The 33 kDa Protein of the Oxygen-Evolving Complex: a Multi-Gene Family in Tomato

A cDNA was isolated by chance from tomato which had a high similarityto a cDNA clone from potato known to code for the 33 kDa proteinof the oxygen-evolving complex [van Spanje et al. (1991) PlantMol. Biol. 17: 157]. The sequence of a previously describedpartial cDNA clone from tomato [Ko et al. (1990) Plant Mol.Biol. 14: 217] which has also a high similarity but is not identicalto the sequence described here indicates that tomato containsat least two genes coding for 33 kDa proteins per haploid genome.This conclusion is supported by Southern blot analysis. Thetissue specific expression of the corresponding genes is described. (Received September 28, 1992; Accepted February 15, 1993)     

Isolation of Genes for Low-Temperature-Induced Proteins in Rice by a Simple Subtractive Method

We have isolated cDNAs that correspond to three distinct low-temperature-inducedmRNAs from rice cells using an effective, simple subtractionmethod. The corresponding genes were designated lip (gene encodinglow-temperature-induced protein) –5,–9 and –19.The expression of these genes was slightly stimulated lip5 andlip19) or unaffected (lip9) by abscisic acid, unlike the expressionof the rabl6A-D genes which are readily induced by either abscisicacid or high osmotic stress. Time-course analysis revealed thatwhile the lip5 gene was induced 3 h after temperature down-shift,and the lip9 and lipl9 genes were induced 6 h after such a shift. ¹Present address: Division of Molecular Medicine, Clinical ResearchCentre, Northwick Park Hospital, Harrow, Middlesex HA1 3UJ,U.K. (Received June 29, 1991; Accepted October 7, 1991)     

Isolation of a Gene for a Metallothionein-Like Protein from Soybean

Using a synthetic oligonucleotide that corresponded to the consensusnucleotide sequence of the N-terminal region of mammalian metallothioneinas probe, we isolated a cDNA clone from a soybean library. Theclone had an ORF that encode a protein of 79 amino acids whichshowed significant homology to both N- and C-terminal regionsof mammalian and Neurospora crassa metallothioneins ⁴Present address: Department of Biosciences, Teikyo University,Toyosatodai, Utsunomiya, Tochigi, 320 Japan (Received March 13, 1991; Accepted June 17, 1991)     

Phosphate Deficiency in Maize. II. Enzyme Activities

Low-phosphate (P) treatment decreased photosynthetic rates ofmaize plants (Zea mays L.) by about 50% 18 to 19 days afterplanting [Usuda and Shimogawara (1991) Plant Cell Physiol. 32:497]. Low-P treatment decreased the enzyme activities differentially(by 0-49%). The significance of the decreased activities ofpyruvate.Pj dikinase (by 29%), phosphoenolpyruvate carboxylase(by 49%), and ribulose 1,5-bisphosphate carboxylase (by 42%)in the detrimental effects of low-P treatments on the ratesof photosynthesis is discussed. (Received August 9, 1991; Accepted October 1, 1991)     

The nifH-Like (frxC) Gene Is Involved in the Biosynthesis of Chlorophyll in the Filamentous Cyanobacterium Plectonema boryanum

The frxC gene found in the DNA of the liverwort chloroplastencodes a protein of unknown function. The deduced amino acidsequence shows significant homology to that of the nitrogenaseFe-protein encoded by the nifH gene. We previously identifiedthe frxC and nifH genes in the filamentous cyanobacterium Plectonemaboryanum. We describe here the isolation of targeted mutantsof frxC (YFC1004) and nifH (YFH201) which were generated byinsertion of a kanamycin-resistance gene into the structuralportion of the respective genes. As expected, YFH201 cannotgrow under nitrogen-fixing conditions. However, YFC1004 growsas well as the wild type does under nitrogen-fixing, photoautotrophicand chemoheterotrophic conditions, indicating that the FrxCprotein is essential neither for nitrogen fixation nor for majorenergytransduction systems, such as photosynthesis and respiration.YFC1004 synthesizes chlorophyll (Chi) normally in the lightbut not in the dark, and it accumulates a precursor to Chi,protochlorophyllide (Pchlide) in the dark. These phenotypiccharacteristics of YFC1004 suggest that the cyanobacterium hastwo pathways for the reduction of Pchlide: a light-dependentand a lightindependent system. The FrxC protein appears to beinvolved in the light-independent reduction of Pchlide. (Received September 24, 1991; Accepted November 11, 1991)     

Plant Growth-promoting Rhizobacteria and Soybean [ Glycine max (L.) Merr.] Growth and Physiology at Suboptimal Root Zone Temperatures

