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Biomedical subjects

W Schuster

Publications and source records attributed to W Schuster.

At least 19 recordsLinked to original sources
PubMed2006

Electromyographic patterns in children with cerebral palsy: do they change after surgery?

The purpose of this study was to investigate the changes in electromyographic (EMG) patterns after multilevel surgical treatment in children with spastic cerebral palsy. Children with diplegia (n=18) and hemiplegia (n=16) aging from 6 to 16 years participated in the study. Twenty healthy children within the same age span are presented as reference. Gait analysis and surface electromyograms of seven major lower limb muscles were assessed before and 1-5 years after the multilevel surgery. The most frequent procedures were equinus correction, distal rectus femoris transfer, femoral derotation osteotomy and hamstrings lengthening. The results showed that the EMG pattern of the soleus, lateral gastrocnemius and tibialis anterior muscles became closer to normal after the surgery, while no differences were detected between diplegic and hemiplegic patients. Furthermore, a subgroup of 10 patients showed an increase in medial hamstrings activation during preswing that decreased postoperatively. These findings indicate that changes in EMG patterns should not be ruled out after surgical treatment, although the extent of these changes is limited compared to changes in the kinematics. Abnormal muscle activation before the operation can be related to a compensatory response in some patients and this can be manipulated after surgery.

Adolescent↗
PubMed2001

Plant growth in elevated CO2 alters mitochondrial number and chloroplast fine structure.

With increasing interest in the effects of elevated atmospheric CO(2) on plant growth and the global carbon balance, there is a need for greater understanding of how plants respond to variations in atmospheric partial pressure of CO(2). Our research shows that elevated CO(2) produces significant fine structural changes in major cellular organelles that appear to be an important component of the metabolic responses of plants to this global change. Nine species (representing seven plant families) in several experimental facilities with different CO(2)-dosing technologies were examined. Growth in elevated CO(2) increased numbers of mitochondria per unit cell area by 1.3-2.4 times the number in control plants grown in lower CO(2) and produced a statistically significant increase in the amount of chloroplast stroma (nonappressed) thylakoid membranes compared with those in lower CO(2) treatments. There was no observable change in size of the mitochondria. However, in contrast to the CO(2) effect on mitochondrial number, elevated CO(2) promoted a decrease in the rate of mass-based dark respiration. These changes may reflect a major shift in plant metabolism and energy balance that may help to explain enhanced plant productivity in response to elevated atmospheric CO(2) concentrations.

Carbon Dioxide↗
PubMed1998

Assays for investigating RNA editing in plant mitochondria.

In plant organelles transcripts are modified posttranscriptionally by RNA editing. This modification process changes almost every protein-coding RNA at specific cytidine and uridine positions. Therefore, mitochondrially encoded protein sequences differ from the genomically fixed information and show, after editing, a higher conservation. To investigate this unusual processing step in plant mitochondria, several assays have been developed. However, compared with the progress made in other RNA editing fields, knowledge about the factors involved in plant mitochondrial editing is limited. One reason for this is the lack of a reliable in vitro system for mitochondria. To reveal the biochemical nature of the RNA editing reaction in plant mitochondria, we developed an in vitro system by which we were able to show that cytidine is specifically modified to uridine by a deamination or transamination process. Here we describe the development of a pea in vitro system and discuss assays to follow the editing process.

Genetic Techniques↗
PubMed1996

Transfer of rps19 to the nucleus involves the gain of an RNP-binding motif which may functionally replace RPS13 in Arabidopsis mitochondria.

The discovery of disrupted rps19 genes in Arabidopsis mitochondria prompted speculation about the transfer to the nuclear compartment. We here describe the functional gene transfer of rps19 into the nucleus of Arabidopsis. Molecular cloning and sequence analysis of rps19 show that the nuclear gene encodes a long N-terminal extension. Import studies of the precursor protein indicate that only a small part of this extension is cleaved off during import. The larger part of the extension, which shows high similarity to conserved RNA-binding domains of the RNP-CS type, became part of the S19 protein. In the Escherichia coli ribosome S19 forms an RNA-binding complex as heterodimer with S13. By using immuno-analysis and import studies we show that a eubacterial-like S13 protein is absent from Arabidopsis mitochondria, and is not substituted by either a chloroplastic or a cytosolic homologue of this ribosomal protein. We therefore propose that either a highly diverged or missing RPS13 has been functionally replaced by an RNP domain that most likely derived from a glycine-rich RNA-binding protein. These results represent the first case of a functional replacement of a ribosomal protein by a common RNA-binding domain and offer a new view on the flexibility of biological systems in using well-adapted functional domains for different jobs.

