Macromolecular physiology of plastids XIV.Viridis mutants in barley: Genetic, fluoroscopic and ultrastructural characterisation

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The properties and composition of bundle sheath and mesophyl! thylakoids from maize leaves are compared. This was possible because of the isolation of large amounts of purified material obtained by enzymatic digestion of mechanically disrupted leaves. Bundle sheath thylakoids from mature leaves, lack the chlorophyll-proteins and polypeptides associated with the reaction centre ofphotosystem II. They do, however, contain significant amounts of LHCII, which transfers excitation energy to photosystem I. LHCII isolated from bundle sheath thylakoids had a different freeze-fracture ultrastructure and a different polypeptide composition from LHCII isolated from mesophyll thylakoids, indicating a differential expression of the LHCII gene family in mesophyll and bundle sheath cells of maize leaves. INTRODUCI'IONThe photosynthetic membranes of higher plants are now thought to be characterised by extreme lateral heterogeneity with respect to composition and function, with all photosystem I (PSI) in the unstacked membrane regions (stroma la'mellae) and most, if not all photosystern II (PSII) in the appressed regions (grana) (1). The locations of some of the minor components remain in doubt because the methods used for the isolation of stroma and grana lamellae requires either mechanical disruption or detergent fractionation. Both procedures can generate artefacts due to cross-contamination or selective removal of some components.Chloroplasts containing no grana lamellae can therefore be very useful for studying thylakoid composition since their isolation avoids these procedures. Such chloroplasts are found in the bundle sheath cells (l l) of some C4 plants (e.g., maize and sorghum) characterised by high levels of NADPH-malic enzyme. In the leaves of such plants, CO2 is fixed by phosphoenolpyruvate c~rboxylase in mesophyll cells and transported as malate to bundle sheath cells, where decarboxylation yields CO2 and NADPH. The Abbreviations: BS = bundle sheath; Chl = chlorophyll; DCPIP = 2,6-di-chlorophenol indophenol; DPC = 1,5-diphenyi carbazide; EDTA = ethylenediamine tetraacetate; Hepes = N-2-(hydroxyethyl)-piperazine-N'-2-ethanesulphonic acid; LHC = light-harvesting complex; Mes = 2-(N-morpholino)-ethanesulphonic acid; PS = photosystem; SDS = sodium dodecyl sulphate; Tricine = N-(tris-(hydroxymethyl)-methyl)-glycine.Springer-Verlag

Section: Othermentioning

confidence: 99%

Differential expression of LHCII genes in mesophyll and bundle sheath cells of maize

Bassi

Simpson

1986

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“…The fluorescence signal was measured using a Hamamatsu R928 photomultiplier and a signal amplifier (Applied Photophysics, London) and recorded on a digital oscilloscope (Nicolet Explorer II Digital Osscilloscope, Madison, Wisconsin). Low temperature fluorescence emission spectra were obtained as in (44). stained for 3 hours at 60 ~ with 2% uranyl acetate.…”

Section: Photochemical Assaysmentioning

confidence: 99%

A thylakoid polypeptide involved in the reconstitution of photosynthetic oxygen evolution

Møller

Høj

1983

Sonication of barley thylakoids in a high salt buffer released three polypeptides of M r 32,000, 23,000, and 13,500 which were purified to homogeneity by chromatofocusing.Highly purified inside-out photosystem II preparations were obtained by French Press treatment or by Triton X-100 fractionation of stacked lamellar systems. Both preparations are composed of pairs of appressed membrane sheets. In the French Press preparation, the majority of these membrane pairs are sealed whereas they are predominantly unsealed in the Triton X-100 preparation. Both preparations are able to evolve oxygen and show reversed proton pumping. The oxygen evolving capacity and variable fluorescence of both preparations were lost upon washing with high salt buffer. This treatment also removed three polypeptides at M r 32,000, 23,000, and 13,500. The inactivated preparations were reconstituted with respect to oxygen evolution and variable fluorescence by rebinding of the isolated M r 23,000 polypeptide. The M r 32,000 and 13,500 polypeptides had no effect on reconstitution.Abbreviations: Chl = chlorophyll; chl-P = chlorophyll-protein; DCCD = dicyclohexyl carbodiimide: DCMU = 3-(3,4-dichlorophenyl)-l,l-dimethylurea; DCPIP = 2,6-dichlorophenol indophenol; EPR = electron paramagnetic resonance; Hepes = N-2-hydroxyethylpiperazine-N'-2-ethane sulfonic acid; PPBQ = phenyl-p-benzoquinone: PS = photosystem; SDS-PAGE = sodium dodecylsulfate polyacrylamide gel electrophoresis; TMBZ = 3.Y,5,5'-tetramethylbenzidine; Tricine = N-(2-hydroxy-l,l-bis(hydroxymethyl)ethyl)glycine: Tris = tris-(hydroxymethyl)-amino methane 0105/1938/83/0048/0161/$05.00

“…While viridis-n 34 is defective in photosystem I activity (17), viridis-e 64 is low in both photosystems (30). The two mutants can be distinguished by studying the variable fluorescence, viridis-e 64 lacking variable fluorescence while viridis-n 34 has variable fluorescence due to a normally functioning photosystem II (26).…”

Section: Photosynthetic Nuclear Gene Mutantsmentioning

confidence: 99%

“…Two areas in particular have benefitted from the development of practical measurements and screening procedures based on chlorophyll fluorescence. Firstly, the detection of photosynthetic mutants (1,11,15,26,32) and secondly, monitoring the development of injury to plant cells resulting from an applied stress including chilling (12,28), freezing (13,19), high temperatures (21,23,27,31), water (33) or oxygen (24) deficits, and ozone toxicity (25).…”

Section: Introductionmentioning

confidence: 99%

Chlorophyll fluorescence photography to detect mutants, chilling injury and heat stress

Gibbons

Smillie

1980

Chlorophyll fluorescence photography with high speed colour film has been used to examine: (i) kinetics of the rise and fall of variable fluorescence in barley leaves, (ii) detection of high fluorescing mendelian and maternally inherited mutants of barley, (iii) detection of protochlorophyllide-deflcient and protochlorophyllideaccumulating mutants of barley, (iv) heat stress by photographing the increase of initial fluorescence in heated leaves, and (v) decrease in variable fluorescence in leaves of peanut, tomato and tomato-potato somatic hybrids associated with the development of chilling injury. Filter combinations and a light source for photographing chlorophyll fluorescence are described. The relative intensity and kinetics of the fluorescence from the plants photographed was also measured, so that this could be correlated with fluorescence as detected by photography, and to facilitate the determination of optimal conditions for fluorescence photography.Abbreviations: Fo = initial fluorescence; Fv = variable fluorescence; Fs = steady state fluorescence.