Microheterogeneity of PSI-E Subunit of Photosystem I in Nicotiana sylvestris

Microheterogeneity of a photosystem I (PSI) subunit encodedby a nuclear gene psaE was examined in Nicotiana sylvestris,with the aid of cDNA cloning, peptide mapping analysis and proteinsequencing. The psaE product of this plant has four isoformswhose mobilities in PAGE are slightly different from each other.We isolated two types of psaE cDNAs from a N. sylvestris cDNAlibrary, and designated the corresponding genes as psaEa andpsaEb, respectively. The psaEa and psaEb genes are 77% homologousat DNA level, and their translation products share 80.4% homologyfor the precursor proteins and 89.1% for the mature forms. Comparativeanalysis of the four isoproteins and the putative products ofthe two psaE genes revealed that two isoproteins out of fourare derived from psaEa gene, and the difference between thesetwo isoproteins lies in the respective presence or absence ofN-terminal alanine. Likewise, the other two proteins are derivedfrom psaEb with similar N-terminal heterogeneity. These resultsindicate that multi-gene organization and heterogeneous N-terminalformation at post-translational level are two possible causesfor PSI subunit polymorphism in isogenic plant lines. (Received October 8, 1993; Accepted November 30, 1993)     

Regulation of Stoichiometry between PSI and PSII in Response to Light Regime for Photosynthesis Observed with Synechocystis PCC 6714: Relationship between Redox State of Cyt b6-f Complex and Regulation of PSI Formation

The effect of the Cyt b₆-f redox state on the PSI formationwas examined with the cyanophyte Synechocystis PCC 6714 by usinga Q-cycle inhibitor, HQNO (2-n-heptyl-4-hydroxyquinoline N-oxide).HQNO inhibited the rapid reduction of flash-oxidized Cyt f,the reaction correlating with the stimulation of PSI formation,on one hand, and accumulated reduced Cyt b₆, on the other, indicatingthat the electron flow in the Q-cycle correlates with regulationof PSI synthesis. HQNO also inhibited the stimulation of PSIformation under PSII light, resulting in a low PSI/PSII ratioeven under PSII light, while the PSI formation under PSI lightwas not suppressed by HQNO. Simultaneous inhibition of Cyt b₆oxidation through the Q-cycle and the stimulated PSI formationby HQNO suggests that an HQNO-sensitive Cyt b₆ oxidation isinvolved in the mechanism of monitoring the state of electrontransport system for regulation of PSI formation. (Received March 3, 1993; Accepted August 9, 1993)     

Crystallization of Photosystem I Complexes from the Thermophilic Cyanobacterium, Synechococcus vulcanus

PSI complexes were isolated from the thermophilic cyanobacterium,Synechococcus vulcanus, by mild detergent treatment of the thylakoidmembranes, purified by sucrose-density gradient centrifugation,and then crystallized. High resolution SDS-PAGE revealed thepresence of the product of the psaI gene in S. vulcanus PSIcomplexes and crystals. Crystals of the PSI complexes retainedall of the components of electron carriers and subunit polypeptides(including PsaX) known in cyanobacteria. (Received July 22, 1998; Accepted October 19, 1998)     

Selective Photoinhibition of Photosystem I in Isolated Thylakoid Membranes from Cucumber and Spinach

The site of photoinhibition at low temperatures in leaves ofa chilling-sensitive plant, cucumber, is photosystem I [Terashimaet al. (1994) Planta 193: 300]. As described herein, selectivephotoinhibition of PSI can also be induced in isolated thylakoidmembranes in vitro. Inhibition was observed both at chillingtemperatures and at 25°C, and not only in the thylakoidmembranes isolated from cucumber, but also in those isolatedfrom a chilling-tolerant plant, spinach. Comparison of theseobservations in vitro to the earlier results in vivo indicatesthat (1) photoinhibition of PSI is a universal phenomenon; (2)a mechanism exists to protect PSI in vivo; and (3) the protectivemechanism is chilling-sensitive in cucumber. The chilling-sensitivecomponent seems to be lost during the isolation of thylakoidmembranes. Very weak light (10–20µmol m^-2 s^-1) wassufficient to cause the inhibition of PSI. About 80% of theoxygen-evolving activity by PSII was maintained even after theactivity of PSI had decreased by more than 70%. This is thefirst report of the selective photoinhibition of PSI in vitro. (Received March 1, 1995; Accepted April 26, 1995)     