Amino Acid Sequence↗
PubMed1996

Loss of RNA editing of rps1 sequences in Oenothera mitochondria.

We have analysed a region downstream from the atp9 gene in Oenothera mitochondrial DNA which contains an open reading frame of 224 codons. This open reading frame, designated orf224, is co-transcribed with the atp9 gene. In wheat mitochondria, a homologous reading frame (orf174) has been described which is considerably shorter at the N-terminus compared to the putative Oenothera gene. The deduced polypeptides from both species show high similarity to the N-terminal third of ribosomal protein S1 of bacteria. Transcripts of orf174 are edited in wheat mitochondria whereas the similarly conserved cytidine positions in orf224 mRNAs of Oenothera are silent. On the other hand, atp9 sequences, which are located upstream on the same co-transcript, are fully edited. Because of this, we conclude that editing sites are selected independently for mRNA sections. Our results suggest that orf224 represents a transcribed pseudogene in Oenothera. The active gene for a functional ribosomal protein S1 for mitochondria is therefore expected to be present in the nucleus. A nuclear localization for this gene is likewise suggested for Arabidopsis due to the fact that rps1 sequences are absent from the mitochondrial genome.

Amino Acid Sequence↗
PubMed1995

Transfer of rps10 from the mitochondrion to the nucleus in Arabidopsis thaliana: evidence for RNA-mediated transfer and exon shuffling at the integration site.

Rps10, a gene coding for ribosomal protein S10 of Arabidopsis mitochondria has been transferred to the nuclear compartment, while in pea and potato the active rps10 is mitochondrially located. The nuclear rps10 gene contains an intron at the junction of the target signal sequence and the mitochondrial-derived sequence, indicating that exon shuffling may have been involved in the addition of the transit peptide signal. Sequence comparison of Arabidopsis rps10 to the plant mitochondrial counterparts shows that the edited version is present in the nucleus of Arabidopsis. This finding corroborates RNA as an intermediate of a functional gene transfer between mitochondria and the nucleus. In vitro-translated RPS10 protein is efficiently imported into potato mitochondria and a presequence of about 7 kDa is removed resulting in a mature protein that is larger compared to organellar and bacterial RPS10 proteins.

Amino Acid Sequence↗
PubMed1995

Evidence for a site-specific cytidine deamination reaction involved in C to U RNA editing of plant mitochondria.

Transcripts of higher plant mitochondria are modified post-transcriptionally by RNA editing. To distinguish between the mechanisms by which the cytidine to uridine transition could occur a combined transcription/RNA editing assay and an in vitro RNA editing system were investigated. Mitochondria isolated from etiolated pea seedlings and potato tubers were supplied with [alpha-32P]CTP to radiolabel the mitochondrial run-on transcripts. High molecular weight run-on transcripts were isolated and hydrolyzed, and nucleotide identities were analyzed by one- and two-dimensional thin layer chromatography. The amount of label comigrating with UMP nucleotides increases with extended incubation times. Analogous products were obtained by incubation of [alpha-32P]CTP or [5-3H]CTP radiolabeled in vitro transcripts with a mitochondrial lysate from pea mitochondria. 5-3H label of the cytosine base was detected in the UMP spot after incubation of in vitro transcripts with mitochondrial lysate. These results are consistent with a deamination reaction involved in this post-transcriptional C to U modification process. To prove that cytidines are deaminated specifically in vitro transcripts were reisolated after incubation and analyzed by reverse transcription-polymerase chain reaction. Sequence analysis clearly shows that only cytidines at editing sites are edited while residual cytidines are not modified and suggests that site-specific factors are involved in RNA editing of plant mitochondria.

Base Sequence↗
PubMed1995

Oenothera mitochondrial orf454, a gene involved in cytochrome c biogenesis corresponds to orf169 and orf322 of Marchantia.

