Involvement of Cyanogens in the Promotion of Germination of Cocklebur Seeds in Response to Various Nitrogenous Compounds, Inhibitors of Respiratory and Ethylene

Nitrogenous inhibitors of respiration, namely, KCN, NaN₃ andNH₂OH, which promote the germination of cocklebur (Xanthiumpennsylvanücum Wallr.) seeds, enhanced the accumulationof cyanogenic compounds. Urea and cyanamide, which were ineffective,did not. Part of the exogenously applied KCN was converted tocyanogenic glycosides and lipids, but these compounds were onlyproduced at low levels in NaN₃-treated seeds. Exposure to NH₂OHcaused a slight accumulation of both types of cyanogenic compounds.Thiourea, effective in stimulating germination, did not increasethe cyanogenic compounds, which suggests that the mechanismof its effect on germination is different from that of othernitrogenous compounds. Thiocyanate had a slightly promotiveeffect and caused minor increases in the levels of cyanogeniccompounds. Ethylene increased the metabolic utilization of the accumulatedcyanogens in response to KCN or NaN₃. The response to ethyleneof seeds in secondary dormancy was restored only after pre-treatmentwith KCN or NaN₃. The response occurred once the level of cyanogenshad increased. By contrast, enhancement of the effects of KCNand NaN₃ on germination by propyl gallate or benzohydroxamate,inhibitors of CN-resistant respiration, was accompanied by theincreased accumulation of cyanogens within the seeds that hadbeen exposed to KCN or NaN₃ in combination with the other typesof respiratory inhibitors. Thus, it is suggested that endogenouscyanogens might be involved in the germination of cockleburseeds. (Received December 20, 1995; Accepted April 9, 1996)     

Abscisic acid both promotes and inhibits photoperiodic flowering of Pharbitis nil

Abscisic acid (ABA) has been reported to have diverse effects on photoperiodic flowering. Activity of a natural ABA, (+)-( S )-abscisic acid (S-ABA), was recently suggested to be somewhat different from that of racemic ABA, which has been used in previous work. Use of S-ABA might enable clarification of the role of ABA in flowering. S-ABA inhibited flowering of the short-day plant Pharbitis nil (cv. Violet) when given before or 4 h after the start of a 14-h inductive dark period, and promoted flowering when given 12 h after the start of the dark period or later. The flower-promoting effect was observed when ABA was applied to the shoot apex. These results indicate that ABA has a dual effect on photoperiodic flowering of P. nil : it may inhibit the time-measuring process as well as promote some processes that proceed after generation of the flowering stimulus.     

Optical isolation of individual mitochondria ofPhysarum polycephalum for PCR analysis

Summary We attempted to amplify a specific region of mitochondrial DNA (mtDNA) using the polymerase chain reaction (PCR) from fewer than ten mitochondria isolated individually by microdissection or use of an optical tweezer. We selected preliminarily isolated mitochondria fromPhysarum polycephalum as the model materials and tried to amplify the mtDNA region corresponding to the specific mitochondrial plasmid of this true slime mould. For separation of a few mitochondria from the mitochondrial population, we initially used a destruction method in which excluded mitochondria were disrupted by a UV laser. However, mtDNA was still amplified, although weakly, from mitochondria that had been destroyed by the UV laser. Therefore, we used an optical tweezer to trap individual mitochondria and separate them from the others. The required number of mitochondria were separated from the mitochondrial suspension through a narrow canal of isolation buffer and used directly for PCR amplification. The results showed that the mtDNA could be amplified from at least 9 mitochondria trapped by the optical tweezer.Abbreviations DAPI 4,6-diamidino-2-phenylindole - EDTA ethylenediaminetetraacetic acid - mtDNA mitochondrial DNA - PCR polymerase chain reaction     

Mapping of the Gene for Machado-Joseph Disease within a 3.6-cM Interval Flanked by D14S291/D14S280 and D14S81, on the Basis of Studies of Linkage and Linkage Disequilibrium in 24 Japanese Families

