Biomedical subjects

Fritz Kreuzaler

Publications and source records attributed to Fritz Kreuzaler.

6 recordsLinked to original sources

PubMed2006

A drastic reduction in DOF1 transcript levels does not affect C4-specific gene expression in maize.

The transcription factor DOF1 has been suggested to regulate photosynthetic gene expression in maize. By screening a RescueMu transposon-tagged mutant library, we identified a maize mutant with a transposon integration in the Dof1 gene 16 bp upstream of the transcription initiation site (TIS). Sequencing of the Dof1 promoter region revealed an unusual promoter structure missing any typical elements. Homozygous (ho) mutant lines were generated by selfing and subsequent PCR and DNA gel blot analyses. The transposon integration reduced Dof1 transcript levels to less than 20% compared to the wild-type and overlapping RT-PCR systems revealed that these transcripts were not initiated from the native transcription start site. Dof1 transcripts transiently accumulate in wild-type plants after illumination of darkened seedlings, but this accumulation cannot be observed in mutant lines. However, the time-course of transcript accumulation from the C(4)-specific phosphoenolpyruvate carboxylase (PEPC) gene, a possible target of DOF1, is not altered. Moreover, no impact on the steady-state levels of five additional transcripts involved in C(4)-metabolism can be observed. The contents of amino acids, glucose, and malate as well as the carbon to nitrogen ratio in the leaves remained unchanged when comparing wild-type and mutant plants. Our data question the importance of DOF1 in the control of photosynthetic gene expression in maize.

Carbon Dioxide↗

PubMed2004

Fusion proteins comprising a Fusarium-specific antibody linked to antifungal peptides protect plants against a fungal pathogen.

In planta expression of recombinant antibodies recognizing pathogen-specific antigens has been proposed as a strategy for crop protection. We report the expression of fusion proteins comprising a Fusarium-specific recombinant antibody linked to one of three antifungal peptides (AFPs) as a method for protecting plants against fungal diseases. A chicken-derived single-chain antibody specific to antigens displayed on the Fusarium cell surface was isolated from a pooled immunocompetent phage display library. This recombinant antibody inhibited fungal growth in vitro when fused to any of the three AFPs. Expression of the fusion proteins in transgenic Arabidopsis thaliana plants conferred high levels of protection against Fusarium oxysporum f.sp. matthiolae, whereas plants expressing either the fungus-specific antibody or AFPs alone exhibited only moderate resistance. Our results demonstrate that antibody fusion proteins may be used as effective and versatile tools for the protection of crop plants against fungal infection.

Agrobacterium tumefaciens↗

PubMed2003

Quantification of photosynthetic gene expression in maize C(3) and C(4) tissues by real-time PCR.

Carbon assimilation in maize follows the C(4) mechanism. This requires the tissue-specific and light-induced expression of a set of different genes involved in CO(2) fixation as well as adaptations in the leaf anatomy including a reduced distance between vascular bundles compared to C(3) plants. However, several maize tissues exist with larger bundle distances and there is significant evidence that CO(2) fixation follows the C(3) mechanism in these tissues. We isolated maize C(3) and C(4) tissues and quantified the accumulation of mRNAs encoding PEPC, ME, the small subunit of Rubisco, and PPDK. For this, primer systems for the specific and sensitive detection by real-time PCR were established. The observed patterns show the expected distribution for foliar leaf tissues. Also in total husk leaves, all transcripts under investigation were detected, albeit at a lower level. When mesophyll cells which are located distant from bundles were isolated from husk leaves, only accumulation of RbcS was observed. Comparing the expression of two genes encoding for isoenzymes of the small subunit of RbcS in the different tissues differential patterns of relative transcript abundance were observed. Transcripts for the DOF1 transcription factor involved in the activation of photosynthetic genes in maize were found in leaf tissues performing both C(4) and C(3) photosynthesis with highest accumulation levels in C(4) mesophyll cells, whereas the homologous DOF2 gene was not expressed in any of the investigated samples. The results provide novel insights into the regulation of C(3) and C(4) carbon fixation pathways in maize.

Journal Article↗

PubMed2002

An engineered phosphoenolpyruvate carboxylase redirects carbon and nitrogen flow in transgenic potato plants.

