Cells of Anacystis nidulans strain R2 and of Synechococcus cedrorum were grown in an iron-deficient medium. Iron starvation induced several pronounced effects without influencing the viability of these cells. The phycocyanin and chlorophyll contents of these cells were depressed, and the absorption maxima of membrane-bound chlorophyll was blue-shifted by 5 nanometers. Cells showed a dramatic increase in original and in maximal chlorophyll fluorescence when monitored at room temperature. Low temperature chlorophyll fluorescence revealed a loss in fluorescence at 696 and 716 nanometers; much of the remaining fluorescence emission was at 686 nanometers. These chanes sugest an alteration of membrane composition and structure. This was documented by an electrophoretic analysis of iron-deficient membranes.%The prominent findings were: (a) large chlorophyll-protein complexes were not observed in iron-deficient membranes, although the chlorophyll-binding proteins were present; (b) the staining of acrylamide gels with 3,3',5,5'-tetramethylbenzidine plus peroxide indicated that iron deficiency led to a decrease in the quantity of cytochromes. These results support a stral model of the relation between fluorescence and chlorophyll organization in Anacystis. In addition, they suggest a method for studying cytochrome and chlorophyll protein assembly in these membranes. thesis of nuclear and chloroplastic gene products, and the subsequent assembly of these products into functional membranes (1 1). MATERIALS AND METHODSCells of Synechococcus cedrorum UTEX 1191 were obtained from the Indiana University culture collection, whereasAnacystis nidulans R2 was kindly provided by G. A. van Arkel, University of Utrecht, The Netherlands. These strains differed notably in such characteristics as cold sensitivity and efficiency of genetic transformation (23). However, no major differences were observed in the photosynthetic parameters described here or in the cellular responses to iron deficiency. Cells were routinely monitored for bacterial contamination by microscopic observation and by plating on nutrient agar.Cells were grown in shaking culture (120 rpm) in a manner described previously (7). Illumination was provided by banks of cool-white fluorescent lights at an intensity of approximately 0.4 mw/cm2 (YSI Radiometer). Optimal growth was obtained using Allen's BG-l 1 growth medium (1). Iron-deficient media was prepared by replacing the ferric ammonium citrate stock solution with equal molar amounts of ammonium citrate. Glassware Cells in buffer A were incubated with 10 mM EDTA and lysozyme (10 mg/ml; Sigma) for 2 to 3 h at 37°C. A membrane fraction was obtained by centrifugation after the osmotic rupture of these spheroplasts (8). The Chl-protein organization and the heme-dependent peroxidase profile of membranes were determined using LiDS-PAGE in a manner previously described (8). Polyacrylamide gradient gels were cast and were loaded with samples (80 ,g protein/well) which had been solubilized for 10 min with 1% LiDS (...

The rice bacterial blight pathogen Xanthomonas oryzae pv. oryzae is a vascular pathogen that elicits a defensive response through interaction with metabolically active rice cells. In leaves of 12-day-old rice seedlings, the exposed pit membrane separating the xylem lumen from the associated parenchyma cells allows contact with bacterial cells. During resistant responses, the xylem secondary walls thicken within 48 h and the pit diameter decreases, effectively reducing the area of pit membrane exposed for access by bacteria. In susceptible interactions and mock-inoculated controls, the xylem walls do not thicken within 48 h. Xylem secondary wall thickening is developmental and, in untreated 65-day-old rice plants, the size of the pit also is reduced. Activity and accumulation of a secreted cationic peroxidase, PO-C1, were previously shown to increase in xylem vessel walls and lumen. Peptide-specific antibodies and immunogold-labeling were used to demonstrate that PO-C1 is produced in the xylem parenchyma and secreted to the xylem lumen and walls. The timing of the accumulation is consistent with vessel secondary wall thickening. The PO-C1 gene is distinct but shares a high level of similarity with previously cloned pathogen-induced peroxidases in rice. PO-C1 gene expression was induced as early as 12 h during resistant interactions and peaked between 18 and 24 h after inoculation. Expression during susceptible interactions was lower than that observed in resistant interactions and was undetectable after infiltration with water, after mechanical wounding, or in mature leaves. These data are consistent with a role for vessel secondary wall thickening and peroxidase PO-C1 accumulation in the defense response in rice to X. oryzae pv. oryzae.

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Photosynthetic capacity decreases rapidly when temperate species are exposed to heat stress during reproductive development. We investigated whether injury in wheat (Triticum aestivum L.) resulted from general acceleration of senescence processes or specific heat-induced lesions. In situ photosynthetic capacity of leaf discs and thylakoid reactions were measured using flag leaf tissue from two cultivars maintained at 20 and 350C during maturation. Photosynthetic rates of leaf discs decreased faster at 35 than at 200C and were more photolabile in cv Len than in cv Waverly at high temperature. Patterns of thylakoid breakdown also differed in the two wheat genotypes at 20°C: intersystem electron transport and photosystem 11 activity decreased linearly during postanthesis development in Len wheat, whereas coupling of photophosphorylation to electron transport declined late during senescence in Waverly wheat. Heat stress induced early loss of intersystem electron transport followed sequentially by decreased silicomolybdic acid, + 3-(3,4-dichlorophenyl)-1-dimethylurea-mediated photosystem 11 activity and 2,5-dichloro-p-benzoquinone-mediated photosystem 11 activity in Len. Stress accelerated the uncoupling process, but loss of intersystem electron transport and photosystem 11 activities was slower in Waverly than in Len

Ubiquitin, a key component in an ATP-dependent proteolytic pathway, participates in the response of various eucaryotic organisms to high temperature stress. Our objective was to determine if ubiquitin serves a similar capacity for metabolizing altered proteins in higher plants during stress. Degradation of total proteins was measured, and ubiquitin pools (free versus conjugated) were extracted with an improved protocol from wheat (Triticum aestivum L. cv Len) roots treated at 22, 27, 32, 37, and 42 degrees C for 1 hour and assayed by western blots and radioimmunoassays. Heat-shock protein synthesis was detected by in vivo labeling and autoradiography. Mean half-life of total root proteins decreased from 51 hours at 22 degrees C to 23 hours at 40 degrees C. Ubiquitin pools were extracted better and proteolysis was slowed more by the improved protocol than by a conventional procedure for plant proteins. Amounts of high molecular mass conjugates were elevated and levels of low molecular mass conjugates and free ubiquitin were depressed when roots were treated at 37 or 42 degrees C than at lower temperatures; the same high temperatures also induced synthesis of heat-shock proteins. We concluded that high temperatures increase breakdown of root proteins, which are degraded via the ubiquitin proteolytic pathway. A conjugate with an apparent molecular mass of 23 kilodaltons was tentatively identified as an ubiquitinated histone.