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Biomedical subjects

Masao Watanabe

Publications and source records attributed to Masao Watanabe.

At least 19 recordsLinked to original sources

Characterization of the SP11/SCR high-affinity binding site involved in self/nonself recognition in brassica self-incompatibility.

In Brassica self-incompatibility, the recognition of self/nonself pollen grains, is controlled by the S-locus, which encodes three highly polymorphic proteins: S-locus receptor kinase (SRK), S-locus protein 11 (SP11; also designated S-locus Cys-rich protein), and S-locus glycoprotein (SLG). SP11, located in the pollen coat, determines pollen S-haplotype specificity, whereas SRK, located on the plasma membrane of stigmatic papilla cells, determines stigmatic S-haplotype specificity. SLG shares significant sequence similarity with the extracellular domain of SRK and is abundant in the stigmatic cell wall, but its function is controversial. We previously showed that SP11 binds directly to its cognate SRK with high affinity (K(d) = 0.7 nM) and induces its autophosphorylation. We also found that an SLG-like, 60-kD protein on the stigmatic membrane forms a high-affinity binding site for SP11. Here, we show that the 60-kD stigmatic membrane protein is a truncated form of SRK containing the extracellular domain, transmembrane domain, and part of the juxtamembrane domain. A transiently expressed, membrane-anchored form of SRK exhibits high-affinity binding to SP11, whereas the soluble SRK (eSRK) lacking the transmembrane domain exhibits no high-affinity binding, as is the case with SLG. The different binding affinities of the membrane-anchored SRK and soluble eSRK or SLG will be significant for the specific perception of SP11 by SRK.

Amino Acid Sequence↗

Dominance relationships between self-incompatibility alleles controlled by DNA methylation.

In crucifers, the pollen S-determinant gene, SP11, is sporophytically expressed in the anther tapetum, and the pollen self-incompatibility phenotype is determined by the dominance relationships between the two S-haplotypes it carries. We report here that 5' promoter sequences of recessive SP11 alleles are specifically methylated in the tapetum before the initiation of SP11 transcription. These results suggest that tissue-specific monoallelic de novo DNA methylation is involved in determining the dominance interactions that determine the cruciferous self-incompatibility phenotype.

Brassica↗

Molecular characterization of mature pollen-specific genes encoding novel small cysteine-rich proteins in rice (Oryza sativa L.).

In our previous cDNA microarray analysis, we identified 53 mature anther-specific genes, whose function was unknown, in rice. We reanalyzed these genes from the viewpoint of the specific amino acid motif. Out of 53 genes, three genes, Os-26, Os-32, and Os-169 (renamed as OsSCP1, OsSCP2, and OsSCP3), encoded cysteine-rich motif (Cys-X3-Cys-X13-Cys-X3-Cys), indicating that they were novel small cysteine-rich proteins. From the search of specific elements in promoter regions, several pollen-specific elements were found. In order to determine whether three promoters were functional in pollen or not, the gene constructs with promoter regions fused to the beta-glucuronidase gene were transformed into tobacco. Histochemical analysis showed that these promoters were active in the mature pollen grains and pollen tubes. Furthermore, OsSCP1 and OsSCP3 formed a multigene family tandemly in the rice genome. From the results, OsSCPs might have important roles in mature pollen development and pollen tube growth.

Amino Acid Sequence↗

MCI-186 administered to the maternal circulation inhibits fetal brain injury resulting from total umbilical cord occlusion in the chronically instrumented fetal lamb.