Application of plant growth-promoting rhizobacteria (PGPR) hasbeen shown to increase legume growth and development under optimaltemperature conditions, and specifically to increase nodulationand nitrogen fixation of soybean [Glycine max (L.) Merr.] overa range of root zone temperatures (RZTs). Nine rhizobacteriaapplied into soybean rooting media were tested for their abilityto reduce the negative effects of low RZT on soybean growthand development by improving the physiological status of theplant. Three RZTs were tested: 25, 17.5, and 15 °C. At eachtemperature some PGPR strains increased plant growth and development,but the stimulatory strains varied with temperature. The strainsthat were most stimulatory at each temperatures were as follows:15 °C—Serratia proteamaculans 1–102; 17.5 °C—Aeromonashydrophila P73, and 25 °C—Serratia liquefaciens 2–68.Because enhancement of plant physiological activities were detectedbefore the onset of nitrogen fixation, these stimulatory effectscan be attributed to direct stimulation of the plant by thePGPR rather than stimulation of plant growth via improvementof the nitrogen fixation symbiosis. Legume; nitrogen fixation; nodulation; root zone temperature; PGPR     

Cloning, Nucleotide Sequences and Differential Expression of the nifH and nifH-Like (frxC) Genes from the Filamentous Nitrogen-Fixing Cyanobacterium Plectonema boryanum

The frxC gene, found in liverwort chloroplast DNA, encodes aprotein of unknown function. The deduced amino acid sequenceof the protein shows significant homology to that of ni-trogenaseFe-protein encoded by the nifH gene. We have cloned the frxCand nifH genes from the nitrogen-fixing cyanobacterium Plectonemaboryanum, using frxC- and nifH-specific probes, and have determinedtheir nucleotide sequences. The amino acid sequence deducedfrom the frxC gene of P. boryanum exhibits 83% homology to thatof the protein encoded by the/rxCgene from liverwort, whereasit exhibits only 34% homology to that encoded by the nifH genefrom the same organism, namely, P. boryanum. Northern blot analysisshowed that the frxC gene was transcribed more actively undernitrogenase-repressed conditions than under nitrogenase-inducedconditions, suggesting that the FrxC protein has a functiondistinct from nitrogen fixation. These results, together withthe phylogenetic relationship between the nifH and frxC genes,indicate that the frxC and nifH genes are derived from a commonancestral gene but have evolved independently to encode proteinswith different functions. (Received April 27, 1991; Accepted August 12, 1991)     

Transformation of Indica Rice (Oryza sativa L.) Mediated by Agrobacterium tumefaciens

A system has been developed for the transfer of genes by Agrobacteriumtumefaciens to indica rice (Oryza sativa L. cv. Taichung Native1), and optimal conditions for such Agrobacterium-mediated transformationhave been determined. Excised stems, leaves, and roots from3- to 4-day-old seedlings were infected with strains of Agrobacteriumthat carried plasmids with chimeric derivatives of genes forß-glucuronidase (GUS) and neomycin phosphotransferase(NPTII). After selection with the antibiotic G418, only rootssurvived and formed calli. The integration of chimeric genesin the rice genome was demonstrated by Southern analysis andassays of enzymatic activity. We obtained a transformation frequencyof 37.5% for transgenic callus. In addition, the absence ofhybridization with vir probes indicated that transformants werenot contaminated with Agrobacterium. The complete failure oftransformation in the absence of medium from potato suspensioncultures (PSC) indicates that PSC is essential for Ti-mediatedtransformation in indica rice. A high efficiency of transformationwas observed upon infection with the cointegrate vector pGV2260::NG1. (Received August 13, 1991; Accepted April 28, 1992)     

The role of thiol compounds in increasing Agrobacterium-mediated transformation of soybean cotyledonary-node cells

Agrobacterium-mediated transformation of soybean cells and the production of fertile transgenic soybean [Glycine max (L.) Merrill] plants using the cotyledonary-node (cot-node) method were improved by amending the solid co-cultivation medium with L-cysteine. The goal of this study was to investigate the role of cysteine and other thiol compounds in increasing the frequency of transformed soybean cot-node cells. The frequency of transformed cells was increased only when L-cysteine was present during co-cultivation of Agrobacterium and cot-node explants. This effect was due to the thiol group since D-cysteine and other thiol compounds also increased the frequency of transformed cells. Copper and iron chelators also increased the frequency of transformed cells, indicating an association with inhibition of polyphenol oxidases and peroxidases. Thiol compounds likely inhibit wound- and pathogen-induced responses, thereby increasing the capacity for Agrobacterium-mediated transformation of soybean cells. The increases in transformed cot-node cells were independent of soybean genotype, Agrobacterium strain, and binary plasmid.     

