Computational analysis of RNA editing sites in plant mitochondrial genomes reveals similar information content and a sporadic distribution of editing sites

A computational analysis of RNA editing sites was performedon protein-coding sequences of plant mitochondrial genomes fromArabidopsis thaliana, Beta vulgaris, Brassica napus, and Oryzasativa. The distribution of nucleotides around edited and uneditedcytidines was compared in 41 nucleotide segments and included1481 edited cytidines and 21,390 unedited cytidines in the 4genomes. The distribution of nucleotides was examined in 1,2, and 3 nucleotide windows by comparison of nucleotide frequencyratios and relative entropy. The relative entropy analyses indicatethat information is encoded in the nucleotide sequences in the5 prime flank (–18 to –14, –13 to –10,–6 to –4, –2/–1) and the immediate 3prime flanking nucleotide (+1), and these regions may be importantin editing site recognition. The relative entropy was largewhen 2 or 3 nucleotide windows were analyzed, suggesting thatseveral contiguous nucleotides may be involved in editing siterecognition. RNA editing sites were frequently preceded by 2pyrimidines or AU and followed by a guanidine (HYCG) in themonocot and dicot mitochondrial genomes, and rarely precededby 2 purines. Analysis of chloroplast editing sites from a dicot,Nicotiana tabacum, and a monocot, Zea mays, revealed a similardistribution of nucleotides around editing sites (HYCA). Thesimilarity of this motif around editing sites in monocots anddicots in both mitochondria and chloroplasts suggests that amechanistic basis for this motif exists that is common in thesedifferent organelle and phylogenetic systems. The preferredsequence distribution around RNA editing sites may have an importantimpact on the acquisition of editing sites in evolution becausethe immediate sequence context of a cytidine residue may rendera cytidine editable or uneditable, and consequently determinewhether a T to C mutation at a specific position may be correctedby RNA editing. The distribution of editing sites in many protein-codingsequences is shown to be non-random with editing sites clusteredin groups separated by regions with no editing sites. The sporadicdistribution of editing sites could result from a mechanismof editing site loss by gene conversion utilizing edited sequenceinformation, possibly through an edited cDNA intermediate.     

Delimitation of Epacridaceae: Preliminary Molecular Evidence

Comparative sequence data for the chloroplast encodedrbc L genehas been obtained for species representing the basal lineagesin the proposed phylogeny of Powellet al. (Annals of Botany77:305–315, 1996), with the aim of testing the delimitationof the family and the validity of the supra-generic taxa proposed,and estimating relationships within the family. Cladistic analysesindicate that EpacridaceaesensuPowellet al. (1996) is not monophyletic.Lebetanthus,a monotypic genus from South America commonly placed withinthe family, mostly near the Tasmanian endemicPrionotes, is shownto be closer toGaultheriaand other members of the ericaceoustribe Andromedeae. The hypothetical phylogeny of Powellet al.(1996) is evaluated in the light of this preliminary analysisof relationships within the family. The data do not supportthe recognition of the two subfamilies, Richeoideae and Epacridoideae,of Watson Epacridaceae; Ericales; molecular systematics; molecular systematics; rbc L     

A COMPARATIVE STUDY OF THE MATURE SPERMATOZOA OF BULINUS AFRICANUS (KRAUSS, 1848) AND B. GLOBOSUS (MORELET, 1866) (GASTROPODA: PLANORBIDAE)

The morphology of the mature spermatozoa of the closely-relatedfreshwater pulmonates, Bulinus africanus and Bulinus globosus,was examined using scanning and transmission electron microscopy.A comparison showed no useful differences that could be usedto distinguish the species though they did differ in one respectfrom Bulinus tropicus, the only other bulinid for which spermmorphology is known. (Received 12 August 1996; accepted 6 December 1996)     

Genetic Analysis of the Effects of Polar Auxin Transport Inhibitors on Root Growth in Arabidopsis thaliana