The gene locus of Machado-Joseph disease (MJD) has recently been mapped within a 29-cM subregion of 14q chromosome. We did a linkage study of 24 multigenerational MJD Japanese pedigrees, in an attempt to narrow the candidate region of this gene. Pairwise and multipoint linkage analysis, together with haplotype segregation analysis, led to the conclusion that the MJD gene is located at the 6.8-cM interval between D14S256 and D14S81 (Z_max = 24.78, multipoint linkage analysis). D14S291 and D14S280, located at the center of this interval, showed no obligate recombination with the MJD gene (Z_max = 5.93 for D14S291 and 9.99 for D14S280). A weak, but significant, linkage disequilibrium of MJD gene was noted with D14S81 (P < .05) but not with D14S291 or D14S280. These results suggest that a 3.6-cM interval flanked by D14S291/D14S280 and D14S81 is the most likely location of the MJD gene and that it is closest to D14S81.     

Erythropoietin processing in erythropoietic system and central nervous system

We describe possible functions of carbohydrates attached to growth factors and strategies to examine the functions, concentrating on erythropoietin, a major regulator of erythropoiesis. Erythropoietin in erythropoiesis functions as an endocrine hormone; it is produced by kidney cells and transferred into the circulation to hemopoietic sites. In the brain, erythropoietin acts on neurons in a paracrine fashion. Comparison of glycosylation has been made between kidney and brain erythropoietins.Abbreviations BHK Baby Hamster Kidney - Epo Erythropoietin - Epo-R erythropoietin receptor     

A monoclonal antibody against T-cell receptor αβ induces endogenous cytokines and prevents mice from a lethal infection with Listeria monocytogenes

Abstract In vivo induction of cytokines by a monoclonal antibody (mAb) against T-cell receptor (TCR) αβ and the protective effect induced by the mAb on a lethal infection with Listeria monocytogenes were studied. Injection of anti-TCR αβ mAb induced rapid production of endogenous tumour necrosis factor in the spleens, and gamma interferon and interleukin-6 in the bloodstreams and spleens of mice. Administration of anti-CD4 mAb, anti-CD8 mAb, or anti-Thy1.2 mAb resulted in suppression of anti-TCR αβ mAb-induced endogenous cytokine production. Mice were protected against lethal L. monocytogenes infection when treated with anti-TCR αβ mAb. The protective effect was not demonstrated in CD4 + cell- or CD8 + cell-depleted mice. These results suggest that anti-TCR αβ mAb shows a protective effect on a lethal infection with L. monocytogenes in mice and that the mAb-induced endogenous cytokines might be involved in the effect of anti-TCR αβ mAb.     

An apolipoprotein CIII marker associated with hypertriglyceridemia in Caucasians also confers increased risk in a west Japanese population

Polymorphisms and haplotypes at the adjacent apolipoprotein (apo) AI and CIII gene loci were investigated in 61 Japanese patients with triglycerides greater than 350 mg/dl and in 66 unrelated normolipidemic subjects. The polymorphic sites were the SstI site in the apoCIII 3 untranslated region, whose presence has previously been shown to be associated with hypertriglyceridemia (HTG) in Caucasians, and the MspI site in the third intron of the apoAI gene. The frequencies of the SstI minor allele (S2) were 0.48 in HTG patients and 0.25 in normolipidemic subjects (P < 0.00015). The frequencies of the MspI minor allele (M2) were 0.61 in HTG patients and 0.33 in normolipidemic subjects (P < 0.00001). The two polymorphic sites were in strong linkage disequilibrium, and maximum likelihood analysis supported the existence of three of the four possible haplotypes: S1-M1, S1-M2, and S2-M2. Since all S2 alleles were estimated to be present on M2-bearing chromosomes, the HTG-associated S2-M2 haplotype conferred the same approximate relative risk as the S2 allele alone when compared with the other two haplotypes (odds ratio 2.8). This study demonstrates that the S2 allele is a marker for HTG among west Japanese subjects as well as among Caucasians. The results suggest that S2-M2 chromosomes carry HTG susceptibility sequences that predate the separation of the Asian and Caucasian races.     