Photoinhibition of Photosynthesis during Rain Treatment: Identification of the Intersystem Electron-Transfer Chain as the Site of Inhibition

Continuous wetness of leaves in the light causes a reductionin the carbon exchange rate (CER) in Phaseolus vulgaris L. [Ishibashiand Terashima (1995) Plant Cell Environ. 18: 431]. In this study,we investigated the initial cause of photoinhibition upon applicationof water, designated rain treatment, and we found a large decreasein the rate of electron transport through the whole chain fromwater to methyl viologen via PSII and PSI. In spite of the decreasein the rate of electron transport, there was no decrease inthe activity of either PSI or PSII when these activities weremeasured separately. The intactness of PSI was also confirmedby the absence of any change in the pho-tooxidizable amountof P-700, the reaction centre of PSI, and the intactness ofPSII was confirmed by measurements of Chi fluorescence. Theresults suggest that the inhibition by the rain treatment, whichoccurs at the site between PSI and PSII, might be a novel typeof photoinhibition, unlike the conventional types of photoinhibitionthat involve PSI and PSII. (Received July 29, 1996; Accepted November 28, 1996)     

Stoichiometries of Photosystem I and Photosystem II in Rice Mutants Differently Deficient in Chlorophyll b

Stoichiometries of photosystem I (PSI) and photosystem II (PSII)reaction centers in a cultivar of rice, Norin No. 8, and threechlorophyll b-deficient mutants derived from the cultivar wereinvestigated. Quantitation of PSI by photooxidation of P-700and chromatographic assay of vitamin K₁ showed that, on thebasis of chlorophyll, the mutants have higher concentrationsof PSI than the wildtype rice. Greater increases were observedin the PSII contents measured by photoreduction of Q_A, bindingof a radioactive herbicide and atomic absorption spectroscopyof Mn. Consequently, the PSII to PSI ratio increased from 1.1–1.3in the wild-type rice to 1.8 in chlorina 2, which contains noChl b, and to 2.0–3.3 in chlorina 11 and chlorina 14,which have chlorophyll a/b ratios of 9 and 13, respectively.Measurement of oxygen evolution with saturating single-turnoverflashes revealed that, whereas at most 20% of PSII centers areinactive in oxygen evolution in the wildtype rice, the non-functionalPSII centers amount to about 50% in the three mutant strains.The fluorescence induction kinetics was also analyzed to estimateproportions of the inactive PSII in the mutants. The data obtainedsuggest that plants have an ability to adjust the stoichiometryof the two photosystems and the functional organization of PSIIin response to the genetically induced deficiency of chlorophyllb. (Received July 29, 1994; Accepted February 7, 1996)     

The Accumulation of Protochlorophyllide in Cells of Synechocystis PCC 6714 with a Low PSI/PSII Stoichiometry

