Molecular mapping of restorer-of-fertility 2 gene identified from a sugar beet (Beta vulgaris L. ssp. vulgaris) homozygous for the non-restoring restorer-of-fertility 1 allele.

KEY MESSAGE: By genetically eliminating the major restorer - of - fertility gene ( Rf ), a weak Rf gene was unveiled. It is an allele of Z , long known as an elusive Rf gene in sugar beet. In the hybrid breeding of sugar beet, maintainer-genotype selection is a laborious process because of the dependence on test crossing, despite the very low occurrence of this genotype. Marker-assisted selection (MAS) of the maintainer genotype is highly desired by sugar beet breeders. The major restorer-of-fertility gene (Rf) was identified as Rf1, and its non-restoring allele (rf1) was discriminated at the DNA level; however, some of the rf1rf1 selections retained an as yet unidentified Rf, another target locus for MAS. The objective of this study was to identify this Rf. An rfrf1 plant was crossed to a cytoplasmic male-sterile sugar beet and then backcrossed to obtain progeny segregating the unidentified Rf. The progeny exhibited partial male-fertility restoration that was unstable in single plants. The segregation ratio of restored vs. non-restored plants suggested the involvement of a single Rf in this male-fertility restoration, designated as Rf2. We confirmed the feasibility of molecular tagging of Rf2 by identifying four shared amplified fragment length polymorphism (AFLP) fragments specific to 17 restored plants. Bulked segregant analysis also was performed to screen the Rf2-linked AFLP markers, which were subsequently converted into 17 sequence-tagged site markers. All the markers, as well two additional chromosome-IV-assigned markers, were linked to each other to form a single linkage map, on which Rf2 was located. Our data suggested that Rf2 is likely an allele of Z, long known as an elusive Rf gene in sugar beet. We also discuss the importance of Rf2 for sugar beet breeding.

Termination of Palliative Chemotherapy Near the End of Life: A Retrospective Study of Gastrointestinal Cancer Patients.

Background: Palliative chemotherapy is commonly used for advanced cancer patients. The timing of chemotherapy termination is crucial for efforts to maintain quality of life. Patients and Methods: This retrospective study included gastrointestinal cancer patients who were treated with chemotherapy and died between 2013 and 2022 at Niigata University Medical and Dental Hospital. Data were reviewed regarding age, gender, cancer type, reason for chemotherapy termination, cause of death, survival after chemotherapy termination, and place of death. Results: In total, 388 patients were included; the median survival after chemotherapy was 73 days. Patients aged <67 years had shorter survival durations (59 days), compared with patients aged >67 years (82 days). Ten (2.6%) patients began a new chemotherapy regimen, whereas 17 (4.4%) patients received chemotherapy, within 4 weeks before death. The most common reason for chemotherapy termination was disease progression, and most deaths occurred in hospitals. Conclusion: The rates of chemotherapy and initiation of new chemotherapeutic regimens near the end of life were lower than previously reported. Most deaths occurred in hospitals, highlighting the need for development of hospices.

Polymorphic minisatellites in the mitochondrial DNAs of Oryza and Brassica.

Polymorphic analyses of angiosperm mitochondrial DNA are rare in comparison with chloroplast DNA, because few target sequences in angiosperm mitochondrial DNA are known. Minisatellites, a tandem array of repeated sequences with a repeat unit of 10 to ~100 bp, are popular target sequences of animal mitochondria, but Beta vulgaris is the only known angiosperm species for which such an analysis has been conducted. From this lack of information, it was uncertain as to whether polymorphic minisatellites existed in other angiosperm species. Ten plant mitochondrial DNAs were found to contain minisatellite-like repeated sequences, most of which were located in intergenic regions but a few occurred in gene coding and intronic regions. Oryza and Brassica accessions were selected as models for the investigation of minisatellite polymorphism because substantial systematic information existed. PCR analysis of 42 Oryza accessions revealed length polymorphisms in four of the five minisatellites. The mitochondrial haplotypes of the 16 Oryza accessions with chromosomal complement (genome) types of CC, BBCC and CCDD were identical but were clearly distinguished from BB-genome accessions, a result consistent with the notion that the cytoplasmic donor parent of the amphidiploid species might be the CC-genome species. Twenty-nine accessions of six major cultivated species of Brassica were classified into five mitochondrial haplotypes based on two polymorphic minisatellites out of six loci. The haplotypes of Brassica juncea and Brassica carinata accessions were identical to Brassica rapa and Brassica nigra accessions, respectively. The haplotypes of Brassica napus accessions were heterogeneous and unique, results that were consistent with previous studies.

