Dataset Information

Transcript levels of orf288 are associated with the hau cytoplasmic male sterility system and altered nuclear gene expression in Brassica juncea.

ABSTRACT: Cytoplasmic male sterility (CMS) is primarily caused by chimeric genes located in the mitochondrial genomes. In Brassica juncea, orf288 has been identified as a CMS-associated gene in the hau CMS line; however, neither the specific abortive stage nor the molecular function of the gene have been determined. We therefore characterized the hau CMS line, and found that defective mitochondria affect the development of archesporial cells during the L2 stage, leading to male sterility. The expression level of the orf288 transcript was higher in the male-sterility line than in the fertility-restorer line, although no significant differences were apparent at the protein level. The toxicity region of ORF288 was found to be located near the N-terminus and repressed growth of Escherichia coli. However, transgenic expression of different portions of ORF288 indicated that the region that causes male sterility resides between amino acids 73 and 288, the expression of which in E. coli did not result in growth inhibition. Transcriptome analysis revealed a wide range of genes involved in anther development and mitochondrial function that were differentially expressed in the hau CMS line. This study provides new insights into the hau CMS mechanism by which orf288 affects the fertility of Brassica juncea.

SUBMITTER: Heng S

PROVIDER: S-EPMC5853284 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Transcript levels of orf288 are associated with the hau cytoplasmic male sterility system and altered nuclear gene expression in Brassica juncea.

Heng Shuangping S Gao Jie J Wei Chao C Chen Fengyi F Li Xianwen X Wen Jing J Yi Bin B Ma Chaozhi C Tu Jinxing J Fu Tingdong T Shen Jinxiong J

Journal of experimental botany 20180101 3

Cytoplasmic male sterility (CMS) is primarily caused by chimeric genes located in the mitochondrial genomes. In Brassica juncea, orf288 has been identified as a CMS-associated gene in the hau CMS line; however, neither the specific abortive stage nor the molecular function of the gene have been determined. We therefore characterized the hau CMS line, and found that defective mitochondria affect the development of archesporial cells during the L2 stage, leading to male sterility. The expression l ...[more]

PMID: 29301015

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Project description:BACKGROUND: The novel chimeric open reading frame (orf) resulting from the rearrangement of a mitochondrial genome is generally thought to be a causal factor in the occurrence of cytoplasmic male sterility (CMS). Both positive and negative correlations have been found between CMS-associated orfs and the occurrence of CMS when CMS-associated orfs were expressed and targeted at mitochondria. Some orfs cause male sterility or semi-sterility, while some do not. Little is currently known about how mitochondrial factor regulates the expression of the nuclear genes involved in male sterility. The purpose of this study was to investigate the biological function of a candidate CMS-associated orf220 gene, newly isolated from cytoplasmic male-sterile stem mustard, and show how mitochondrial retrograde regulated nuclear gene expression is related to male sterility. RESULTS: It was shown that the ORF220 protein can be guided to the mitochondria using the mitochondrial-targeting sequence of the ? subunit of F1-ATPase (atp2-1). Transgenic stem mustard plants expressed the chimeric gene containing the orf220 gene and a mitochondrial-targeting sequence of the ? subunit of F1-ATPase (atp2-1). Transgenic plants were male-sterile, most being unable to produce pollen while some could only produce non-vigorous pollen. The transgenic stem mustard plants also showed aberrant floral development identical to that observed in the CMS stem mustard phenotype. Results obtained from oligooarray analysis showed that some genes related to mitochondrial energy metabolism were down-regulated, indicating a weakening of mitochondrial function in transgenic stem mustard. Some genes related to pollen development were shown to be down-regulated in transgenic stem mustard and the expression of some transcription factor genes was also altered. CONCLUSION: The work presented furthers our understanding of how the mitochondrially-targeted expression of CMS-associated orf220 gene causes male sterility through retrograde regulation of nuclear gene expression in Brassica juncea.

Project description:BACKGROUND: Cytoplasmic male sterility (CMS) is not only important for exploiting heterosis in crop plants, but also as a model for investigating nuclear-cytoplasmic interaction. CMS may be caused by mutations, rearrangement or recombination in the mitochondrial genome. Understanding the mitochondrial genome is often the first and key step in unraveling the molecular and genetic basis of CMS in plants. Comparative analysis of the mitochondrial genome of the hau CMS line and its maintainer line in B. juneca (Brassica juncea) may help show the origin of the CMS-associated gene orf288. RESULTS: Through next-generation sequencing, the B. juncea hau CMS mitochondrial genome was assembled into a single, circular-mapping molecule that is 247,903 bp in size and 45.08% in GC content. In addition to the CMS associated gene orf288, the genome contains 35 protein-encoding genes, 3 rRNAs, 25 tRNA genes and 29 ORFs of unknown function. The mitochondrial genome sizes of the maintainer line and another normal type line "J163-4" are both 219,863 bp and with GC content at 45.23%. The maintainer line has 36 genes with protein products, 3 rRNAs, 22 tRNA genes and 31 unidentified ORFs. Comparative analysis the mitochondrial genomes of the hau CMS line and its maintainer line allowed us to develop specific markers to separate the two lines at the seedling stage. We also confirmed that different mitotypes coexist substoichiometrically in hau CMS lines and its maintainer lines in B. juncea. The number of repeats larger than 100 bp in the hau CMS line (16 repeats) are nearly twice of those found in the maintainer line (9 repeats). Phylogenetic analysis of the CMS-associated gene orf288 and four other homologous sequences in Brassicaceae show that orf288 was clearly different from orf263 in Brassica tournefortii despite of strong similarity. CONCLUSION: The hau CMS mitochondrial genome was highly rearranged when compared with its iso-nuclear maintainer line mitochondrial genome. This study may be useful for studying the mechanism of natural CMS in B. juncea, performing comparative analysis on sequenced mitochondrial genomes in Brassicas, and uncovering the origin of the hau CMS mitotype and structural and evolutionary differences between different mitotypes.