Changes in intracellular levels of Chl a precursors were examinedin relation to changes in the PSI/PSII stoichiometry in thecyanophyte Synechocystis PCC 6714. Protochlorophyllide (Pchlide)accumulated markedly in cells with a low PSI/PSII stoichiometrygrown under light that is absorbed by Chl a (PSI light) whereasno accumulation occurred in cells with a high PSI/PSII stoichiometrygrown under light absorbed by phycobilisomes (PSII light). Levelsof Pchlide in cells grown under PSI light decreased rapidlyupon a shift to PSII light. The rapid decrease in Pchlide accompanieda transient increase in chlorophyllide a, indicating that reductionof Pchlide was enhanced by shift to PSII light. The action spectrumindicated that the Pchlide decrease upon the shift to PSII lightdepended on excitation of Pchlide, suggesting that the accumulationof Pchllide was due to limited excitation of Pchlide, so thatPchlide photoreduction, under PSI light. However, comparisonof levels of Pchlide and the photosystem complexes in wild-typePlectonema boryanum with those in a mutant that lacked the darkPchlide reductase (YFC 1004) indicated that dark reduction compensatedfor the limited photoreduction under PSI light. Similar compensationby dark reduction was confirmed with Synechocystis PCC 6714.In cultures of Synechocystis under conditions where Pchlidecould not be photoreduced, accumulation of Pchlide and low PSI/PSIIstoichiometry occurred only when cells were illuminated withlight that preferentially excited PSI. The results indicatethat the low PSI/PSII stoichiometry in cells grown under PSIlight is not a result of inefficient synthesis of Chl a witha reduced rate of Pchlide photoreduction. They suggest furtherthat accumulation of Pchlide under PSI light results from retardationof the Chl a synthesis due to suppression of PSI synthesis. ¹Present address: Tsurukawa 5-15-11, Machida, Tokyo, 195 Japan.     

Nuclear Receptors in Mosquito Vitellogenesis

Vitellogenesis in insects involves the coordinated activityof the fat body, which produces large amounts of yolk proteinprecursors (YP), and oocytes, which specifically accumulatethese proteins. The expression of YP genes is achieved throughstrict sex-, tissue-, and hormone-specific control in the femalefat body. In mosquitoes, expression of YP genes is controlledby 20-hydroxyecdysone (20E). To elucidate the role of 20E inmosquito vitellogenesis, we cloned cDNAs encoding the Aedesaegypti ecdysteroid receptor (AaEcR) and two isoforms of itsheterodimeric partner, the Ultraspiracle homologue (AaUSP).The two AaUSP isoforms differ in their A/B domains and havedistinct expression patterns. The ecdysone regulation of YPgenes likely involves products of early genes. We cloned thegene of the mosquito homologue to the Drosophila early geneE75 (AaE75) belonging to the nuclear receptor superfamily. Kineticsof AaE75 expression correlate with the expression of YP genes,suggesting that AaE75 may have a regulatory role in YP geneexpression. A second nuclear receptor superfamily member, theNGFI-B homologue AaHR38 is implicated in repression of the ecdysone-signalingpathway in the fat body of the previtellogenic female mosquitoat the state-of-arrest. Finally, three isoforms of the hepatocytenuclear factor 4 (HNF-4) homologue AaHNF-4 are differentiallyexpressed in the mosquito fat body during vitellogenesis, suggestingtheir involvement in regulating vitellogenic events in thistissue.     

Changes in Stoichiometry among PSI, PSII and Cyt b6-f Complexes in Response to Chromatic Light for Cell Growth Observed with the Red Alga Porphyra yezoensis

Stoichiometry among 3 thylakoid components, PSI and PSII andCyt b6-f complexes, was determined with the red alga Porphyrayezoensis with special reference to the regulation of PSI/PSIIstoichiometry in response to light regime. The ratio of PSIto PSII abundance was four times greater in thalli grown underorange light which excites mainly phycobilisome, thus PSII,than that under red light which excites preferentially Chl a,thus PSI. Cyt b₆-f abundance remained almost constant. The PSIand PSII content was regulated separately under the two growthlight conditions as was also observed with the red alga Porphyridiumcruentum by Cunningham et al. [(1990) Plant Physiol. 93: 888].This differs from the cyanophyte Synechocystis PCC 6714 whereadjustment occurs only in the PSI content [(1987) Plant CellPhysiol. 28: 1547]. However, results on the marine cyanophyteSynechococcus NIBB 1071 indicate that changes in the PSI/PSIIsoichiometry is similar to red algae. In this species, as inthe red algae, more than one PSII is associated with each phycobilisome.The light regime also induced changes in the phycobiliproteincomposition in Porphyra yezoensis. Under PSII light, phycoerythrinincreased, and phycocyanin decreased, while under PSI lightthe response was reversed. The change suggests an occurrenceof complementary chromatic adaptation. (Received April 8, 1994; Accepted June 1, 1994)     

