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S A Rensing

Publications and source records attributed to S A Rensing.

14 recordsLinked to original sources

Representation and high-quality annotation of the Physcomitrella patens transcriptome demonstrates a high proportion of proteins involved in metabolism in mosses.

To gain insight into the transcriptome of the well-used plant model system Physcomitrella patens, several EST sequencing projects have been undertaken. We have clustered, assembled, and annotated all publicly available EST and CDS sequences in order to represent the transcriptome of this non-seed plant. Here, we present our fully annotated knowledge resource for the Physcomitrella patens transcriptome, integrating annotation from the production process of the clustered sequences and from a high-quality annotation pipeline developed during this study. Each transcript is represented as an entity containing full annotations and GO term associations. The whole production, filtering, clustering, and annotation process is being modelled and results in seven datasets, representing the annotated Physcomitrella transcriptome from different perspectives. We were able to annotate 63.4 % of the 26 123 virtual transcripts. The transcript archetype, as covered by our clustered data, is compared to a compilation based on all available Physcomitrella full length CDS. The distribution of the gene ontology annotations (GOA) for the virtual transcriptome of Physcomitrella patens demonstrates consistency in the ratios of the core molecular functions among the plant GOA. However, the metabolism subcategory is over-represented in bryophytes as compared to seed plants. This observation can be taken as an indicator for the wealth of alternative metabolic pathways in moss in comparison to spermatophytes. All resources presented in this study have been made available to the scientific community through a suite of user-friendly web interfaces via www.cosmoss.org and form the basis for assembly and annotation of the moss genome, which will be sequenced in 2005.

3' Untranslated Regions↗

Large-scale analysis of 73 329 physcomitrella plants transformed with different gene disruption libraries: production parameters and mutant phenotypes.

Gene targeting in the moss Physcomitrella patens has created a new platform for plant functional genomics. We produced a mutant collection of 73 329 Physcomitrella plants and evaluated the phenotype of each transformant in comparison to wild type Physcomitrella. Production parameters and morphological changes in 16 categories, such as plant structure, colour, coverage with gametophores, cell shape, etc., were listed and all data were compiled in a database (mossDB). Our mutant collection consists of at least 1804 auxotrophic mutants which showed growth defects on minimal Knop medium but were rescued on supplemented medium. 8129 haploid and 11 068 polyploid transformants had morphological alterations. 9 % of the haploid transformants had deviations in the leaf shape, 7 % developed less gametophores or had a different leaf cell shape. Other morphological deviations in plant structure, colour, and uniformity of leaves on a moss colony were less frequently observed. Preculture conditions of the plant material and the cDNA library (representing genes from either protonema, gametophore or sporophyte tissue) used to transform Physcomitrella had an effect on the number of transformants per transformation. We found correlations between ploidy level and plant morphology and growth rate on Knop medium. In haploid transformants correlations between the percentage of plants with specific phenotypes and the cDNA library used for transformation were detected. The number of different cDNAs present during transformation had no effect on the number of transformants per transformation, but it had an effect on the overall percentage of plants with phenotypic deviations. We conclude that by linking incoming molecular, proteome, and metabolome data of the transformants in the future, the database mossDB will be a valuable biological resource for systems biology.

Bryopsida↗

Visualization of a cytoskeleton-like FtsZ network in chloroplasts.

It has been a long-standing dogma in life sciences that only eukaryotic organisms possess a cytoskeleton. Recently, this belief was questioned by the finding that the bacterial cell division protein FtsZ resembles tubulin in sequence and structure and, thus, may be the progenitor of this major eukaryotic cytoskeletal element. Here, we report two nuclear-encoded plant ftsZ genes which are highly conserved in coding sequence and intron structure. Both their encoded proteins are imported into plastids and there, like in bacteria, they act on the division process in a dose-dependent manner. Whereas in bacteria FtsZ only transiently polymerizes to a ring-like structure, in chloroplasts we identified persistent, highly organized filamentous scaffolds that are most likely involved in the maintenance of plastid integrity and in plastid division. As these networks resemble the eukaryotic cytoskeleton in form and function, we suggest the term "plastoskeleton" for this newly described subcellular structure.

Arabidopsis↗

Substitution rate calibration of small subunit ribosomal RNA identifies chlorarachniophyte endosymbionts as remnants of green algae.

