Biology subjects

Kuwabara, K.

Publications and source records attributed to Kuwabara, K..

5 recordsLinked to original sources

bioRxiv2025

Characterization and transcriptome analysis reveal abnormal pollen germination in cytoplasmic male sterile tomato

Cytoplasmic male sterility (CMS) is a phenomenon resulting from mitochondrial-nuclear incompatibility that is widely used in hybrid breeding. Although the mitochondrial gene orf137 has been identified as the causal CMS gene in tomatoes, its function remains unclear. In this study, we characterized the male-sterile phenotypes and analyzed the CMS pollen transcriptome. Microscopic and calcium imaging analyses revealed that CMS pollen exhibited abnormal germination from multiple apertures, accompanied by elevated calcium concentrations and vesicle accumulation, which are typically observed in pollen tube tips. RNA-Seq analysis revealed 440 differentially expressed genes, including four pectin methylesterase inhibitor (PMEI) genes that were highly expressed in the pollen. PME activity was significantly reduced in CMS pollen, suggesting its association with abnormal pollen germination. ATP and reactive oxygen species (ROS) levels, which are key mediators of mitochondrial retrograde signaling (MRS), remained unchanged in CMS pollen, and the expression of the mitochondrial stress marker AOX1a was not elevated. These findings suggest that orf137 triggers an alternative MRS pathway independent of ATP or ROS, potentially leading to PMEI upregulation and abnormal pollen germination. Our results reveal a previously unrecognized mechanism of CMS-induced male sterility in tomatoes involving nuclear gene regulation through unconventional mitochondrial signaling.

plant biology↗

bioRxiv2025

Relationship Between Open Reading Frame 320, a Gene Causing Male Sterility in Tomatoes, and Cytoplasmic Male Sterility in Potatoes

Cytoplasmic male sterility (CMS) is a phenotype wherein plants cannot develop normal male organs because of mitochondrial genes. Several cytoplasmic sterility factors are suggested to be involved in CMS phenotypes in potatoes (Solanum tuberosum); however, reports on the relevant mitochondrial genes remain scarce. Many potato cultivars carry mitochondrial genes that cause CMS, resulting in pollen sterility, thereby limiting their use as male parents in breeding. Therefore, identifying the causal genes is crucial for potato breeding. In this study, we focused on the T/{beta}-type cytoplasm, the most prevalent genome type of potato worldwide, to explore mitochondrial genes involved in CMS in potatoes. We identified a novel gene, open reading frame 320 (orf320), in potato genotypes with T/{beta}-type cytoplasm by comparing the mitochondrial genomes of potatoes. Functional analysis of tomatoes showed that overexpression of orf320 with a mitochondrial transit peptide induced male sterility phenotypes, including abnormal anther development and pollen abortion. Furthermore, an investigation of orf320 in 126 potato cultivars revealed that this gene is specific to the T/{beta}-type cytoplasm and is absent in investigated cultivars with other cytoplasmic types. These findings provide strong evidence that orf320 is a candidate CMS-causing gene in male sterility of T/{beta}-type cytoplasm, offering valuable insights for future potato breeding.

plant biology↗

bioRxiv2025

Artificial mutations in the nuclear gene encoding mitochondrial RNA polymerase restore pollen fertility in cytoplasmic male sterile tomato

Cytoplasmic male sterility (CMS) and restorer of fertility (Rf) are important traits in F1 hybrid breeding. However, the CMS/Rf system has not been used in tomatoes because of the limited resources of Rf lines. In this study, we performed mutagenesis in tomato CMS seeds and successfully obtained 13 suppressor mutants with pollen fertility. Using bulked segregant analysis and whole-genome sequencing for each suppressor mutant, we detected mutations associated with fertility restoration in the nuclear-encoded gene for the mitochondrial RNA polymerase termed SlRPOTm in four independent mutants created through mutagenesis. Furthermore, we found that the loss of function of SlRPOTm was associated with fertility restoration in the tomato CMS line. Expression analysis of orf137, a tomato CMS-causing gene, revealed that reduced expression of orf137 was associated with fertility restoration in tomato CMS. In addition, F1 plants carrying mutations in SlRPOTm were generated, and tomato fruit formation was comparable to that of normal F1 plants. This study demonstrates for the first time that the loss of function of mitochondrial RNA polymerase contributes to fertility restoration in CMS lines. Furthermore, it is possible to replace various tomato varieties with Rf lines using genome editing technology, which will promote tomato F1 breeding in the future.

plant biology↗

bioRxiv2025

Multiple fertility restorer loci for cytoplasmic male sterility caused by orf137 in tomato

Cytoplasmic male sterility (CMS) in plants is caused by incompatibility between nuclear and cytoplasmic genetic information. Fertility can be restored through the action of fertility restoration (RF) genes, which are usually present in the nucleus. CMS lines of tomato (Solanum lycopersicum) have been developed from asymmetric cell fusions, in these lines, cultivated tomato served as a nuclear donor and its wild relative, S. acaule, as a cytoplasm donor. Although RF genes are present in wild relatives of tomato, no genetic or genomic information on the RF genes is yet available. This study reports an RF genetic locus, RF1, on chromosome 1 of S. pimpinellifolium LA1670 and S. lycopersicum var. cerasiforme LA1673 that was revealed by bulked segregant analysis and sequencing. An additional RF locus, RF2, was identified on chromosome 2 of LA1670. A transgenic approach identified two other candidate genes that also restored fertility. The genomic sequence of S. cheesmaniae LA0166 was assembled using high-fidelity, long-read sequencing technology. Sequence comparisons identified further candidate RF genes on chromosome 1 of S. cheesmaniae LA0166. These results suggested that multiple gene loci control the fertility restoration trait in wild relatives of tomato.

plant biology↗

bioRxiv2021

Organelle genome assembly uncovers the dynamic genome reorganization and cytoplasmic male sterility associated genes in tomato.

To identify cytoplasmic male sterility (CMS)-associated genes in tomato, we determined the genome sequences of mitochondria and chloroplasts in three CMS tomato lines derived from independent asymmetric cell fusions, their nuclear and cytoplasmic donors, and male fertile weedy cultivated tomato and wild relatives. The structures of the CMS mitochondrial genomes were highly divergent from those of the nuclear and cytoplasmic donors, and genes of the donors were mixed up in these genomes. On the other hand, the structures of CMS chloroplast genomes were moderately conserved across the donors, but CMS chloroplast genes were unexpectedly likely derived from the nuclear donors. Comparative analysis of the structures and contents of organelle genes and transcriptome analysis identified three genes that were uniquely present in the CMS lines, but not in the donor or fertile lines. RNA sequencing analysis indicated that these three genes transcriptionally expressed in anther, two of which were also expressed in pollen. They could be potential candidates for CMS-associated genes. This study suggests that organelle reorganization mechanisms after cell fusion events differ between mitochondria and chloroplasts, and provides insight into the development of new F1 hybrid breeding programs employing the CMS system in tomato.

plant biology↗