Properties of Two N-Linked Glycoproteins Associated with the Nuclear Envelope in Mung Bean Hypocotyls

Nuclei from mung bean (Vigna radiata) hypocotyls contained twoglycoproteins of 50 and 49 kDa, respectively, that reacted withconcanavalin A. The glycoproteins were released from the nuclearenvelope by treatment with 2 M KCl but not with nucleases. Theglycoproteins, tentatively named gp50 and gp49, were isolatedand characterized. Gel-permeation chromatography suggested thatgp50 and gp49 seem to exist as a complex with other components.The glycoproteins could be detected only in the nuclear fractionby immunoblot analysis with specific antibodies, and they werenot detected in endoplasmic reticulum, plasma membrane, vacuolarmembrane or mitochondria. Agglutinin I from Ulex europeaus,peanut agglutinin, soybean agglutinin and wheat germ agglutininall failed to bind to the glycoproteins. Treatment with glycopeptidaseF removed all oligosaccharides from the glycoproteins and decreasedtheir molecular masses by about one thousand daltons each. Theseresults suggest that the glycoproteins contained N-linked, high-mannose-typeoligosaccharides with six or seven hexose residues. gp50 andgp49 seemed to be isoforms of a single glycoprotein becausethe two proteins had some common properties. Nuclear fractionsfrom azuki bean (Phaseolus angularis) and soybean (Glycine max)contained proteins that were immunologically similar to gp50and gp49. (Received March 18, 1995; Accepted May 24, 1995)     

Changes in Photosystem Stoichiometry in Response to Environmental Conditions for Cell Growth Observed with the Cyanophyte Synechocystis PCC 6714

Changes in photosystem stoichiometry in response to shift ofenvironments for cell growth other than light regime were studiedwith the cyanophyte Synechocystis PCC 6714 in relation to thechange induced by light-quality shift. Following two environment-shiftswere examined: the shift of molecular form of inorganic carbonsource for photosynthesis from CO₂ to HCO₃^– (CO₂ stress)and the increase in salinity of the medium with NaCl (0.5 M)(Na⁺ stress). Both CO₂ and Na⁺ stresses induced the increasein PSI abundance resulting in a higher PSI/PSII stoichiometry.CO₂ stress was found to elevate simultaneously Cyt c oxidaseactivity (V_max). The feature was the same as that caused bylight-quality shift from preferential excitation of PSI to PSII(light stress) though the enhancement by either stress was smallerthan that by light stress. Under our experimental conditions,PSI/PSII stoichiometry appeared to increase at a fairly constantrate to the basal level even when the basal level had been differentlydetermined by the light-quality. Enhancing rates for PSI/PSIIstoichiometry and for Cyt c oxidase activity were also similarto each other. Since the two stresses affect the thylakoid electrontransport similarly to the shift of light-quality, we interpretedour results as follows: three environmental stresses, CO₂, Na⁺,and light stresses, cause changes in electron turnover capacityof PSI and Cyt c oxidase under a similar, probably a common,mechanism for monitoring redox state of thylakoid electron transportsystem. ¹On leave from Department of Biology, College of Natural Science,Kyngpook National University, Taegu 702-701, Korea. ₂Present address: Department of Marine Bioscience, Fukui Pre-fecturalUniversity, Obama, Fukui, 917 Japan.     