We have characterized a mitochondrial gene in Oenothera, designated orf454, capable of encoding a component of the cytochrome c biogenesis system. This open reading frame is interrupted by an intron of 941 nucleotides showing high similarity to a group II intron residing in the rpl2 gene. RNA editing, which is observed at 18 cytidine positions within the orf454 reading frame, improves the similarity to protein-coding sequences in bacteria and higher plants and removes the last 16 amino acids. orf454 also shows high sequence similarity to two overlapping reading frames (orf169 and orf322) of Marchantia mitochondria. These ORFs belong to an operon-like cluster of genes in the liverwort that is not conserved in Oenothera mitochondria. However, in bacteria these reading frames are organized like the Marchantia gene cluster. It has been shown by genetical analysis in Rhodobacter capsulatus that these genes are essential for cytochrome c biogenesis. Genes of bacterial operons-ccl1 in Rhodobacter and yejR and nrfE in Escherichia coli - show high sequence similarity to the mitochondrial reading frames orf577 and orf454 of Oenothera. orf454, which we describe here, is homologous to the C-terminal region of these bacterial genes, while the previously described orf577 is homologous to the N-terminal region.

Amino Acid Sequence↗
PubMed1995

orf250 encodes a second subunit of an ABC-type heme transporter in Oenothera mitochondria.

A highly transcribed region in Oenothera mitochondria codes for an open reading frame comprising 250 condons (orf250). This open reading frame shows high sequence similarity to the helC gene of Rhodobacter capsulatus which encodes a subunit of a proposed ABC-type heme transporter. Transcripts of orf250 are edited by cytidine to uridine transitions at 29 sites, altering 10% of all encoded amino acids. Genes homologous to helC have also been found in the bacteria Bradyrhizobium japonicum and Escherichia coli, and are conserved in mitochondria of Marchantia polymorpha, Daucus carota, and Arabidopsis thaliana. In bacteria these genes belong to operons that are involved in the biogenesis of c-type cytochromes. The bacterial gene organization is partly conserved in Marchantia, but altered in the mitochondrial genome of Oenothera.

ATP-Binding Cassette Transporters↗
PubMed1995

RNA editing in higher plant mitochondria: analysis of biochemistry and specificity.

RNA editing alters genomically encoded cytidines to uridines posttranscriptionally in higher plant mitochondria. Most of these editing events occur in translated regions and consequently alter the amino acid sequence. In Oenothera berteriana more than 500 editing sites have been detected and the total number of editing sites exceeds 1000 sites in this mitochondrial genome. To identify the components involved in this process we investigated the factors determining the specificity of RNA editing and the apparent conversion of cytidine to uridine residues. The possible biochemical reactions responsible for RNA editing in plant mitochondria are de- or transamination, base substitution and nucleotide replacement. In order to discriminate between these different biochemical mechanisms we followed the fate of the sugar-phosphate backbone by analysing radiolabeled nucleotides after incorporation into high molecular mass RNA. Plant mitochondria were supplied with [alpha-32P]CTP to radiolabel CMP residues in newly synthesized transcripts. Radiolabeled mtRNA was extracted and digested with nuclease P1 to hydrolyse the RNA to monophosphates. The resulting monophosphates were analysed on one- and two-dimensional TLC systems to separate pC from pU. Radiolabeled pU was detected in increasing quantities during the course of incubation. These results suggest that RNA editing in plant mitochondria involves either a deamination or a transglycosylation reaction. The editing product was identified as uridine and not as a hypermodified nucleotide which is recognized as uridine. Similar results have been obtained by incubating in vitro transcribed mRNAs with mitochondrial lysates indicating that RNA editing and transcription is not directly linked in plant mitochondria.(ABSTRACT TRUNCATED AT 250 WORDS)

Biological Evolution↗
PubMed1995

Potato mitochondrial manganese superoxide dismutase is an RNA-binding protein.

An RNA-binding protein present in potato mitochondrial lysates was purified and identified as manganese-containing superoxide dismutase (MnSOD). Using a gel mobility shift assay we found that proteins from mitochondrial lysates bind with high affinity to in vitro transcripts of mitochondrial orf206, encoding a subunit of the ABC-type heme transporter. By ammonium sulfate fractionation and two subsequent chromatographic steps on MonoQ columns we purified a 28 kDa protein to apparent homogeneity. Protein sequencing identified the purified polypeptide as manganese-containing superoxide dismutase, which is a specific enzymatic scavenger of superoxides in mitochondria. Using gel mobility shift and competition assays, we show that RNA-binding of MnSOD of potato is not influenced by 400 mM KCl or heparin and is specific to heteropolymeric RNAs. The labeled mitochondrial transcript could be competed with low amounts of unlabeled transcript while binding was stable to competition with large amounts of tRNA or high concentrations of NADH and NADPH. The purified MnSOD of potato mitochondria was UV-cross-linked to the mitochondrial transcript. The Mn- and Fe-containing SODs from Escherichia coli showed no binding to the RNA by either gel mobility shift or UV-cross-linking. Enzyme activity assays revealed that binding of RNA to the mitochondrial MnSOD does not significantly influence enzyme activity. This indicates that the RNA-binding feature of MnSOD of potato mitochondria is probably not involved in modulating SOD enzyme activity and suggests a function different from superoxide degradation as ist biological role.