What Does the Molecular Genetics of Different Types of Restorer-of-Fertility Genes Imply?

Cytoplasmic male sterility (CMS) is a widely used trait for hybrid seed production. Although male sterility is caused by S cytoplasm (male-sterility inducing mitochondria), the action of S cytoplasm is suppressed by restorer-of-fertility (Rf), a nuclear gene. Hence, the genetics of Rf has attained particular interest among plant breeders. The genetic model posits Rf diversity in which an Rf specifically suppresses the cognate S cytoplasm. Molecular analysis of Rf loci in plants has identified various genes; however, pentatricopeptide repeat (PPR) protein (a specific type of RNA-binding protein) is so prominent as the Rf-gene product that Rfs have been categorized into two classes, PPR and non-PPR. In contrast, several shared features between PPR- and some non-PPR Rfs are apparent, suggesting the possibility of another grouping. Our present focus is to group Rfs by molecular genetic classes other than the presence of PPRs. We propose three categories that define partially overlapping groups of Rfs: association with post-transcriptional regulation of mitochondrial gene expression, resistance gene-like copy number variation at the locus, and lack of a direct link to S-orf (a mitochondrial ORF associated with CMS). These groups appear to reflect their own evolutionary background and their mechanism of conferring S cytoplasm specificity.

High-quality sugar production by osgcs1 rice.

Carbohydrates (sugars) are an essential energy-source for all life forms. They take a significant share of our daily consumption and are used for biofuel production as well. However, sugarcane and sugar beet are the only two crop plants which are used to produce sugar in significant amounts. Here, we have discovered and fine-tuned a phenomenon in rice which leads them to produce sugary-grain. We knocked-out GCS1 genes in rice by using CRISPR technology, which led to fertilization failure and pollen tube-dependent ovule enlargement morphology (POEM) phenomenon. Apparently, the POEMed-like rice ovule ('endosperm-focused') can grow near-normal seed-size unlike earlier observations in Arabidopsis in which gcs1 ovules ('embryo-focused') were aborted quite early. The POEMed-like rice ovules contained 10-20% sugar, with extremely high sucrose content (98%). Trancriptomic analysis revealed that the osgcs1 ovules had downregulation of starch biosynthetic genes, which would otherwise have converted sucrose to starch. Overall, this study shows that pollen tube content release is sufficient to trigger sucrose unloading at rice ovules. However, successful fertilization is indispensable to trigger sucrose-starch conversion. These findings are expected to pave the way for developing novel sugar producing crops suited for diverse climatic regions.

How did a duplicated gene copy evolve into a restorer-of-fertility gene in a plant? The case of Oma1.

Restorer-of-fertility (Rf) is a suppressor of cytoplasmic male sterility (CMS), a mitochondrion-encoded trait that has been reported in many plant species. The occurrence of CMS is considered to be independent in each lineage; hence, the question of how Rf evolved was raised. Sugar beet Rf resembles Oma1, a gene for quality control of the mitochondrial inner membrane. Oma1 homologues comprise a small gene family in the sugar beet genome, unlike Arabidopsis and other eukaryotes. The sugar beet sequence that best matched Arabidopsis atOma1 was named bvOma1; sugar beet Rf (RF1-Oma1) was another member. During anther development, atOma1 mRNA was detected from the tetrad to the microspore stages, whereas bvOma1 mRNA was detected at the microspore stage and RF1-Oma1 mRNA was detected during the meiosis and tetrad stages. A transgenic study revealed that, whereas RF1-Oma1 can bind to a CMS-specific protein and alter the higher-order structure of the CMS-specific protein complex, neither bvOma1 nor atOma1 show such activity. We favour the hypothesis that an ancestral Oma1 gene duplicated to form a small gene family, and that one of the copies evolved and acquired a novel expression pattern and protein function as an Rf, i.e. RF1-Oma1 evolved via neofunctionalization.