OBJECTIVE: To evaluate the transplacental effects of MCI-186 (edaravone), a potent hydroxyl radical scavenger, administered to the maternal circulation to inhibit fetal brain injury caused by umbilical cord occlusion. METHODS: Nine chronically instrumented lambs were prepared. In three cases, 10-min persistent total umbilical cord occlusion (group A) was performed. Another three cases underwent occlusion and were administered 60 mg of MCI-186 through the maternal femoral vein prior to the end of occlusion (group B). The remaining three cases underwent sham operation (group C). On day 3 after insult, fetal brains were extirpated. Paraffin-embedded brain tissue sections were stained with hematoxylin and eosin, Bodian, Kluver-Barrera, and TUNEL. Neuronal cellular damage was evaluated by two pathologists blinded to the experimental conditions. RESULTS: Group A displayed numerous cells with eosinophilic condensation of nuclear chromatin and proliferation of microglia in the hippocampus and basal ganglia. TUNEL-positive cells were observed in the periventricular area. Group B showed microglial proliferations, but no marked changes. No pathological changes were apparent in group C. CONCLUSIONS: MCI-186 administered to the maternal circulation could inhibit fetal brain injury resulting from hypoxia-reperfusion induced by umbilical cord occlusion.

Animals↗

Pathological and biochemical studies on a case of Pick disease with severe white matter atrophy.

We report on a male patient with Pick disease who had shown severe white matter atrophy and dilatation of the lateral ventricle in the frontal lobe from an early stage. Upon admission to our hospital 2 years after disease onset, the patient showed apathy, and MRI revealed severe atrophy of the cortex and white matter of the frontal lobe. He died at age 74, 11 years after disease onset. Autopsy revealed severe atrophy of the frontal and temporal lobes, severe loss of white matter in the frontal lobe, dilatation of the lateral ventricles, and cortical thinning. Histopathological examination showed severe loss of myelinated fibers in the frontal white matter and severe neuronal loss with gliosis in the frontal and temporal cortices. Many Pick bodies were seen. Our patient had a rare case of Pick disease predominantly affecting the frontal lobe with severe involvement of the white matter from an early stage. This case suggests that myelinated fibers in the white matter as well as cerebral neurons are primarily affected in Pick disease.

Aged↗

The production of hydroxyl radicals in the fetal lamb brain resulting from occlusion of the umbilical circulation and the transplacental effect of MCI-186 to inhibit hydroxyl radical production.

The present study evaluated hydroxyl radical production in fetal lamb brain during and after umbilical cord occlusion and examined the effects of injecting MCI-186 (3-metyl-1-phenyl-2-pyrazolin-5-one; Edaravone), a hydroxyl radical scavenger, into the maternal circulation. In 11 chronically instrumented lambs, intermittent total umbilical cord occlusions 1 min out of every 3 min for 1 h and 10-min persistent total umbilical cord occlusion were performed with brain microdialysis using 5 mM of sodium salicylate. In the remaining four lambs, 60 mg of MCI-186 was administered into the maternal circulation from shortly before the end of 10-min persistent total umbilical cord occlusion. Concentrations of 2,3-dihydroxy-benzoic acid (2,3-DHBA), produced by hydroxyl radical reactions with sodium salicylate, were measured in perfusate by HPLC. Concentration of 2,3-DHBA in perfusate was 23.05 +/- 10.95 nM before umbilical cord occlusion. Levels of 2,3-DHBA tended to increase slightly during and after intermittent umbilical cord occlusion, and were significantly increased by the end of 10-min occlusion (40.06 +/- 21.36 nM) and after occlusion (93.74 +/- 29.17 nM). Infusion of MCI-186 suppressed 2,3-DHBA concentration to 29.35 +/- 14.95 nM after occlusion. Administration of MCI-186 into the maternal circulation reduces hydroxyl radical production induced by umbilical cord occlusion in the fetal lamb brain.

Animals↗

Molecular characterization of two anther-specific genes encoding putative RNA-binding proteins, AtRBP45s, in Arabidopsis thaliana.

