Characterization of a Sorghum bicolor gene family encoding putative protein kinases with a high similarity to the yeast SNF1 protein kinase

C₄ photosynthesis is functionally dependent on metabolic interactions between mesophyll and bundle-sheath cells. Although the C₄ cycle is biochemically well understood many aspects of the regulation of enzyme activities, gene expression and cell differentiation are elusive.Protein kinases are likely involved in these regulatory processes providing links to hormonal, metabolic and developmental signal transduction pathways. We have identified several protein kinases that are differentially expressed in mesophyll and bundle-sheath cells of the C₄ plant Sorghum bicolor. Here we describe the characterization of two putative protein kinases that show high similarity to the SNF1/AMPK family of protein serine/threonine kinases. The mRNA of both kinases accumulates to much higher levels in mesophyll cells than in the bundle-sheath and can also be detected in root tissue. Complementation experiments with a snf1 mutant of Saccharomyces cerevisiae indicate that the S. bicolor protein kinase SNFL1 does not represent a functional homologue of the yeast SNF1 protein kinase.     

SbRLK1, a receptor-like protein kinase of Sorghum bicolor (L.) Moench that is expressed in mesophyll cells

To study the metabolic interactions between mesophyll and bundle-sheath cells of C₄ plants, protein kinases possibly involved in the regulatory processes and signal transduction pathways have been cloned and characterized. A receptor-like protein kinase (RLK) cDNA clone from the C₄ plant Sorghum bicolor (L.) Moench has been identified. The deduced protein was designated SbRLK1 for receptor-like protein kinase from S. bicolor. The putative cytoplasmic domain of SbRLK1 contains all amino acids that are characteristic of protein kinases. The extracellular domain contains five leucine-rich repeats. The mRNA of the SbRLK1 gene accumulated to much higher levels in mesophyll cells than in the bundle-sheath and was almost undetectable in roots. This expression pattern indicates that SbRLK1 might be involved in the regulation of specific processes in mesophyll cells. Received: 13 August 1998 / Accepted: 22 December 1998     

Antioxidants and antioxidative enzymes in wild-type and transgenic Lycopersicon genotypes of different chilling tolerance

The Mehler–Ascorbate–Peroxidase cycle is a protection system against reactive oxygen species (ROS) occurring during over-excitation of the photosynthetic apparatus. In the cultivated tomato, Lycopersicon esculentum, long-term chilling under moderate light leads to oxidation of the Calvin cycle key enzyme, ribulose-1,5-bisphosphate carboxylase (rubisco), presumably by generation of ROS. In contrast, high-altitude lines of the wild tomato species L. peruvianum were tolerant against the same chilling stress. In the present study, we analysed leaf contents of antioxidants (ascorbate, glutathione) and activities of enzymes of the Mehler–Ascorbate–Peroxidase cycle in the two Lycopersicon species. While antioxidant levels and activities of chloroplast superoxide dismutase (SOD) and ascorbate peroxidase (APX), both inducible by chilling stress, were similar in chilling-tolerant and chilling-sensitive genotypes, chilled L. esculentum showed lower glutathione reductase (GR) activities than high-altitude L. peruvianum. We constructed transgenic plants overexpressing an Escherichia coli GR in the chloroplast (approximately 60-fold of the wild-type (WT) activity). However, these plants resembled identical chilling sensitivity of the photosynthetic apparatus as WT plants as measured after a photoinhibition treatment and by the effect of long-term chilling on rubisco activity. We conclude that the Mehler–Ascorbate–Peroxidase cycle is not the limiting factor for the sensitivity of the photosynthetic apparatus of L. esculentum towards long-term chilling under moderate light. We suggest that a possible cause for the higher chilling tolerance of L. peruvianum is prevention of ROS formation by better conversion of light energy to photochemistry at suboptimal temperatures.     

