Changes in the Amounts of Ribulose Bisphosphate Carboxylase Synthesized and Degraded during the Life Span of Rice Leaf (Oryza sativa L.)

In situ synthesis and degradation of ribulose bisphosphate carboxylase(RuBPCase) were studied quantitatively in the 12th leaf bladeof the rice plant during the life span of the leaf. Levels ofRuBPCase protein were determined by rocket immunoelectrophoresis.The amounts of RuBPCase synthesized and degraded were estimatedusing ¹⁵N tracer. RuBPCase was scarcely recognized in the leaf when the tip ofthe leaf had just emerged from the 1 lth leaf sheath. Then itincreased rapidly and reached its maximum content a week afterthe completion of leaf expansion. At this time RuBPCase accountedfor 56% of the soluble leaf protein N (26% of the total leafN). The total amount of RuBPCase synthesized up to this timewas about 90% of the amount synthesized throughout the leaf'slife. Degradation of RuBPCase started about the time when it reachedthe maximum content and proceeded at a faster rate during senescencethan that of the remaining soluble protein. When the leaf hadsenesced completely, it contained little measurable RuBPCasealthough the total leaf N was about 30% of the maximum level.These results clearly suggest that RuBPCase is a major N componentwhich is used as remobilized N for the growth of young tissues. Influx and efflux of N and the synthesis and degradation ofRuBPCase are discussed in relation to leaf age. (Received February 18, 1983; Accepted June 16, 1983)     

Enteroviral and Mumps Meningitis in Toronto, 1963

Virological investigations of 115 children with the aseptic meningitis syndrome during 1963 resulted in the isolation of enteroviruses from cerebrospinal fluid (CSF) and/or feces of 21 of 48 children who had no association with mumps. For the third successive year, Echo 9 was the dominant enterovirus in cases of aseptic meningitis in Toronto children, but no rashes were associated with Echo 9 meningitis during 1963, in contradistinction to previous years. Mumps virus was isolated from CSF of 25 patients by inoculation of rhesus monkey kidney cultures, and rising or elevated mumps antihemagglutinin titres in paired sera from a further 33 cases provided laboratory evidence of infection with mumps virus in 58 of 67 patients with mumps meningoencephalitis. No enlargement of salivary glands was noted in 20 laboratory-proved cases of mumps meningoencephalitis. Enteroviral meningitis occurred principally during summer, but the peak of mumps meningoencephalitis occurred during late winter.     

Glutelin accumulation and changes in the levels of its mRNA in the superior and inferior spikelets of rice ear during ripening

Glutelin accumulation in the apical spikelet of the top primary branch (superior spikelet) and the second spikelet of the lowest secondary branch (inferior spikelet) of the ear of the rice plant (Oryza sativa L.) was characterized during grain filling.In the superior spikelet, the accumulation of dry matter and nitrogen started immediately after flowering and rapidly reached the maturation level by 20 days after heading (DAH). At 7 DAH, total RNA content had already reached its maximum level and glutelin mRNA content 70% of its maximum. The increase in glutelin mRNA was followed by a rapid increase in glutelin between 7 and 16 DAH.In the inferior spikelet dry matter, nitrogen and glutelin accumulation were low immediately after flowering and increased only after grain filling of the superior spikelet was almost complete. Total RNA and glutelin mRNA increased much later at slower rates than in the superior spikelet.It is very likely that the retardation of dry matter, total nitrogen and glutelin accumulation in the inferior spikelet is due to retardation of differentiation and development of endosperm tissue, and to glutelin gene expression in endosperm cells. It is suggested that the delayed development resulted from limited partitioning of nutrients to the inferior spikelet at the early stage of ripening.     

Changes in the Content of Two Glutamate Synthase Proteins in Spikelets of Rice (Oryza sativa) Plants during Ripening

Nitrogen accumulation in the apical spikelets on the primary branches of the main stem of rice plants have been studied during the ripening process (0-35 d after flowering). The level of NADH-dependent glutamate synthase (GOGAT) protein and activity increased 4- and 6-fold, respectively, in the first 15 d after flowering. Maximum levels of NADH-GOGAT were found at that time when the spikelets had just begun to increase in dry weight and to accumulate storage proteins. Subsequently, both the level of NADH-GOGAT protein and its activity in spikelets declined rapidly. Although changes in ferredoxin (Fd)-dependent GOGAT paralleled changes in NADH-GOGAT, the relative abundance of NADH-GOGAT protein in the spikelets was about 3 times higher than that of Fd-GOGAT from 5 to 15 d after flowering. When the chaff (lemma and palea) was separated from the spikelets 10 d after the flowering, 16% of the NADH-GOGAT protein was found in the chaff and 84% in the young grain tissues (endosperm, testae, aleurone tissues, and embryo). On the other hand, Fd-GOGAT protein was distributed 52% in the chaff and 48% in the young grain tissues in spikelets of the same age. Activity of NADP-isocitrate dehydrogenase, which may generate the 2-oxoglutarate required for the GOGAT reactions, was much higher than that of total GOGAT activities on a spikelet basis during the ripening process. These results suggest that in rice plants NADH-GOGAT is responsible for the synthesis of glutamate from the glutamine that is transported from senescing tissues to the spikelets.     

