Biomedical subjects

J Lerchl

Publications and source records attributed to J Lerchl.

4 recordsLinked to original sources

PubMed2000

Cloning and functional expression of the first plant fatty acid elongase specific for Delta(6)-polyunsaturated fatty acids.

In order to elucidate the biosynthesis of long-chain polyunsaturated fatty acids (PUFAs) in plants we searched for a cDNA encoding a Delta(6)-specific PUFA elongase from Physcomitrella patens, which is known to contain high proportions of arachidonic acid (20:4 Delta(5,8,11,14)). An EST clone from P. patens was identified by its low homology to the yeast gene ELO1, which is required for the elongation of medium-chain fatty acids. We functionally characterized this cDNA by heterologous expression in Saccharomyces cerevisiae grown in the presence of several fatty acids. Analysis of the fatty acid profile of the transgenic yeast revealed that the cDNA encodes a protein that leads to the elongation of the C(18) Delta(6)-polyunsaturated fatty acids gamma-linolenic acid (18:3 Delta(6,9,12)) and stearidonic acid (18:4 Delta(6,9,12,15)), which were recovered to 45-51% as their elongation products. In contrast, linoleic and alpha-linolenic acids were hardly elongated and we could not measure any elongation of saturated and mono-unsaturated fatty acids (including 18:1 Delta(6)), indicating that the elongase is highly specific for the polyunsaturated nature of the fatty acid acting as substrate.

Acetyltransferases↗

PubMed1999

Spinach hexokinase I is located in the outer envelope membrane of plastids.

The subcellular localization of hexokinase activities in plant cells has been a matter of debate for a long time. We have isolated a hexokinase cDNA fragment from glucose-fed spinach leaves using a differential display reverse transcription-PCR approach. The corresponding cDNA was expressed in Escherichia coli and an antiserum, raised against the recombinant protein, was used in subcellular localization studies. The spinach hexokinase could be localized primarily to the outer envelope membrane of chloroplasts where it is inserted via its N-terminal membrane anchor. We suggest that the chloroplast envelope hexokinase is involved in the energization of glucose export from plastids rather than in the sugar-sensing pathway of the plant cell.

Base Sequence↗

PubMed1995

Molecular cloning, characterization and expression analysis of isoforms encoding tonoplast-bound proton-translocating inorganic pyrophosphatase in tobacco.

The tonoplast-bound proton-translocating inorganic pyrophosphatase (V-type PPase) and the proton ATPase (V-type ATPase) are electrogenic proton pumps guaranteeing the energization of solute transport across the tonoplast. Using an Arabidopsis thaliana PCR cDNA fragment corresponding to clone ATAVP3 we have isolated 24 cDNA clones encoding tonoplast-bound inorganic pyrophosphatase of tobacco. Based on restriction analysis the cDNA clones could be grouped into three different classes. The complete nucleotide sequence of one member of each class (TVP5, TVP9 and TVP31) was determined. The cDNA clones contain an uninterrupted open reading frame of 2292 bp (TVP5), 2295 bp (TVP9) and 2298 bp (TVP31) coding for polypeptides of 764, 765 and 766 amino acids, respectively. The nucleotide sequence of the different clones is highly homologous within the coding region (79-89% identity) but differs strongly in the untranslated regions. The individual classes are encoded by single- or low-copy genes as judged from genomic gel blot experiments using 3'-specific probes. RNA analysis revealed that the accumulation of the specific transcripts is differentially regulated during leaf development.

Amino Acid Sequence↗

PubMed1995

Impaired photoassimilate partitioning caused by phloem-specific removal of pyrophosphate can be complemented by a phloem-specific cytosolic yeast-derived invertase in transgenic plants.

Constitutive expression of the Escherichia coli ppa gene encoding inorganic pyrophosphatase resulted in sugar accumulation in source leaves and stunted growth of transgenic tobacco plants. The reason for this phenotype was hypothesized to be reduced sucrose utilization and loading into the phloem. To study the role of PPi in phloem cells, a chimeric gene was constructed using the phloem-specific rolC promoter of Agrobacterium rhizogenes to drive the expression of the ppa gene. Removal of cytosolic PPi in those cells resulted in photoassimilate accumulation in source leaves, chlorophyll loss, and reduced plant growth. From these data, it was postulated that sucrose hydrolysis via sucrose synthase is essential for assimilate partitioning. To bypass the PPi-dependent sucrose synthase step, transgenic plants were produced that express various levels of the yeast suc2 gene, which encodes cytosolic invertase, in their phloem cells. To combine the phloem-specific expression of the ppa gene and the suc2 gene, crosses between invertase- and pyrophosphatase-containing transgenic plants were performed. Analysis of their offspring revealed that invertase can complement the phenotypic effects caused by the removal of PPi in phloem cells.

Base Sequence↗