Spectrophotometric and Molecular Properties of Mutated Rice Phytochrome A

A cDNA (PHYA) for the phytochrome A apoprotein (PHYA) of riceand three mutated sequences (phyA S/A, the first ten serineresidues in the N-terminal domain of PHYA were changed to alanineresidues; phyA ND, the first 80 N-terminal amino acids weredeleted; phyA CD, the amino acids of the C-terminal domain from689 to 1,128 were deleted) were expressed in yeast, and thewild-type and mutant apophytochromes were allowed to combinein vitro with the chromophore phycocyanobilin (PCB). The PCB-attachedproduct of phyA S/A gave very similar spectrophotometric peaksto the PhA_fr and PhyA_fr forms of wild-type product. By contrast,the peak of the product of phyA CD in the P_fr form was significantlyshifted towards a shorter wavelength, an indication that, whereasthe C-terminal domain is not crucial for the PCB attachment,it greatly influences the absorption maximum of PhyA_fr. Therate of 50% reversion from PhyA_fr to PhA_r in darkness was 3h at 27°C with all of the samples, showing that the S/Aand CD mutations did not affect this property. No photoreversibilitywas detected with the product of phyA ND. Gel-filtration analysisof the wild-type PHYA and the product of phyA S/A showed thatthe apparent molecular mass of each was 330 kDa, suggestingthat both exists as dimers in solution. ⁴Present address: Institut für Entwicklungs- und Molekularbiologieder Pflanzen, Heinrich-Heine-Universität-Düsseldorf,Universitätsstr. 1, 40225 Düsseldorf, Germany     

Characterization of 14 different putative protein kinase cDNA clones of the C4 plantSorghum bicolor

C₄ photosynthesis is functionally dependent on metabolic interactions between mesophyll- and bundle-sheath cells. Although the C₄ cycle is biochemically well understood, many aspects of the regulation of enzyme activities, gene expression and cell differentiation are elusive. Protein kinases are likely involved in these regulatory processes, providing links to hormonal, metabolic and developmental signal-transduction pathways. Here we describe the cloning and characterization of 14 different putative protein kinase leaf cDNA clones from the C₄ plant Sorghum bicolor. These genes belong to three different protein kinase subfamilies: ribosomal protein S6 kinases, SNF1-like protein kinases, and receptor-like protein kinases. We report the partial cDNA sequences, mesophyll/bundle-sheath steady-state mRNA ratios, mesophyll/etiolated leaf steady-state mRNA ratios, and the positions of 14 protein kinase genes on the genetic map of S. bicolor. Only three of the protein kinase genes described here are expressed preferentially in mesophyll cells as compared with the bundle-sheath.     

Characterization of 14 different putative protein kinase cDNA clones of the C4 plantSorghum bicolor

C₄ photosynthesis is functionally dependent on metabolic interactions between mesophyll- and bundle-sheath cells. Although the C₄ cycle is biochemically well understood, many aspects of the regulation of enzyme activities, gene expression and cell differentiation are elusive. Protein kinases are likely involved in these regulatory processes, providing links to hormonal, metabolic and developmental signal-transduction pathways. Here we describe the cloning and characterization of 14 different putative protein kinase leaf cDNA clones from the C₄ plant Sorghum bicolor. These genes belong to three different protein kinase subfamilies: ribosomal protein S6 kinases, SNF1-like protein kinases, and receptor-like protein kinases. We report the partial cDNA sequences, mesophyll/bundle-sheath steady-state mRNA ratios, mesophyll/etiolated leaf steady-state mRNA ratios, and the positions of 14 protein kinase genes on the genetic map of S. bicolor. Only three of the protein kinase genes described here are expressed preferentially in mesophyll cells as compared with the bundle-sheath. Received: 16 January 1998 / Accepted: 3 April 1998     

Ectopic expression of a single homeotic gene, the Petunia gene green petal, is sufficient to convert sepals to petaloid organs.

Genetic studies in Arabidopsis and Antirrhinum showed that petal determination requires the concomitant expression of two homeotic functions, A and B, whereas the A function alone determines sepal identity. The B function is represented by at least two genes. The Petunia homeotic gene green petal (gp) is essential for petal determination as demonstrated by a Petunia gp mutant that has sepals instead of petals. We have used ectopic expression of the gp gene as a tool to study flower development in Petunia. CaMV 35S-gp expression leads to homeotic conversion of sepals into petaloid organs when expressed early in development. This demonstrates that a single homeotic gene is sufficient to induce homeotic conversion of sepals to petals, suggesting that other petal determining genes are regulated in part by ectopically expressed gp. Indeed, two other MADS-box-containing genes, pmads 2 and fbp 1, which show homology to the Antirrhinum B function gene globosa, are activated in the converted petal tissue. Furthermore, our data provide evidence for autoregulation of gp expression in the petaloid tissue and uncover the role of gp in fusion of petal tissues.