Amino Acid Sequence↗
PubMed1994

The highly edited orf206 in Oenothera mitochondria may encode a component of a heme transporter involved in cytochrome c biogenesis.

A highly transcribed region in Oenothera mitochondria codes for a reading frame (orf206) which shows high homology to the Marchantia encoded mitochondrial open reading frame orf277 and is also conserved in the mitochondrial genomes of Arabidopsis thaliana and Daucus carota. Transcripts of orf206 are modified by cytidine to uridine changes in 46 positions by RNA editing, affecting 30% of all cytidines and 15% of the total encoded amino acids. This ORF is cotranscribed with an upstream reading frame and with the downstream rps 14 gene. The orf206 deduced protein shows high similarity to polypeptides which are proposed to be part of an ABC-type heme transporter involved in cytochrome c biogenesis in Bradyrhizobium and Rhodobacter.

Amino Acid Sequence↗
PubMed1994

Rps3 and rpl16 genes do not overlap in Oenothera mitochondria: GTG as a potential translation initiation codon in plant mitochondria?

Characterization of the Oenothera mitochondrial ribosomal gene cluster rps19-rps3-rpl16 shows the two genes rps3 and rpl16 to be separated by 9 nucleotides. The first codon of rpl16 is a GTG codon for valine and the only potential translational start. This GTG codon is conserved at the same position in maize, Petunia and Marchantia mitochondria, while sequences diverge upstream. These observations suggest that GTG at least at this position may act as translation initiation codon in plant mitochondria. Analysis of RNA editing suggests both genes to code for functional ribosomal proteins in Oenothera mitochondria. A duplication/recombination event at a decanucleotide in the intron of rps3 created a pseudogene missing part of the intron and the 3' exon.

Amino Acid Sequence↗
PubMed1994

Physical mapping of the mitochondrial genome of Arabidopsis thaliana by cosmid and YAC clones.

As part of the worldwide efforts at molecular analysis of Arabidopsis thaliana as a model plant the complete structure of the mitochondrial genome has been determined. The mitochondrial DNA molecules were mapped by restriction fragment analysis of more than 300 cosmid clones and purified mitochondrial DNA. The entire genome of 372 kb is contained in three different configurations of circular molecules and is split into two additional subgenomic molecules of 234 kb and 138 kb, respectively. These arrangements result from recombinations of the two sets of repeats present in combinations of inverted and/or direct orientation. Alignment of YAC clones confirms the in vivo presence of continuous DNA molecules of more than 300 kb in A. thaliana mitochondria. The presence of this comparatively large mitochondrial genome in a plant with one of the smallest nuclear genomes shows that different size constraints act upon the different genomes in plant cells.

Arabidopsis↗
PubMed1993

The mitochondrial genome on its way to the nucleus: different stages of gene transfer in higher plants.

The vast majority of mitochondrial proteins are in all eukaryotes encoded in the nuclear genomes by genes which have been transferred from the original endosymbiont. DNA as well as RNA was and is exchanged between organelles. A functionally successful information transfer, however, requires complex structural and regulatory alterations of the concerned gene. The recently identified variations of the information content in mitochondrial genomes of different plant species represent different stages of the transfer process. These evolutionary intermediates allow a definition of requirements and chances of successful gene transfers.

Cell Nucleus↗
PubMed1993

An adenine nucleotide translocator gene from Arabidopsis thaliana.

The sequence of an adenine nucleotide translocator (ANT) gene of Arabidopsis contains three introns, the first of which is located upstream of the assumed initiation codon. The presequence characteristic for plant ANTs is processed also in Arabidopsis as suggested by Western blot analysis, most likely at the conserved cleavage site.

Amino Acid Sequence↗
PubMed1993

RNA editing in plant mitochondria.

RNA editing in plant mitochondria alters nearly all mRNAs by C to U and U to C transitions. In some species more than 400 edited sites have been identified with significant effects on the encoded proteins. RNA editing occurs in higher and lower plants and presumably has evolved before the differentiation of land plants. Current research focuses on the elucidation of the biochemistry and the specificity determinants of RNA editing in plant mitochondria.

Amino Acid Sequence↗