A fertility-restoring genotype of beet (Beta vulgaris L.) is composed of a weak restorer-of-fertility gene and a modifier gene tightly linked to the Rf1 locus.

Cytoplasmic male sterility (CMS) is a plant trait that involves interactions between nuclear- and mitochondrial genomes. In CMS, the nuclear restorer-of-fertility gene (Rf), a suppressor of male-sterility inducing mitochondria, is one of the best known genetic factors. Other unidentified genetic factors may exist but have not been well characterized. In sugar beet (Beta vulgaris L.), CMS is used for hybrid seed production, but few male-sterility inducing nuclear genotypes exist. Such genotypes could be introduced from a closely related plant such as leaf beet, but first the fertility restoring genotype of the related plant must be characterized. Here, we report the discovery of a Japanese leaf beet accession 'Fukkoku-ouba' that has both male-sterility inducing and fertility restoring genotypes. We crossed the leaf beet accession with a sugar beet CMS line, developed succeeding generations, and examined the segregation of two DNA markers that are linked to two sugar beet Rfs, Rf1 and Rf2. Only the Rf2 marker co-segregated with fertility restoration in every generation, implying that the Rf1 locus in leaf beet is occupied by a non-restoring allele. Fertility restoration was incomplete without a genetic factor closely linked to Rf1, leading to the assumption that the Rf1 locus encodes a modifier that cannot restore fertility by itself but perhaps strengthens another Rf. We sequenced the apparently non-restoring 'Fukkoku-ouba' rf1 gene-coding region and found that it closely resembles a restoring allele. The protein product demonstrated its potential to suppress CMS in transgenic suspension cells. In contrast, 'Fukkoku-ouba' rf1 transcript abundance was highly reduced compared to that of the restoring Rf1. Consistently, changes in protein complexes containing CMS-associated mitochondrial protein in anthers were very minor. Accordingly, we concluded that 'Fukkoku-ouba' rf1 is a hypomorph that acts as a non-restoring allele but has the potential to support another Rf, i.e. it is a modifier candidate.

Pollen tube contents initiate ovule enlargement and enhance seed coat development without fertilization.

In angiosperms, pollen tubes carry two sperm cells toward the egg and central cells to complete double fertilization. In animals, not only sperm but also seminal plasma is required for proper fertilization. However, little is known regarding the function of pollen tube content (PTC), which is analogous to seminal plasma. We report that the PTC plays a vital role in the prefertilization state and causes an enlargement of ovules without fertilization. We termed this phenomenon as pollen tube-dependent ovule enlargement morphology and placed it between pollen tube guidance and double fertilization. Additionally, PTC increases endosperm nuclei without fertilization when combined with autonomous endosperm mutants. This finding could be applied in agriculture, particularly in enhancing seed formation without fertilization in important crops.

He and Ar beam scatterings from bare and defect induced graphite surfaces.

The effect of the local electronic modification of the graphite surface on the gas-graphite interaction has been investigated by the molecular beam scattering technique. The angular intensity distributions of He and Ar beams scattered from pristine and defect induced graphite surfaces have been measured at various surface temperatures. From the He scattering results, the cross-section for the He diffuse scattering per defect is estimated as being as much as 113 nm(2). The origin of the extremely large cross-section is ascribed to the modulated electronic states of graphite around the defect based on the STM measurements, which is due to the local breaking of the π conjugated system of graphite. From the Ar scattering results, the effective mass of the graphite surface for the Ar collision has been estimated as M = 114 u, based on the analysis with the hard cube model. The new component appears in the scattering distribution of Ar for the defect induced graphite surface. The component has a larger peak position angle than that for the pristine graphite surface, indicating that the normal component of the translational energy of the Ar atom was greatly lost by the collision at the electronically modified area of graphite.