Circadian rhythms of corneal mitotic rate,retinal melatonin and immunoreactive visual pigments,and the effects of melatonin on the rhythms in the Japanese quail

We investigated circadian ocular rhythms in the Japanese quail, Coturnix coturnix japonica. The birds were placed under light-dark cycles (LD 1212), constant light (LL) and constant darkness (DD), and the retinas were dissected out at four-hour intervals throughout 24 h. Following measurements were performed. (1) Melatonin content in the retina was measured by radioimmunoassay. It was low in light and several folds higher in darkness under LD 1212. The rhythm continued in DD, but disappeared in LL. (2) Mitotic figures in the corneal epithelium were counted. Similar rhythms to the melatonin content were observed in the corneal mitotic rate with a slight phase delay. (3) The retinas were fixed at 4-h intervals and immunostained with anti-bovine rhodopsin serum and anti-chicken iodopsin monoclonal antibodies. The outer segments of photoreceptor cells were stained intensively throughout 24 h in LD 1212, LL and DD. In contrast, the stainability of the locus close to the outer limiting membrane where the Golgi apparatus exists changed diurnally. Scores showing the ratio of cells with positive staining indicated high values from 4 h after the onset of light to the beginning of dark phase under LD 1212. The values were high throughout 24 h in LL and intermediate or low in DD. (4) To investigate the effect of melatonin on the corneal mitotic rate and visual pigments at the Golgi region, melatonin was injected into one eye and saline into the contralateral eye. Melatonin induced a phase advance in the corneal mitotic rate under LD 1212, but did not induce a rhythm under LL. The ratio of photoreceptor cells with positive staining to anti-visual pigment antibodies at the Golgi region was not affected by melatonin injection.Abbreviations ANOVA analysis of variance - BSA bovine serum albumin - DD constant darkness - Io-mAb monoclonal antibodies against chicken iodospin - LD light-dark - LL constant light - mRNA messenger ribonucleic acid - PBS phosphate buffer solution - Rh-As antiserum against bovine rhodopsin - SCN suprachiasmatic nucleus - T transducin - T transducin      

Dinucleotide repeat polymorphism in the human ceruloplasmin gene

We have identified a GT dinucleotide repeat polymorphism in intron 14 of the human ceruloplasmin gene. Observed heterozygosity for the polymorphism is 0.84.     

Loss ofHindIII cleavage sites in thed-amino acid oxidase gene in some inbred strains of mice

Summary d-Amino acid oxidase cDNA was amplified by a polymerase chain reaction using RNA extracted from the mouse kidney. When digested withHindIII, the cDNAs of the BALB/c and ddY/DAO^– mice were cleaved into two fragments whereas the cDNA of the ddY/DAO⁺ mice was not. Sequencing revealed that nucleotide-471 of the cDNAs was G in the BALB/c and ddY/DAO^– mice whereas it was substituted for C in the ddY/DAO⁺ mice. This base substitution was the cause of the failure of the cleavage of the cDNA of the ddY/DAO⁺ mice.Examination of other strains of inbred mice showed thatd-amino-acid oxidase cDNAs of A/J, AKR, C57BL/6, CD-1, CF#1, ICR, DBA/2, NZB and NZW mice were cleaved withHindIII into two fragments whereas those of C3H/He, CBA/J and NC mice were not. Genomic DNAs extracted from the mice of these 15 strains were digested withHindIII and hybridized withd-amino-acid oxidase cDNA. A 18.2-kb fragment hybridized with the probe in the C3H/He, CBA/J, ddY/DAO⁺ and NC mice whereas two fragments of 12 kb and 6.2 kb hybridized in the other mice. These results are consistent with those of the cDNAs, confirming the loss of theHindIII cleavage site in the C3H/He, CBA/J, ddY/DAO⁺ and NC mice. The Southern hybridization revealed a loss of a differentHindIII cleavage site in the A/J, AKR, C57BL/6, DBA/2, ICR and NZB mice.The substitution at nucleotide-471 should cause a substitution of an amino acid residue. However, this substitution did not affect catalytic activity ofd-amino acid oxidase.