Chlorarachniophytes are amoeboid algae with chlorophyll a and b containing plastids that are surrounded by four membranes instead of two as in plants and green algae. These extra membranes form important support for the hypothesis that chlorarachniophytes have acquired their plastids by the ingestion of another eukaryotic plastid-containing alga. Chlorarachniophytes also contain a small nucleus-like structure called the nucleomorph situated between the two inner and the two outer membranes surrounding the plastid. This nucleomorph is a remnant of the endosymbiont's nucleus and encodes, among other molecules, small subunit ribosomal RNA. Previous phylogenetic analyses on the basis of this molecule provided unexpected and contradictory evidence for the origin of the chlorarachniophyte endosymbiont. We developed a new method for measuring the substitution rates of the individual nucleotides of small subunit ribosomal RNA. From the resulting substitution rate distribution, we derived an equation that gives a more realistic relationship between sequence dissimilarity and evolutionary distance than equations previously available. Phylogenetic trees constructed on the basis of evolutionary distances computed by this new method clearly situate the chlorarachniophyte nucleomorphs among the green algae. Moreover, this relationship is confirmed by transversion analysis of the Chlorarachnion plastid small subunit ribosomal RNA.

Amoeba↗

Characterization of a maize ribosomal P2 protein cDNA and phylogenetic analysis of the P1/P2 family of ribosomal proteins.

The nucleotide sequence of a full-length ribosomal P2 protein cDNA from maize was determined and used for a sequence comparison with the P2 and P1 proteins from other organisms. The integration of these data into a phylogenetic tree shows that the P proteins separated into the subspecies P1 and P2 before the eukaryotic kingdoms including plants developed from their ancestor.

Base Sequence↗

Twintrons are not unique to the Euglena chloroplast genome: structure and evolution of a plastome cpn60 gene from a cryptomonad.

Introns within introns (twintrons) are known only from the Euglena chloroplast genome. Twintrons are group II or III introns, into which another group II or III intron has been transposed. In this paper we describe a non-Euglena twintron structure within a plastid-encoded chaperone gene (cpn60) of the cryptomonad alga Pyrenomonas salina. In addition, the evolutionary relationships between members of the Cpn60 protein family are determined. Our findings permit the inclusion of cryptomonad plastomes in phylogenetic studies of intron evolution and present further evidence for the origin of modern plastids from a cyanobacterial ancestor.

Amino Acid Sequence↗

The cryptomonad histone H4-encoding gene: structure and chromosomal localization.

Cryptomonads are unicellular flagellates whose plastids are surrounded by four membranes. A periplastidal compartment, containing eukaryote-type ribosomes, starch grains and a so-called nucleomorph, is located between the inner and outer membrane pairs. The nucleomorph has been shown to be the vestigial nucleus of a eukaryotic endosymbiont. In order to obtain more information about the chromatin structure of the nucleomorph and the host nuclear chromosomes, we studied the distribution of the histone, H4. H4 was not detectable in the nucleomorph by immunolocalization, thus supporting earlier findings by Gibbs [In: Wiesner et al. (Eds.), Experimental Phycology 1, 1990, pp. 145-157]. Likewise, no H4 DNA was demonstrable in the nucleomorph by Southern hybridization. Sequence analysis, and Southern and Northern blot data of a cryptomonad gene, H4, indicate an intermediate position for these genes between animals and plants.

Amino Acid Sequence↗

The smallest known eukaryotic genomes encode a protein gene: towards an understanding of nucleomorph functions.

Cryptomonads are unicellular algae with plastids surrounded by four membranes. Between the two pairs of membranes lies a periplastidal compartment that harbours a DNA-containing organelle, termed the nucleomorph. The nucleomorph is the vestigial nucleus of a phototrophic, eukaryotic endosymbiont. Subcloning of parts of one nucleomorph chromosome revealed a gene coding for an Hsp70 protein. We demonstrate the expression of this nucleomorph protein-coding gene and present a model for protein transport from the host to the endosymbiont compartment.

Biological Transport↗

The SecY protein family: comparative analysis and phylogenetic relationships.

We have cloned and sequenced the plastome encoded secY homologue of the cryptomonad alga Pyrenomonas salina. In this study we have carried out a comparative analysis of all but one fully sequenced proteins of the SecY family. We present an alignment of 16 SecY family proteins, containing family signatures, putative transmembrane domains, consensus sequence, and conservation grade. A phylogenetic tree derived from the conserved blocks of the alignment reveals the relationships among this protein family. The tree shows division into two broad subfamilies, one consisting of prokaryotic and plastidal sequences and the other of eukaryotic as well as archaeal sequences.

Amino Acid Sequence↗

Evidence that stilbene synthases have developed from chalcone synthases several times in the course of evolution.