Characterization of {beta}-amylase isozymes in soybean seeds by isoelectric focusing

Distributions of the isozymes of ß-amylase [EC 3.2.1.2 [EC] ]in seeds of the cultivated soybean, Glycine max, the semicultivatedtype, Glycine gradlis, and the wild type, Glycine ussuriensis,were studied by the isoelectric focusing method. Four isozymes(1 to 4) were found in seeds of the soybean and related species;their respective isoelectric points were 5.12, 5.25, 5.38 and5.52?0.02. Crystalline components I and II described in a previouspaper corresponded to isozymes 2 and 4, respectively. Each varietyor species of soybean contained one or two isozymes, the typeof isozyme pattern being characteristic of the variety of soybeanand the related species. Taxonomically, the isozyme patternsseem to correlate pardy with some morphological features, e.g.the determinate growth habit, the semitwinning property of thestem and the color of the seed episperm. (Received January 9, 1979; )     

Localizing a-Amylase Gene Expression in Germinated Rice Grains

In situ hybridization was used to localize the sites of expressionfor two a-amylase genes (RAmy1A and RAmy3D) in rice (Oryza sativaL.) over five days of germination. Messenger RNAs from bothgenes were initially detected in the scutellar epithelium andappeared at later stages of germination in the aleurone layer.RAmy3D mRNA reached its peak of accumulation 2 days(d) earlierin the scutellar epithelium than RAmy1A mRNA. Both mRNAs continuedto accumulate in the aleurone layer up to 5 d of germination,although RAmy1A mRNA reached significantly higher levels thanRAmy3D mRNA. Overall, the aleurone layer was responsible forproducing the majority of the total grain a-amylase mRNA. (Received July 27, 1991; Accepted November 6, 1991)     

Inhibition of ATPase Activity in Pea Plasma Membranes In Situ by a Suppressor from a Pea Pathogen, Mycosphaerella pinodes

A pathogenic fungus of pea, Mycosphaerella pinodes, secretesa so-called "suppressor" in its pycnospore germination fluid.The suppressor blocks the defense responses and induces localsusceptibility (accessibility) in pea plants to agents thatare not pathogenic in pea. The suppressor nonspecifically inhibitsthe ATPase activity in plasma membranes prepared from pea, soybean,kidney bean, cowpea and barley plants. However, cytochemicalstudies by electron microscopy indicate that the suppressorspecifically inhibits the ATPase in pea cell membranes, butnot in those of four other plant species tested. That is, thespecificity of the suppressor appears at the cell and/or tissuelevel, but is not evident in vitro. Furthermore, the inhibitoryeffect of the suppressor is temporary because the ATPase activityrecovers 9 h after the treatment. A similar effect was observedafter inoculation with M. pinodes but not with a nonpathogenof pea, M. ligulicola. The role of the suppressor in host-parasitespecificity is discussed. (Received April 9, 1991; Accepted August 6, 1991)     

Characters of Pod Anatomy Associated with Resistance to Pod-Shattering in Soybean

Seven characteristics of pod anatomy were studied for theirassociation with resistance to pod-shattering in 16 soybean[Glycine max (L.) Merrill] varieties and strains. The thicknessand length of the bundle cap on the dorsal side of the pod andpod-wall thickness were found to be significantly negativelycorrelated with the degree of pod-shattering. Further statisticalanalyses confirmed that these three anatomical characters werealmost equally important and could potentially serve as criteriafor the selection of resistance to pod-shattering. The identifiedtraits/sites in the pod represent sclerenchymous structuresand may provide the structural basis of resistance to pod-shatteringin soybean.Copyright 1995, 1999 Academic Press Pod, anatomy, shattering, soybean, Glycine max (L.) Merrill     

Effects of low root zone temperatures on the early stages of symbiosis establishment between soybean [Glycine max (L.) Merr.] and Bradyrhizobium japonicum

It is known that low root zone temperatures (RZT) have moreeffect on infection and early nodule development than on nitrogenfixation by soybean [Glycine max (L.) Merr.]. However, therehave been no studies regarding how the low RZT inhibit the infectionstages of soybean. Two controlled environment experiments wereconducted to examine the effect of low RZT on bacterial attachmentto, and infection thread penetration of, soybean root hairs.The experimental designs were (1) plants maintained at 25, 17.5or 15C RZT, or transferred from 25 or 17.5 to 15C RZT at either0.5, 1, 2, or 7d after inoculation (DAI), (2) early symbioticestablishment between soybean and Bradyrhizobium japonicum wasexamined microscopically under three RZT (15, 17.5 and 25C).These results indicated that (1) keeping plants at 25C only0.5 DAI prior to transfer to a 15C RZT accelerates the onsetof N₂ fixation at 15C RZT by 6 d, (2) at RZT between 25 and17.5C the infection processes were progressively delayed astemperature declined, (3) RZT less than 17C strongly inhibitedinfection steps, such that when RZT dropped 8.5C from 25 to17.5C infection initiation was delayed 1 d, while when RZTdropped only 2.5C from 17.5 to 15C, infection initiation wasdelayed another 2 d. Key words: Bradyrhizobium japonicum, low temperature, nodulation, soybean