Relation between Nitrogen and Ribulose-1,5-bisphosphate Carboxylase in Rice Leaves from Emergence through Senescence

The relation between N content and ribulose-l,5-bisphosphate(RuBP) carboxylase protein was examined in the 12th leaf bladeof rice. Plants were grown under different amounts of N afterthe emergence of the 12th leaf blade. RuBP carboxylase proteinincreased with leaf N during leaf expansion. The synthesis ofRuBP carboxylase predominated during this period, and changesin the amounts of carboxylase synthesized until leaf death paralleledchanges in the N influx to the leaves. When the carboxylasereached its maximum content, the proportion of RuBP carboxylaseto leaf N was 27 to 28% irrespective of N treatment. As theleaf senesced, however, this proportion differed significantlywith the treatment. It was higher in the N-deficient leaf thanin the N-sufficient leaf. This was due to different patternsof RuBP carboxylase degradation for the treatments during senescence.RuBP carboxylase was degraded actively during the early stageof senescence in the N-sufficient leaf, whereas its degradationproceeded almost constantly in the N-deficient leaf during senescence. (Received October 17, 1983; Accepted January 27, 1984)     

The Remobilization of Nitrogen Related to Leaf Growth and Senescence in Rice Plants (Oryza sativa L.)

Rice plants (Oryzae sativa L.) grown in a nutrient solutionwere fed with (¹⁵NH₄)₂SO₄ during the 5 days of their young panicleformation. At the end of that time in the youngest leaf blade, which hadstarted to emerge during the labelling, absorbed-nitrogen accountedfor 37% of the increased nitrogen of the tissue; in the nextdeveloping leaf blade it accounted for 55%. Thus, remobilized-nitrogenoriginating from older patrs of the plant made up 63 and 45%,respectively, of their total nitrogen. The important contributionof the remobilized-nitrogen to the development of a leaf isevident. The remobilization of nitrogen in the 12th leaf blade on themain stem was examined in detail after labelling during itsdeveloping stage. The ¹⁵N level started to decrease soon afterthe end of the labelling period and continued to decrease untilfull senescence, although the total nitrogen in the same leafincreased until just after its complete expansion, suggestingthat even a young leaf plays a role as a supplier of remobilized-nitrogen. During the rapid decrease in the total nitrogen after its peakat full expansion of the leaf, the actual proportion of labelledabsorbed nitrogen remained nearly the same, indicating thatinflux of new nitrogen into a senescing leaf is very limited. (Received March 13, 1981; Accepted July 13, 1981)     

1-Acylglycerophosphocholine O-acyltransferase in maturing safflower seeds

The activity of 1-acylglycerophosphocholine (1-acyl-GPC) O-acyltransferase (EC 2.3.1.23) varied during maturation of safflower (Carthamus tinctorius L.) seeds, and activity per seed was highest in the middle period of seed development when triacylglycerol (TAG) is most rapidly synthesized. The specific activity of acyl transfer in a 20000·g particulate preparation exceeded 500nmol·min^-1·(mg protein)^-1 and was higher than those of any other enzymes involved in TAG synthesis (K. Ichihara et al., 1993, Plant Cell Physiol. 34, 557–566). This suggested the presence of a large flux of acyl-CoA to phosphatidylcholine in the cell. The reaction was specific to C₁₆ and C₁₈ acyl-CoAs with a double bond at position 9. Lauroyl- and erucoyl-CoA were completely ineffective, while ricinoleoyl- and elaidoyl-CoA were utilized efficiently. The relative order of specificity for native acyl-CoA species was linoleoyl > oleoyl stearoyl = palmitoyl. When acyl-CoA mixtures were presented, preference for the unsaturated species rather than the saturated species was even more apparent. The enzyme preferentially utilized 1-C₁₆-acyl- and 1-C₁₈-acyl-GPC molecular species, and 1-palmitoyl-, 1-stearoyl-, 1-oleoyl-and 1-linoleoyl-GPC equally served as acyl acceptor. No activity was detected with 1-octanoyl-GPC, and 1-erucoyl-GPC produced little effect. The effectiveness of 1-alkyl-GPC was comparable to that of 1-acyl-GPC. It was thus concluded that the enzyme recognizes the chain lengths of the acyl donor and acceptor, and the double bond at position 9 of the acyl donor.Abbreviations DAG diacylglycerol - DTNB 5,5-dithiobis(2-nitrobenzoic acid) - GP sn-glycerol 3-phosphate - GPC sn-glycero-3-phosphocholine - GPE sn-glycero-3-phosphoethanolamine - GPI sn-glycero-3-phosphoinositol - PC phosphatidylcholine - TAG triacylglycerol     

A metabolomics approach exploring the function of the ESX-3 type VII secretion system of M. smegmatis

The genome of Mycobacterium, including Mycobacterium tuberculosis, contains five copies of a cluster of genes encoding a novel type VII secretion system, named the ESX gene cluster region. This ESX-3 gene cluster is essential for in vitro growth and is thought to play a role in iron and zinc homeostasis, however, its exact functionality remains an enigma. A metabolomics research approach was subsequently used to compare the metabolite profiles of a M. smegmatis ESX-3 knockout strain to that a wild type parental strain, in order to elucidate its functionality from a metabolic perspective. Statistical analysis of the GC–MS generated data showed a clear separation between the wild type and knockout sample groups, based on the analysed metabolite profiles of these organisms. Of all the metabolite markers identified, various amino acids and metabolite pathways related to these, appeared to be most affected by the ESX-3 knockout, especially those with enzymes regulated by iron and zinc, supporting previous genomics and proteomics generated hypotheses and findings. This study is the first to demonstrate the capacity of using metabolomics, in conjunction with previous genomics and proteomic findings, to identify underlying metabolic changes and confirm previous hypotheses related to the functionality of ESX-3 in Mycobacterium growth and survival.