Polar auxin transport inhibitors, including N-1-naphthylphthalamicacid (NPA) and 2,3,5-triiodobenzoic acid (TIBA), have variouseffects on physiological and developmental events, such as theelongation and tropism of roots and stems, in higher plants.We isolated NPA-resistant mutants of Arabidopsis thaliana, withmutations designated pir1 and pir2, that were also resistantto TIBA. The mutations specifically affected the root-elongationprocess, and they were shown ultimately to be allelic to aux1and ein2, respectively, which are known as mutations that affectresponses to phytohormones. The mechanism of action of auxintransport inhibitors was investigated with these mutants, inrelation to the effects of ethylene, auxin, and the polar transportof auxin. With respect to the inhibition of root elongationin A. thaliana, we demonstrated that (1) the background levelof ethylene intensifies the effects of auxin transport inhibitors,(2) auxin transport inhibitors might act also via an inhibitorypathway that does not involve ethylene, auxin, or the polartransport of auxin, (3) the hypothesis that the inhibitory effectof NPA on root elongation is due to high-level accumulationof auxin as a result of blockage of auxin transport is not applicableto A. thaliana, and (4) in contrast to NPA, TIBA itself hasa weak auxin-like inhibitory effect. (Received April 12, 1996; Accepted September 2, 1996)     

Mechanisms of Oxidation of Inorganic Electron Donors in Autotrophic Bacteria

The enzymatic reactions involved in the oxidation of sulfide,sulfite, thiosulfate, ferrous ions, ammonia, and nitrite arereviewed for Chlorobium limicola f. thiosulfato philum, Thiobacillusnovellus, Thiobacillus ferrooxidans, Nitrosomonas europaea,and Nitrobacter winogradskyi. The properties of the purifiedredox enzymes and of proteins that participate in the oxidationof the inorganic compounds in these autotrophic bacteria aresummarized, and the mechanisms of the oxidation of the inorganiccompounds are discussed on the basis of the interactions betweenthe redox enzymes and carriers. (Received December 21, 1995; Accepted May 29, 1996)     

Cloning and Functional Expression of a Novel Geranylgeranyl Pyrophosphate Synthase Gene from Arabidopsis thaliana in Escherichia coli

A gene encoding a novel geranylgeranyl pyrophosphate (GGPP)synthase from Arabidopsis thaliana has been identified and termedGGPS5. The gene has been sequenced and expressed in Escherichiacoli. The deduced amino acid sequence showed 64.5% and 57.5%identity with a putative GGPP synthase from Arabidopsis andCapsicum annuum, respectively. GGPP enzymatic activity was detectedin E. coli cells expressing the GGPS5 gene in two differentways. One was the direct measurement of GGPP synthase activityin cell extracts and the other was the yellow color productionof cells when the GGPS5 gene was co-expressed with crtB, crtI,crtX, crtY and crtZ genes derived from Erwinia uredovora. (Received May 20, 1996; Accepted December 14, 1996)     

Distribution of Calanus species in Kongsfjorden, a glacial fjord in Svalbard

The distribution of Calanus species was investigated in Kongsfjordenin summer of 1996 and 1997. In both years Calanus finmarchicusand Calanus glacialis dominated, although the boreal C. finmarchicuswas more abundant than the Arctic C. glacialis in 1997. Thiscoincided with a 2°C higher water temperature at 50 m in1997, indicating stronger influence of Atlantic origin waterthat year. Advected Calanus finmarchicus occurred in deep andsubsurface layers of the outer fjord in 1996 (200 ind. m^-3,mainly CIII). A less abundant local population aggregated insurface layers of the inner fjord (100 ind. m^-3). Similarly,advected C. finmarchicus occurred in subsurface layers in 1997(446 ind. m^-3, mainly CIII and CIV) and a local population insurface layers (183 ind. m^-3, mainly CI). Calanus glacialisin 1996 aggregated as CII and CIII in the deep layers of theouter fjord (272 ind. m^-3), whereas CIII–CV were abundant(216 ind. m^-3) in cold surface waters of the inner fjord. In1997 C. glacialis (mostly CIII–CV) was more abundant inthe outer than in the inner part of the fjord (40 and 192 ind.m^-3, respectively). Within Kongsfjorden, Calanus finmarchicusneeds one year to complete its life cycle, whereas Calanus glacialisneeds two. Calanus hyperboreus seems to be an expatriate inthe fjord system.     