Project description:BACKGROUND:Nsa cytoplasmic male sterility (CMS) is a novel alloplasmic male sterility system derived from somatic hybridization between Brassica napus and Sinapis arvensis. Identification of the CMS-associated gene is a prerequisite for a better understanding of the origin and molecular mechanism of this CMS. With the development of genome sequencing technology, organelle genomes of Nsa CMS line and its maintainer line were sequenced by pyro-sequencing technology, and comparative analysis of the organelle genomes was carried out to characterize the organelle genome composition of Nsa CMS as well as to identify the candidate Nsa CMS-associated genes. RESULTS:Nsa CMS mitochondrial genome showed a higher collinearity with that of S. arvensis than B. napus, indicating that Nsa CMS mitochondrial genome was mainly derived from S. arvensis. However, mitochondrial genome recombination of parental lines was clearly detected. In contrast, the chloroplast genome of Nsa CMS was highly collinear with its B. napus parent, without any evidence of recombination of the two parental chloroplast genomes or integration from S. arvensis. There were 16 open reading frames (ORFs) specifically existed in Nsa CMS mitochondrial genome, which could not be identified in the maintainer line. Among them, three ORFs (orf224, orf309, orf346) possessing chimeric and transmembrane structure are most likely to be the candidate CMS genes. Sequences of all three candidate CMS genes in Nsa CMS line were found to be 100% identical with those from S. arvensis mitochondrial genome. Phylogenetic and homologous analysis showed that all the mitochondrial genes were highly conserved during evolution. CONCLUSIONS:Nsa CMS contains a recombined mitochondrial genome of its two parental species with the majority form S. arvensis. Three candidate Nsa CMS genes were identified and proven to be derived from S. arvensis other than recombination of its two parental species. Further functional study of the candidate genes will help to identify the gene responsible for the CMS and the underlying molecular mechanism.

Project description:Cytoplasmic male sterility (CMS) lines are widely used for hybrid production in Brassica napus. The Shaan2A CMS system is one of the most important in China and has been used for decades; however, the male sterility mechanism underlying Shaan2A CMS remains unknown. Here, we performed transcriptomic and proteomic analysis, combined with additional morphological observation, in the Shaan2A CMS. Sporogenous cells, endothecium, middle layer, and tapetum could not be clearly distinguished in Shaan2A anthers. Furthermore, Shaan2A anther chloroplasts contained fewer starch grains than those in Shaan2B (a near-isogenic line of Shaan2A), and the lamella structure of chloroplasts in Shaan2A anther wall cells was obviously aberrant. Transcriptomic analysis revealed differentially expressed genes (DEGs) mainly related to carbon metabolism, lipid and flavonoid metabolism, and the mitochondrial electron transport/ATP synthesis pathway. Proteomic results showed that differentially expressed proteins were mainly associated with carbohydrate metabolism, energy metabolism, and genetic information processing pathways. Importantly, nine gene ontology categories associated with anther and pollen development were enriched among down-regulated DEGs at the young bud (YB) stage, including microsporogenesis, sporopollenin biosynthetic process, and tapetal layer development. Additionally, 464 down-regulated transcription factor (TF) genes were identified at the YB stage, including some related to early anther differentiation such as SPOROCYTELESS (SPL, also named NOZZLE, NZZ), DYSFUNCTIONAL TAPETUM 1 (DYT1), MYB80 (formerly named MYB103), and ABORTED MICROSPORES (AMS). These results suggested that the sterility gene in the Shaan2A mitochondrion might suppress expression of these TF genes in the nucleus, affecting early anther development. Finally, we constructed an interaction network of candidate proteins based on integrative analysis. The present study provides new insights into the molecular mechanism of Shaan2A CMS in B. napus.

Dataset Information

Transcript levels of orf288 are associated with the hau cytoplasmic male sterility system and altered nuclear gene expression in Brassica juncea.

Publications

Transcript levels of orf288 are associated with the hau cytoplasmic male sterility system and altered nuclear gene expression in Brassica juncea.

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