RRM (RNA-recognition motif) domain is important for the post-transcriptional regulation of gene expression including RNA processing. In our previous study, we found one anther- and/or pollen-specific gene (LjRRM1, previously named as LjMfb-U93) in model legume, Lotus japonicus. Because of the richness of genomic information of another model plant, Arabidopsis thaliana, for functional analysis, we identified and characterized the orthologous genes in A. thaliana. By comparison of the partial nucleotide sequence of LjRRM1 to the public database, we identified three homologous genes (AtRBP45a, AtRBP45b, and AtRBP45c) in A. thaliana genome. Based on promoter analysis, both AtRBP45a and AtRBP45c were specifically expressed in immature anther tissues (tapetum cells) and mature pollen grains of transgenic plants. This expression pattern of AtRBP45a and AtRBP45c is quite similar to that of LjRRM1, indicating that AtRBP45a and AtRBP45c would be orthologous to LjRRM1. Because in another previous experiment, it was shown that proteins having RRM domains were related to pre-mRNA maturation, and as a conclusion, it is possible that LjRRM1, AtRBP45a, and AtRBP45c genes encoding RNA-binding proteins are functionally involved in the repression of translation in mature pollen grains in L. japonicus and A. thaliana.

Amino Acid Sequence↗

Anther-specific genes, which expressed through microsporogenesis, are temporally and spatially regulated in model legume, Lotus japonicus.

Pollen germination and pollen tube elongation are important for pollination and fertilization in higher plants. To date, several pollen-specific genes have been isolated and characterized. However, there is little information about the precise spatial and temporal expression pattern of pollen-specific genes in higher plants. In our previous study, we identified 132 anther-specific genes in the model legume Lotus japonicus by using cDNA microarray analysis, though their precise expression sites in the anther tissues were not determined. In this study, by using in situ hybridization experiments, we determined the spatial and temporal expression sites of 46 anther-specific genes (ca. 35%), which were derived from two groups, cluster I-a and cluster II-a, according to flower developmental stages. In the case of the genes grouped into cluster I-a, thirteen clones were characterized. The specific hybridized signals were varied among the clones, and were observed in tapetum cells, microspores, and anther walls at the early developmental stage of anther tissues. In the case of the genes classified into cluster II-a, we used thirty three different cDNA clones encoding primary and secondary metabolism-related proteins, cell wall reconstruction-related proteins, actin reorganization-related proteins, and sugar transport-related proteins, etc., as a probe. Interestingly, all genes in these thirty three clones examined were specifically expressed in the bicellular pollen grains, though the signal intensity was varied among clones. From the data of the cluster II-a genes, the mRNAs related to pollen germination and pollen tube elongation were specifically transcribed and preserved in mature pollen grains.

Gene Expression Regulation, Plant↗

Comparative analysis of the S-intergenic region in class-II S haplotypes of self-incompatible Brassica rapa (syn. campestris).

In the Brassica self-incompatibility (SI) system, a pollen determinant, SP11, is involved in dominance/recessive relationships in pollen SI phenotypes. In order to gain some insights into the genomic structure around the SP11 and the mechanisms that give dominance/recessive relationships, we characterized the genomic region containing SP11 and SRK genes in three pollen recessive class-II S haplotypes. The direction of transcription of S genes was completely conserved among class-II S haplotypes. However, the region between SP11 and SRK (S-intergenic region) was highly polymorphic without short repetitive sequences. In addition, we found a sequence similarity between the short repetitive sequence and 5'-upstream region of SP11. This sequence similarity was found to be potentially related to the expression of dominance relationships through the change of chromatin structure.

Base Sequence↗

Colloid (enterogenous) cyst in the frontal lobe.

A 60-year-old man presented with a left frontal mass lesion incidentally detected at a health check without apparent symptoms. Computed tomography revealed the lesion as homogeneous high density and magnetic resonance (MR) imaging showed the lesion as hyperintense on T(1)-weighted images, isointense on T(2)-weighted images, and hypointense on diffusion-weighted images. T(1)-weighted MR images with gadolinium showed no enhancement of the mass lesion. Cerebral angiography revealed an avascular area around the left frontal lesion. Total removal of the lesion was achieved through a craniotomy without complications. Histological, immunohistochemical, and electron microscopy examinations established the definite diagnosis of colloid cyst.