A Lectin from Leaves of Neoregelia flandria Recognizes D-Glucose, D-Mannose and N-Acetyl D-glucosamine, Differing from the Mannose-Specific Lectins of Other Monocotyledonous Plants

A mannose-specific lectin was isolated from leaves of Neoregeliaflandria, an ornamental plant that belongs to Bromeliaceae,a family of monocotyledons. The amino acid composition and molecularmass of the lectin were similar to those of mannose-specificlectins from other monocotyledons. However, in a test to examinethe inhibition of hemagglutination, it became apparent thatthe isolated lectin recognized D-glucose and N-acetyl D-glucosaminein addition to D-mannose, unlike mannose-specific lectins fromthe monocotyledons that have been reported to date. (Received May 17, 1996; Accepted August 19, 1996)     

Transient Increase in the Level of mRNA for a Germin-Like Protein in Leaves of the Short-Day Plant Pharbitis nil during the Photoperiodic Induction of Flowering

A cDNA corresponding to an in vivo labeled protein whose levelincreased during flower-inductive darkness in the cotyledonof the short-day plant Pharbitis nil Choisy cv. Violet was isolatedand characterized. The deduced amino-acid sequence of the protein(designated PnGLP; P. nil germin-like protein) showed homologyto that of a GLP of Sinapis alba, a leaf protein, the mRNA accumulationof which showing circadian oscillations. PnGLP mRNA was detectedspecifically in the cotyledon and leaf, in particular, in theyoung expanded cotyledon and leaf. Accumulation of PnGLP mRNAincreased transiently during flower-inductive darkness and peakedat a time that corresponded approximately to the critical nightlength. This mRNA peak was reduced by a brief exposure to redlight at the 8th hour of darkness. The level of PnGLP mRNA peakedabout 10 h from the beginning of the dark period, whereas itwas reported that the level of mRNA for GLP of a long-day plantS. alba increased about 14 h from the beginning of the lightperiod. Thus, the different time courses of accumulation ofthe mRNAs for leaf-specific GLPs appear to reflect the differencesin photoperiodic responses of each plant. (Received May 16, 1996; Accepted July 5, 1996)     

The Level of mRNA Transcribed from psaL, Which Encodes a Subunit of Photosystem I, Is Increased by Cytokinin in Darkness in Etiolated Cotyledons of Cucumber

A cDNA clone for an mRNA whose level increased within 2 h ofthe start of treatment with N⁶-benzyladenine in etiolated cotyledonsof cucumber was isolated by differential hybridization. ThecDNA was homologous to psaL, which encodes subunit XI (PSI-L)of photosystem I. The accumulation of psaL mRNA was specificallyinduced by cytokinins or light. (Received April 10, 1996; Accepted July 31, 1996)     

The role of apical development around the time of leaf initiation in determining leaf width at maturity in wheat seedlings (Triticum aestivum L.) with impeded roots

High soil resistance to root penetration (measured as penetrometerresistance, R_s slows down leaf growth and reduces mature leafsize in wheat seedlings {Triticum aestivum L.). Underlying changesin the kinetics of cell partitioning and expansion and in thesize and organization of mature cells were reported in companionpapers (Beemster and Masle, 1996; Beemster et al., 1996). Inthe present study, the relationships between apex growth, primordiuminitiation and expansion were analysed for plants grown at contrastingR_s, focusing on a leaf whose whole development proceeded afterthe onset of root impedance (leaf 5). High R_s reduced the rates of apex and leaf development, butdid not appear to have immediate effects on the pattern of developmentof the newly initiated phytomers. During an initial short period,the rate of development of a leaf primordium and associatednode were related to plastochronic age, according to similarrelationships (slopes) at the two R_s. Effects on developmentalpatterns were first detected on phytomer radial expansion duringplastochron 2. The ontogenetic pattern of leaf elongation wasaffected later, during the next few plastochrons preceding leafemergence (‘post-primordial stage’). It is concludedthat a reduction in the number of formative divisions and inthe number of proliferative cells along the intercalary mer-istemreported earlier (Beemster and Masle, 1996; Beemster et al.,1996) is not related to the size of the apical dome at leafinitiation nor to the size and number of meristematic cellsinitially recruited to the leaf primordium, which were all unaffectedby R_s. Rather they are generated at the primordial and post-primordialstages. Key words: Wheat, apex development, leaf primodium development, mature leaf width, root impedance     