Phosphoenolpyruvate carboxylase (PEPC) plays a central role in the anaplerotic provision of carbon skeletons for amino acid biosynthesis in leaves of C3 plants. Furthermore, in both C4 and CAM plants photosynthetic isoforms are pivotal for the fixation of atmospheric CO2. Potato PEPC was mutated either by modifications of the N-terminal phosphorylation site or by an exchange of an internal cDNA segment for the homologous sequence of PEPC from the C4 plant Flaveria trinervia. Both modifications resulted in enzymes with lowered sensitivity to malate inhibition and an increased affinity for PEP. These effects were enhanced by a combination of both mutated sequences and pulse labelling with 14CO2 in vivo revealed clearly increased fixation into malate for this genotype. Activity levels correlated well with protein levels of the mutated PEPC. Constitutive overexpression of PEPC carrying both N-terminal and internal modifications strongly diminished plant growth and tuber yield. Metabolite analysis showed that carbon flow was re-directed from soluble sugars and starch to organic acids (malate) and amino acids, which increased four-fold compared with the wild type. The effects on leaf metabolism indicate that the engineered enzyme provides an optimised starting point for the installation of a C4-like photosynthetic pathway in C3 plants.

Biological Transport↗

PubMed2002

Overexpression of C(4)-cycle enzymes in transgenic C(3) plants: a biotechnological approach to improve C(3)-photosynthesis.

The process of photorespiration diminishes the efficiency of CO(2) assimilation and yield of C(3)-crops such as wheat, rice, soybean or potato, which are important for feeding the growing world population. Photorespiration starts with the competitive inhibition of CO(2) fixation by O(2) at the active site of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and can result in a loss of up to 50% of the CO(2) fixed in ambient air. By contrast, C(4) plants, such as maize, sugar cane and Sorghum, possess a CO(2) concentrating mechanism, by which atmospheric CO(2) is bound to C(4)-carbon compounds and shuttled from the mesophyll cells where the prefixation of bicarbonate occurs via phosphoenolpyruvate carboxylase (PEPC) into the gas-tight bundle-sheath cells, where the bound carbon is released again as CO(2) and enters the Calvin cycle. However, the anatomical division into mesophyll and bundle-sheaths cells ("Kranz"-anatomy) appears not to be a prerequisite for the operation of a CO(2) concentrating mechanism. Submerged aquatic macrophytes, for instance, can induce a C(4)-like CO(2) concentrating mechanism in only one cell type when CO(2) becomes limiting. A single cell C(4)-mechanism has also been reported recently for a terrestrial chenopod. For over 10 years researchers in laboratories around the world have attempted to improve photosynthesis and crop yield by introducing a single cell C(4)-cycle in C(3) plants by a transgenic approach. In the meantime, there has been substantial progress in overexpressing the key enzymes of the C(4) cycle in rice, potato, and tobacco. In this review there will be a focus on biochemical and physiological consequences of the overexpression of C(4)-cycle genes in C(3) plants. Bearing in mind that C(4)-cycle enzymes are also present in C(3) plants, the pitfalls encountered when C(3) metabolism is perturbed by the overexpression of individual C(4) genes will also be discussed.

Carbon Dioxide↗

PubMed1986

Light-induced expression of the chimeric chalcone synthase-NPTII gene in tobacco cells.

A chimeric gene was constructed containing the light-inducible chalcone synthase (chs) promoter from Antirrhinum majus, the neomycin phosphotransferase structural sequence from Tn5 as a reporter gene (NPTII) and the termination region from chs gene 1 from Petroselinum hortense. This gene was introduced into tobacco plants with the help of Ti plasmid-derived plant vectors and NPTII expression was measured. Analysis of the chs promoter sequence indicated the position of several possible regulatory regions. These were deleted to test their influence on the expression of the chs-NPTII gene. The different chimeric genes were all shown to be active after transfer to tobacco with the exception of one, in which the entire 5' upstream sequence from -1200 to -39 was deleted. The transcription of a chimeric gene with a 1.2-kbp 5' upstream promoter sequence was shown to be light inducible in tobacco plants. The analysis of various deletions of this 5' upstream sequence indicates that a number of sequence motifs have a quantitative effect on gene expression. One of these sequence motifs (-564 to -661) contains a direct repeat of 47 bp and the sequence GTGGTTAG which corresponds to the consensus core sequences observed in animal gene enhancer sequences. Deletion of a fragment containing this direct repeat resulted in a reduction of NPTII expression by a factor of 5.

Journal Article↗