Comparative Study on Coat Proteins of an Attenuated, a Temperature Sensitive and Their Parent Strains of Tobacco Mosaic Virus

An attenuated strain (L₁₁A) of tobacco mosaic virus (TMV) induces no remarkable symptoms on tomato plants (Goto and Nemoto 1971) and has been used to protect tomato against virulent strains of TMV (Oshima 1981), A temperature sensitive strain (Ls1) of TMV was isolated and found to have a malfunction of virus movement from cell to cell (NISHI-GUCHI et al. 1978, 1980). Those two strains are derived from a wild virulent strain (L). Coat proteins of them were compared with one another and with that of Dahlemense (D) strain of TMV, in order to see whether coat protein was associated with their respective characters. The coat proteins of the four strains behaved similar in both SDS-polyacrylamide gel and 8 M urea polyacrylamide gel electrophoresis, suggesting that they are similar in molecular weight and charging effect in the gels. There was no significant difference in chromatographic pattern of tryptic peptides among the four strains. Amino acid compositions of tryptic peptides revealed that three strains, L₁₁A, Ls1 and L, were identical to one another and that they differed from D slightly. These results suggest that coat protein is related neither to virus attenuation of L₁₁A nor to the malfunction of Ls1.     

The bumblebee Bombus ardens ardens (Hymenoptera: Apidae) visits white clover in orchards before Oriental persimmon blooms

Flowers on the ground of orchards can provide substantial resources for wild pollinators of orchard trees. Few studies, however, have examined the relative importance of groundcover flowers to orchard pollination by analyzing pollen on the body surface of pollinators. Oriental persimmon trees bloom within the longer blooming period of white clover, which is occasionally found as a flowering plant on the ground of persimmon orchards in Japan. The present study compared the insect species assemblage collected on persimmon flowers with that on clover. Before persimmon bloomed, Bombus ardens ardens and Apis cerana japonica were the major visitors of clover flowers. Once persimmon bloomed, the former was the most abundant bee that visited persimmon flowers over the flowering period. Apis mellifera was captured only on clover flowers. We found numerous clover pollen grains on the body surface of bumblebees captured on persimmon flowers, but far fewer persimmon pollen grains on bees that visited clover. These findings show that B. ardens ardens utilized the clover flowers under the orchards before persimmon bloomed.     

Physical and Functional Interaction of Transmembrane Thioredoxin-related Protein with Major Histocompatibility Complex Class I Heavy Chain: Redox-based Protein Quality Control and Its Potential Relevance to Immune Responses

In the endoplasmic reticulum (ER), a variety of oxidoreductases classified in the thioredoxin superfamily have been found to catalyze the formation and rearrangement of disulfide bonds. However, the precise function and specificity of the individual thioredoxin family proteins remain to be elucidated. Here, we characterize a transmembrane thioredoxin-related protein (TMX), a membrane-bound oxidoreductase in the ER. TMX exists in a predominantly reduced form and associates with the molecular chaperon calnexin, which can mediate substrate binding. To determine the target molecules for TMX, we apply a substrate-trapping approach based on the reaction mechanism of thiol-disulfide exchange, identifying major histocompatibility complex (MHC) class I heavy chain (HC) as a candidate substrate. Unlike the classical ER oxidoreductases such as protein disulfide isomerase and ERp57, TMX seems not to be essential for normal assembly of MHC class I molecules. However, we show that TMX–class I HC interaction is enhanced during tunicamycin-induced ER stress, and TMX prevents the ER-to-cytosol retrotranslocation of misfolded class I HC targeted for proteasomal degradation. These results suggest a specific role for TMX and its mechanism of action in redox-based ER quality control.     

Detection of Specific RNAs by in situ Hybridization in Plant Protoplasts

Combined exposure of fixed tomato protoplasts to 0.2 N HCl atroom temperature, heating at 70?C in 2 ? SSC for 30 min, anddigestion with 10 µg/ml proteinase K allowed localizationof viroid RNA by in situ hybridization, without destructionof cellular structures. The technique also allowed localizationof viral RNA in tobacco protoplasts. ⁴Present address: National Institute of Animal Health, Tsukuba,Ibaraki, 305 Japan (Received March 1, 1989; Accepted February 21, 1990)     

Mechanism of post-irradiation degradation of deoxyribonucleic acid in a radiosensitive Escherichia coli (NG30) irradiated with ultraviolet light

The post-irradiation degradation of DNA in a radiosensitive mutant, NG30, of Escherchia coli, was investigated. An extensive degradation of DNA corresponding to the ultraviolet dose was observed. The post-irradiation degradation of DNA having no replicating points is not initiated from a specific point in the DNA, but does occur simultaneously at many sites of the irradiated DNA.     

A Rhipicephalus (Boophilus) microplus cathepsin with dual peptidase and antimicrobial activity

The cattle tick, Rhipicephalus (Boophilus) microplus, is a haematophagous arthropod responsible for considerable losses in the livestock industry. Immunological control with vaccines is a promising alternative to replace chemical acaricides. Due to their importance in parasite physiology, cysteine endopeptidases are potential targets. In a previous study, native Vitellin Degrading Cysteine Endopeptidase (VTDCE) was successfully tested as a vaccine antigen for bovines against R. microplus. In this work, nucleotide and amino acid VTDCE sequences were obtained from cDNA databanks, based on data from Edman sequencing and mass spectrometry. Subsequently, cloning and expression, purification, immunological and biochemical characterisation of the recombinant protein were performed to determine the biological importance of VTDCE. By Western blot, polyclonal antibodies produced against recombinant VTDCE recognised native VTDCE. Interestingly, molecular analysis showed that the VTDCE sequence has similarity to antimicrobial peptides. Indeed, experimental results revealed that VTDCE has an antimicrobial activity which is independent of endopeptidase activity. We believe that this is the first known study to show that an arthropod enzyme has antimicrobial activity.     

Simultaneous UPLC MS/MS analysis of endogenous jasmonic acid,salicylic acid,and their related compounds

Jasmonic acid (JA) and salicylic acid (SA) are plant hormones involved in plant growth and development. Recent studies demonstrated that presence of a complex interplay between JA and SA signaling pathways to response to pathogenesis attack and biotic stresses. To our best knowledge, no method has existed for simultaneous analyses of JA, SA, and their related compounds. Especially, the glucosides are thought to be the storages or the inactivated compounds, but their contribution should be considered for elucidating the amount of the aglycons. It is also valuable for measuring the endogenous amount of phenylalanine, cinnamic acid, and benzoic acid that are the biosynthetic intermediates of SA due to the existence of isochorismate pathway to synthesize SA. We established this method using deuterium labeled compounds as internal standards. This is the first report of simultaneous analysis of endogenous JA, SA, and their related compounds. Measuring the endogenous JA, SA, and their related compounds that had been accumulated in tobacco plants proved the practicality of the newly developed method. It was demonstrated that accumulation of JA, SA and their related compounds were induced in both case of TMV infection and abiotic stresses.