RMA1 an Arabidopsis thaliana Gene Whose cDNA Suppresses the Yeast secl5 Mutation, Encodes a Novel Protein with a RING Finger Motif and a Membrane Anchor

To identify molecules that function in the plant secretory pathway,we screened for Arabidopsis thaliana cDNA clones that complementthe temperature-sensitive (ts), secretion-deficient sec15 mutationof yeast Saccha-romyces cerevisiae. RMA1, one of the genes obtainedin this screening, suppressed not only the ts growth of sec15but also its secretory defect. RMA1 is not a structural homologueof SEC15 but encodes a novel 28 kDa protein with a RING fingermotif and a C-terminal membrane-anchoring domain. Mutationalanalysis indicates that the RING finger motif of RMA1 is importantfor its suppression activity. In Arabidopsis plant, RMA1 isubiquitously expressed. A search for homologous proteins inthe database revealed that Arabidopsis, nematode, mouse andhuman possess close homologues of RMA1. (Received January 5, 1998; Accepted March 9, 1998)     

Nuclear DNA Amounts in Angiosperms

Bennett and Smith (Philosophical Transactions of the Royal Societyof London B274:227-274; B334: 309-345) and Bennett, Smith andHeslop-Harrison (Proceedings of the Royal Society of London,B216: 179-199) published lists of nuclear DNA amounts estimatedfor 1612 angiosperm species collected from 163 sources datedbetween 1951 and 1986. Subsequently, interest in genome sizein angiosperms and its significance has continued, and manynew DNA estimates were published during 1986-1994. Their inaccessibility,and the flow of enquiries for such information, shows that afurther compilation is needed. This paper presents a supplementarylist of nuclear DNA C-values for 105 sources for 899 angiospermspecies not listed in the above-mentioned compilations, plus284 additional estimates for 208 species already listed by them.The data are assembled primarily for reference purposes, withspecies listed in alphabetical order, rather than by any taxonomicscheme. Some advantages and limitations of flow cytometry, nowincreasingly used to quantify DNA C-values in plants, are reviewed.Recent reports regarding the occurrence and extent of intraspecificvariation in genome size are also discussed. While some examplesare real, others reflect technical shortcomings. Work has begunto combine the genome size data compiled in this and the above-mentionedpapers into a unified data base, and to present the informationin separate lists, with species in alphabetical and systematicorders, respectively. DNA C-values are now known for 1% of theworld's angiosperm flora, but improved representation of taxonomicgroups, geographical regions and plant life forms is urgentlyneeded.Copyright 1995, 1999 Academic Press Angiosperm DNA amounts, DNA C-values, nuclear genome sizes, intraspecific variation, flow cytometry, reference lists, genome size database     

The role of cis-regulatory motifs and genetical control of expression in the divergence of yeast duplicate genes

Expression divergence of duplicate genes is widely believedto be important for their retention and evolution of new function,although the mechanism that determines their expression divergenceremains unclear. We use a genetical genomics approach to exploredivergence in genetical control of yeast duplicate genes createdby a whole-genome duplication that occurred about 100 MYA andthose with a younger duplication age. The analysis reveals thatduplicate genes have a significantly higher probability of sharingcommon genetic control than pairs of singleton genes. The expressionquantitative trait loci (eQTLs) have diverged completely fora high proportion of duplicate pairs, whereas a substantiallylarger proportion of duplicates share common regulatory motifsafter 100 Myr of divergent evolution. The similarity in bothgenetical control and cis motif structure for a duplicate pairis a reflection of its evolutionary age. This study revealsthat up to 20% of variation in expression between ancient duplicategene pairs in the yeast genome can be explained by both cismotif divergence (8%) and by trans eQTL divergence (10%). Initially,divergence in all 3 aspects of cis motif structure, trans-geneticalcontrol, and expression evolves coordinately with the codingsequence divergence of both young and old duplicate pairs. Thesefindings highlight the importance of divergence in both cismotif structure and trans-genetical control in the diverse setof mechanisms underlying the expression divergence of yeastduplicate genes.     