Brain Neoplasms↗

Restoration of prehension using double free muscle technique after complete avulsion of brachial plexus in children: a report of three cases.

PURPOSE: Brachial plexus injury in children, excluding birth palsy, is relatively rare and seldom reported. We report our technique, the results of this procedure, and problems we encountered in treating children with brachial plexus injury. METHODS: From 1999 through 2002, we treated 3 children with complete avulsion of the brachial plexus due to trauma by using double free muscle technique (DFMT) with a nerve transfer procedure using the contralateral seventh cervical nerve root transfer to reconstruct prehensile function. There were 2 boys aged 5 and 11 years and a girl aged 4 years. All patients were followed up for at least 3 years after the surgery. RESULTS: All the transferred muscles survived without any vascular complications and were reinnervated successfully. The average active range of elbow flexion was 125 degrees (range, 90 degrees - 145 degrees ). The average total active range of motion of the fingers was 69 degrees (range, 40 degrees -102 degrees ). All patients obtained voluntary prehensile function and could use the reconstructed hand for activities of daily living. They were able to lift and carry light objects with the reconstructed hand and heavy objects with both hands. CONCLUSIONS: The results of DFMT for reconstruction of BPI in children were encouraging. Appropriate postoperative rehabilitation under close supervision is important to obtain useful prehensile function.

Accessory Nerve↗

Environmental factors affecting black/white coloration of the silken girdle in the swallowtail butterfly, Atrophaneura alcinous (Lepidoptera: Papilionidae).

The silken girdles of pupae of the swallowtail butterfly Atrophaneura alcinous show black and white color diphenism. Field observations revealed that all pupae observed on non-food plants and the leaves and stems of the larval food plant Aristolochia debilis were classified as a silken girdle of a black type, while a large portion of pupae pupating on the twigs and trunks of cherry trees in close proximity to A. debilis were classified as a silken girdle of a black type. Additionally, all pupae observed on the surfaces of artificial objects in areas where there are no surrounding plants or trees were classified as a silken girdle of a white type. We demonstrated the effect of day length and the texture, light, plant odor and humidity of pupation sites on the coloration of the silken girdle in A. alcinous. Regardless of long-day or short-day day length conditions, light conditions of constant light or dark, or the presence of a plant odor of A. debilis as environmental cues, all larvae placed at over 80% relative humidity (R.H.) developed into pupae with a silken girdle of a black type. However, all larvae developed into pupae with a silken girdle of a white type when R.H. was below 75%. Furthermore, when pupae with a silken girdle of a white type were transferred to conditions of 90% R.H. within 24 hr of pupation, the white color of the silken girdle changed into a black type within 24 hr of the transfer. The present data suggest that the induction of a black coloration of the silken girdle in A. alcinous requires a R.H. of approximately 80% or more as an environmental factor.

Animals↗

A membrane-anchored protein kinase involved in Brassica self-incompatibility signaling.

Self-incompatibility (SI) response in Brassica is initiated by haplotype-specific interactions between the pollen-borne ligand S locus protein 11/SCR and its stigmatic S receptor kinase, SRK. This binding induces autophosphorylation of SRK, which is then thought to trigger a signaling cascade that leads to self-pollen rejection. A recessive mutation of the modifier (m) gene eliminates the SI response in stigma. Positional cloning of M has revealed that it encodes a membrane-anchored cytoplasmic serine/threonine protein kinase, designated M locus protein kinase (MLPK). Transient expression of MLPK restores the ability of mm papilla cells to reject self-pollen, suggesting that MLPK is a positive mediator of Brassica SI signaling.

Amino Acid Sequence↗

Genetic characterization of a novel Tib-derived variant of soybean Kunitz trypsin inhibitor detected in wild soybean (Glycine soja).