Fluxes of diatoms in the Dona Paula Bay, west coast of India

Sediment traps were deployed at a station in the Dona PaulaBay to collect sedimenting particles at weekly intervals fromNovember to May during 1995–1997. Sedimented particleswere analysed for total diatom flux, chlorophyll a (Chl a) andparticulate organic carbon (POC). The highest diatom flux wasrecorded in April–May for both the years. Fluxes of diatomsvaried from0.6 x 10⁴ cells m^–2 day^–1 (November 1995)to 121.47 x 10⁴ cells m^–2 day^–1 (December 1996).In all, 19 diatom genera were identified in the sedimented material.Navicula, Nitzschia, Pleurosigma, Licmophora, Coscinodiscus,Rhizosolenia and Surirella were the most abundant genera inthe sedimented material throughout the sampling period. Meanflux of POC and diatom carbon was 251 and 0.39 mg C m^–2day^–1, respectively. The diatom carbon accounted for 0.15%of the POC flux. Mass flux of diatoms showed significant negativecorrelation with the concentration of nitrate and phosphate.This suggests that the nutrient concentration played an importantrole in influencing the sedimentation of diatoms at this coastalstation.     

Identification of the chlB Gene and the Gene Product Essential for the Light-Independent Chlorophyll Biosynthesis in the Cyanobacterium Plectonema boryanum

We cloned a 6.0-kb HindIII fragment from the cyanobacteriumPlectonema boryanum using the chloroplast chlB (ORF513) geneof the liverwort (Marchantia polymorpha) as a probe. An openreading frame (ORF508) encoding a polypeptide of 508 amino acidresidues was found within the nucleotide sequence of the 4,437-bpHindIII-EcoRV subfragment. The deduced amino acid sequence ofORF508 shows very high similarity to that encoded by the liverwortchlB gene (72.7%). A mutant, YFB14, in which ORF508 was inactivatedby the insertion of a kanamycin-resistance cartridge, was unableto synthesize chlorophyll, accumulating protochlorophyllidein darkness while synthesizing chlorophyll normally in the light.Thus, the chlB gene is the third gene that is essential forthe light-independent reduction of protochlorophyllide. Theother two genes are chlL and chlN, and the results suggest thatthe light-independent protochlorophyllide reductase consistsof at least three subunits, which are encoded by chlL, chlNand chlB. Using an antiserum prepared against a ChlB-6xHis fusionprotein expressed in Escherichia coli, we detected a proteinwith an apparent molecular weight of 58,000 in the membranefraction of the cyanobacterium. These results indicate thateither the cytoplasmic or thylakoid membranes could be the siteof the light-independent reduction of protochlorophyllide. (Received November 16, 1995; Accepted February 7, 1996)     

Mutations in the SGR4, SGR5 and SGR6 Loci of Arabidopsis thaliana Alter the Shoot Gravitropism

Shoots of higher plants grow upward in response to gravity.To elucidate the molecular mechanism of this response, we haveisolated shoot gravitropism (sgr) mutants in Arabidopsis thaliana.In this report, we describe three novel mutants, sgr4-1, sgr5-1and sgr6-1 whose inflorescence stems showed abnormal gravitropicresponses as previously reported for sgr1, sgr2 and sgr3. Thesenew sgr mutations were recessive and occurred at three independentgenetic loci. The sgr4-1 mutant showed severe defect in gravitropismof both inflorescence stem and hypocotyl but were normal inroot gravitropism as were sgr1 and sgr2. The sgr5-1 and sgr6-1mutants showed reduced gravitropism only in inflorescence stemsbut normal in both hypocotyls and roots as sgr3. These resultssupport the hypothesis that some mechanisms of gravitropismare genetically different in these three organs in A. thaliana.In addition, these mutants showed normal phototropic responses,suggesting that SGR4, SGR5 and SGR6 genes are specifically involvedin gravity perception and/or gravity signal transduction forthe shoot gravitropic response. (Received November 21, 1996; Accepted February 17, 1997)     