Chromatic Regulation in Chlamydomonas reinhardtii: Time Course of Photosystem Stoichiometry Adjustment Following a Shift in Growth Light Quality

Photosystem stoichiometry adjustments in Chlamydomonas reinhardtiiwere induced upon a sudden shift in the light quality duringcell growth. Reversible changes in the PSI/PSII ratio were acompensation response to changes in the balance of light absorptionby the two photosystems. Quantitations of PSII, Cyt b₆-f complexand PSI revealed a constancy in the cellular content of PSIIand the Cyt b₆-f complex, and variable amounts of PSI in C.reinhardtii. These results strengthen the notion that PSI isthe thyla-koid component subject to chromatic regulation andresponsible for the adjustment and optimization of the PSI/PSII ratio in the thylakoid of oxygenic photosynthesis. Additionalresults, obtained upon the use of protein biosynthesis translationinhibitors (chloramphenicol and cyclohex-imide), suggested thata chromatically-induced lowering of the PSI/PSII ratio in C.reinhardtii occurs by suppression of de novo biosynthesis ofPSI components and, therefore, by dilution of the PSI complexin the thylakoid membrane, rather than by active degradationof assembled PSI in chlo-roplasts. (Received November 8, 1996; Accepted December 6, 1996)     

The Regulatory Functions of the rolB and rolC Genes of Agrobacterium rhizogenes are Conserved in the Homologous Genes (Ngrol) of Nicotiana glauca in Tobacco Genetic Tumors

To compare patterns of expression between the Ngrol genes ofN. glauca and the Rirol genes of Agrobacterium rhizogenes, weperformed fluorometric and histochemical analysis of transgenicgenetic tumors on the hybrid of Nicotiana glauca x N. langsdorffü(Fl) that harbored a rß- glucuronidase (GUS) reportergene fused to the promoter of NgrolB, NgrolC, RirolB or RirolC The promoters of NgrolB and NgrolCNgrolC had 2- to 3-fold loweractivity than those of RirolB and RirolC However, the changesin patterns of GUS activity caused by deletion of NgrolB andNgrolCpromoters were similar to those of RirolB and RirolC promoters.This result suggests that the cis-acting sequences that regulatethe level of expression of RirolB and RirolC are conserved inthe NgrolB and NgrolC promoters. Furthermore, an auxin dependent(NAA-dependent) increase in GUS activity was observed in thecase of NgrolB-GUS and RirolB-GUS. Histochemical analysis showedGUS activity encoded by both NgrolB-GUS and RirolB-GUS in normal-typeFl transgenic plants was located in meristematic zones, whilethat encoded by NgrolC-GUS and RirolC-GUS was detected mainlyin vascular systems of various organs. Thus, the patterns ofexpression of the Ngrol genes were the same as those of theRirol genes in terms of promotion by auxin and tissue-specificity,indicating that regulatory mechanisms for both sets of geneshave been conserved during the evolution of the genus Nicotianaafter transfer from a progenitor of Agrobacterium to that ofNicotiana. (Received May 2, 1995; Accepted June 13, 1995)     

Photoinhibition of Photosystem I: Its Physiological Significance in the Chilling Sensitivity of Plants

Photoinhibition was defined originally as the decrease in photosyntheticactivity that occurs upon excess illumination. The site of photoinhibitionhas generally been considered to be located in PSII. However,a novel type of photoinhibition has recently been characterizedin chillingsensitive plants. This photoinhibition occurs underrelatively weak illumination at chilling temperatures and themain site of damage is in PSI. The photoinhibition of PSI isinitiated by the inactivation of the acceptor side, with thesubsequent destruction of the reaction center and the degradationof the product of the psaB gene, which is one of the two majorsubunit polypeptides of the PSI reaction center complex. Chillingand oxidative stress (the presence of reactive species of oxygen)are characteristic requirements for the photoinhibition of PSIin vivo. (Received December 4, 1995; Accepted March 11, 1996)