140-Kb interval ranging from 15.62 Mb to 15.76 Mb. According to reference genome annotation, there were 27 genes in the interval, of which BnaA08g20960D had an SNP type variation in the intron between the mutant and its parent, which may be the candidate gene corresponding to BND2. The hybrid line derived from a cross between the mutant bnd2 and the commercial cultivar L329 had similar plant height but higher grain yield compared to the commercial cultivar, suggesting that the allele bnd2 is beneficial for hybrid breeding of lodging resistant and high yield rapeseed. CONCLUSION: In this study, we identified a novel dwarf mutant of rapeseed with a new locus, which may be useful for functional analyses of genetic mechanisms of plant architecture and grain yield in rapeseed.

Phenotype and TMT-based quantitative proteomics analysis of Brassica napus reveals new insight into chlorophyll synthesis and chloroplast structure.

The conversion of light energy into chemical energy in leaves is very important for plant growth and development. During this process, chlorophylls and their derivatives are indispensable as their fundamental role in the energy absorption and transduction activities. Chlorophyll variation mutants are important materials for studying chlorophyll metabolism, chloroplast biogenesis, photosynthesis and related physiological processes. Here, a chlorophyll-reduced mutant (crm1) was isolated from ethyl methanesulfonate (EMS) mutagenized Brassica napus. Compared to wild type, crm1 showed yellow leaves, reduced chlorophyll content, fewer thylakoid stacks and retarded growth. Quantitative mass spectrometry analysis with Tandem Mass Tag (TMT) isobaric labeling showed that totally 4575 proteins were identified from the chloroplast of Brassica napus leaves, and 466 of which displayed differential accumulations between wild type and crm1. The differential abundance proteins were found to be involved in chlorophyll metabolism, photosynthesis, phagosome and proteasome. Our results suggest that the decreased abundance of chlorophyll biosynthetic enzymes, proteins involved in photosynthesis might account for the reduced chlorophyll content, impaired thylakoid structure, and reduction of plant productivity. The increased abundance of proteins involved in phagosome and proteasome pathways might allow plants to adapt the proteome to environmental conditions to ensure growth and survival due to chlorophyll reduction. BIOLOGICAL SIGNIFICANCE: Photosynthesis, which consists of light and dark reactions, is fundamental to biomass production. Chloroplast is regarded as the main site for photosynthesis. During photosynthesis, the pigment chlorophyll is essential for light harvesting and energy transfer. This work provides new insights into protein expression patterns, and enables the identification of many attractive candidates for investigation of chlorophyll biosynthesis, chloroplast structure and photosynthesis in Brassica napus. These findings may be applied to improve the photosynthetic efficiency by genetic engineering in crops.