DIETARY INDUCTION OF OPISTHOBRANCH MORPHOLOGY: PLACIDA DENDRITICA (ALDER & HANCOCK, 1843) ON DIFFERENT GREEN ALGAL HOSTS

The relationship of phenotypic morphological and behaviouralvariation was investigated for the ascoglossan (=sacoglossan)opisthobranch Placida dendritica. Morphological attributes weredocumented for individual slugs collected from three algal hostspecies along the central Oregon coast, USA, the green algaeCodium setchellii, C. fragile, and Bryopsis corticulans. Becauseindividuals on B. corticulans generally were substantially largerthan conspecifics on Codium spp., the species appeared polymorphic.Comparably sized slugs from different host genera also differedsignificantly in morphology: individuals from Codium spp. hadmore numerous, shorter cerata on wider bodies than did conspecificsfrom B. corticulans. Principal components analysis indicatedthat despite considerable overlap of morphological characters,ceratal shape features were diagnostic of algal host. When slugscollected from C. fragile were maintained on C. fragile andB. corticulans for 4 days in the laboratory, the size and shapeof slugs fed different diets diverged, and changes were consistentwith field data. Thus, phenotype was labile and changed as aresult of modified diet. Although a variety of molluscs exhibitdietary induction of morphology, a general awareness of thephenomenon has only recently begun to emerge, despite its importantecological and taxonomic implications. (Received 5 February 1996; accepted 2 July 1996)     

4-Methylumbelliferyl glycosides of N-acetyl 4-thiochito-oligosaccharides as fluorogenic substrates for chitodextrinase from Vibrio furnissii

The degradation of chitin involves a diverse array of enzymes,some with overlapping substrate specificities. In order to distinguishbetween different types of enzymes, specific substrates areneeded. Toward this end, two new fluorogenic substrates containingthio-glycosidic linkages, 4-methylumbelliferyl N,N'-diacetyl-4-thio-ß-chitobioside(Mu-TCB) and N,N',N'-triacetyl-4,4'-dithio-ß- chitotrioside(Mu-TCT) are described. The substitution of the glycosidic oxygens(except the one that links oligosaccharide with the fluorogenicaglycon) with a sulfur atom resulted in resistance of thesecompounds to N-acetyl-hexosaminidases while they were specificsubstrates for the newly discovered chitodextrinase from Vibriofurnissii (Keyhani,N.O. and Roseman,S. (1996) J. Biol. Chem.,271, 33414–33424) and some bacterial chitinases. The enzymekinetics of these 4-S-linked substrates, Mu-TCB and MuTCT, aswell as the O-linked 4-methylumbelliferyl N,N'diacetyl-ß-chitobioside(Mu-CT) and N,N',N'-triacetyl-ß-chitotrioside (Mu-CT)with the chitodextrinase were studied and compared. The usefulnessof the substrates for screening for chitodextrinase and/or chitinaseactivity was demonstrated. chitodextrinase chitinase fluorometric assay 4-methylumbelliferyl glycoside thiochitooligosaccharide     

The Mode of Expression and Promoter Analysis of the arcA Gene, an Auxin-Regulated Gene in Tobacco BY-2 Cells

The arcA, a member of the G protein rß-subunit family,was isolated from tobacco BY-2 cells as an auxin-responsivegene. Characterization of arcA, which should help to elucidatethe function of the gene product in the plant cells, was performedwith emphasis on the mode of expression and the analysis ofits promoter. Accumulation of the arcA message was detectedonly after treatments with auxins and not after treatments withother phytohormones or CdCl₂, implying that responsiveness ofarcA was exclusive to auxin. The putative arcA promoter regionwas fused to a reporter gene for rß-glucuronidase(GUS), and transient expression was analyzed in tobacco BY-2cells. Two series of arcA promoter/GUS chimeric genes were constructed.One consisted of a set of 5' nested deletions of the arcA promoterconnected to the gene for GUS and the other consisted of a varietyof the arcA promoter fragments fused to a minimal promoter-GUSconstruct. The results indicated that the promoter sequencecovering four sets of direct repeats (– 562 to –167)was necessary for the sufficient response of arcA promoter toauxin in BY-2 cells. Moreover, irrespective of auxin treatment,elevated activity of GUS driven by this promoter fragment wasdetected, a result that implies that this region behaves anenhancer in BY-2 cells. (Received September 30, 1995; Accepted March 1, 1996)