A novel variant of soybean Kunitz trypsin inhibitor (SKTI) was detected in 530 lines of wild soybean (Glycine soja). This variant showed an intermediate electrophoretic mobility between the Tia and Tic types. In isoelectric focusing polyacrylamide gel electrophoresis gels containing urea, this variant had a similar isoelectric point as that of Tia. The genetic analysis of SKTI bands in F2 seeds from crosses of the new variant type with Tia or Tic type showed that this variant type is controlled by a codominant allele at the SKTI locus. We propose the genetic symbol Tif for this novel variant. When the nucleotide sequence of the Tif gene was compared with those of other types of SKTI genes (Tia, Tib, and Tic), the sequence of Tif was identical to that of Tib with the exception of one A-->G transitional mutation occurring at position 676 of Tif. This mutation resulted in an amino acid change from Lys to Glu at the 178 residue. These results suggest that this variant is derived from Tib through a point mutation. In addition, we settled an inconsistency in the number of amino acid differences between Tia and Tib (eight or nine). Analysis of nucleotide and amino acid sequences revealed that Tib was different from Tia by nine amino acids.

Amino Acid Sequence↗

Identification and molecular characterization of novel anther-specific genes in Oryza sativa L. by using cDNA microarray.

The complicated genetic pathway regulates the developmental programs of male reproductive organ, anther tissues. To understand these molecular mechanisms, we performed cDNA microarray analyses and in situ hybridization to monitor gene expression patterns during anther development in rice. Microarray analysis of 4,304 cDNA clones revealed that the hybridization signal of 396 cDNA clones (271 non-redundant groups) increased more than six-fold in every stage of the anthers compared with that of leaves. Cluster analysis with the expression data showed that 259 cDNA clones (156 non redundant groups) were specifically or predominantly expressed in anther tissues and were regulated by developmental stage-specific manners in the anther tissues. These co-regulated genes would be important for development of functional anther tissues. Furthermore, we selected several clones for RNA in situ hybridization analysis. From these analyses, we found several novel genes that show temporal and spatial expression patterns during anther development in addition to anther-specific genes reported so far. These results indicate that the genes identified in this experiment are controlled by different programs and are specialized in their developmental and cell types.

Cluster Analysis↗

Cloning and expression pattern of a novel microspore-specific gene encoding hypersensitive-induced response protein (LjHIR1) from the model legume, Lotus japonicus.

In order to understand the microspore and pollen development, recently, we have isolated a number of anther-specific genes in the model legume, Lotus japonicus. From these anther-specific genes, we identified one novel microspore-specific gene, LjImfb-c82. In order to determine the molecular characterization of LjImfb-c82, full-length cDNA clone was first isolated and sequenced. It encoded a protein of 286 amino acids (LjHIR1), which had sequence similarity to Hypersensitive-Induced Response like protein. LjHIR1 was specifically expressed in microspore on the in situ hybridization experiment. From the sequence similarity to prohibitin-domain protein, the LjHIR1 might be related to ion channel regulation in microspore development.

Amino Acid Sequence↗

Duplicated SP11 genes produce alternative transcripts in the S15 haplotype of Brassica oleracea.

Self-incompatibility (SI) discriminating self and non-self pollen is regulated by S-locus genes in Brassica. In most S haplotypes, a set of three highly polymorphic genes, SLG, SRK, and SP11, is located at the S-locus region. In the present study, we found duplicated SP11 genes, S15-SP11a, S15-SP11b, and S15-SP11b', in the self-incompatible S15 haplotype of B. oleracea. RNA gel blot and reverse transcription polymerase chain reaction (RT-PCR) analyses showed that two different sizes of SP11 transcripts were specifically detected in anther tissues: a 0.65-kb transcript corresponded to S15-SP11a (an exon-1 region of S15-SP11b was also co-transcribed in some cases), and a 1.4-kb transcript contained the duplicated three genes, S15-SP11a, S15-SP11b, and an exon-1 region of S15-SP11b', all three of which were connected to intergenic spacer regions.

Alternative Splicing↗