Chalcone (CHS) and stilbene (STS) synthases are related plant-specific polyketide synthases that are key enzymes in the biosynthesis of flavonoids and of stilbene phytoalexins, respectively. A phylogenetic tree constructed from 34 CHS and four STS sequences revealed that the STS formed no separate cluster but grouped with CHS from the same or related plants. This suggested that STS evolved from CHS several times independently. We attempted to stimulate this by site-directed mutagenesis of an interfamily CHS/STS hybrid, which contained 107 amino acids of a CHS from Sinapis alba (N-terminal) and 287 amino acids of a STS from Arachis hypogaea. The hybrid had no enzyme activity. Three amino acid exchanges in the CHS part (Gln-100 to Glu, Val-103 to Met, Val-105 to Arg) were sufficient to obtain low STS activity, and one additional exchange (Gly-23 to Thr) resulted in 20-25% of the parent STS activity. A kinetic analysis indicated (1) that the hybrids had the same Km for the substrate 4-coumaroyl-CoA but a lower Vmax than the parent STS, and (2) that they had a different substrate preference than the parent STS and CHS. Most of the other mutations and their combinations led to enzymatically inactive protein aggregates, suggesting that the subunit folding and/or the dimerization was disturbed. We propose that STS evolved from CHS by a limited number of amino acid exchanges, and that the advantage gained by this enzyme function favored the selection of plants with improved STS activity.

Acyltransferases↗

Phylogenetic analysis of the stress-70 protein family.

The eukaryotic cyto-/nucleoplasmatic 70-kDa heat-shock protein (HSP70) has homologues in the endoplasmic reticulum as well as in bacteria, mitochondria, and plastids. We selected a representative subset from the large number of sequenced stress-70 family members which covers all known branches of the protein family and calculated and manually improved an alignment. Here we present the consensus sequence of the aligned proteins and putative nuclear localization signals (NLS) in the eukaryotic HSP70 homologues. The phylogenetic relationships of the stress-70 group family members were estimated by use of different computation methods. We present a phylogenetic tree containing all known stress-70 subfamilies and demonstrate the usefulness of stress-70 protein sequences for the estimation of intertaxonic phylogeny.

Amino Acid Sequence↗

Phylogenetic relationships of HMG box DNA-binding domains.

HMG boxes were initially identified as DNA-binding domains of the human RNA polymerase I (pol I) transcription factor hUBF and the animal high-mobility-group (HMG) protein family HMG1. Since then, numerous sequences of HMG-box-containing HMG proteins and other DNA-binding proteins from several species have become available. By sequence comparisons of a selected range of HMG boxes from these proteins and the construction of phylogenetic trees we show that the HMG box is highly conserved between DNA-binding proteins of organisms from all three eukaryotic kingdoms and that HMG boxes are linked by distinct evolutionary relationships. In addition, most HMG boxes display comparable hydropathy profiles and amino acid arrangements, which could serve as nuclear targeting sequences.

Amino Acid Sequence↗

cDNA cloning of a Sec61 homologue from the cryptomonad alga Pyrenomonas salina.

Sec61 is an endoplasmic reticulum transmembrane protein involved in the process of translocation of proteins across this membrane. To-date, the only cloned genes for Sec61 are derived from mammals and yeast. In this paper, we present the first full-length cDNA from a sec61 gene of a plant cell. Comparison of the predicted protein sequence with all known Sec61 proteins, as well as with the bacterial/plastome-encoded homologue SecY, demonstrates a high degree of similarity among the SecY/Sec61 family.

Amino Acid Sequence↗

The presence of a nucleomorph hsp70 gene is a common feature of Cryptophyta and Chlorarachniophyta.

Cryptomonad algae and Chlorarachniophyta are evolutionary chimaeras derived from the engulfment of an eukaryotic phototrophic endosymbiont by a eukaryotic host cell. Although much reduced, the endosymbiont's eukaryotic plasmatic compartment still contains a nucleus, the so-called nucleomorph. These nucleomorphs carry the smallest known eukaryotic genomes. We have characterized the genomes of several cryptomonads and a Chlorarachnion species by means of PFGE (pulsed-field gel electrophoresis). Hybridization studies with small subunit rDNA were used to identify the nucleomorph chromosomes. We also performed hybridization experiments with an hsp70 probe to estimate the distribution of this gene among the different algal species. The evolutionary, genetical, and physiological implications of our studies are discussed. A model on the possible function of the nucleomorph hsp70 gene products is presented.

Blotting, Southern↗