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Genetic and biochemical analysis of the TLA1 gene in Chlamydomonas reinhardtii.

ABSTRACT: The Chlamydomonas reinhardtii genomic DNA database contains a predicted open reading frame (ORF-P) without an apparent stop-codon and unknown coding sequence, located in close proximity and immediately upstream of the TLA1 gene (GenBank Accession No. AF534570). The latter was implicated in the regulation of the light-harvesting chlorophyll antenna size of photosynthesis (Tetali et al. Planta 225:813-829, 2007). To provide currently lacking information on ORF-P and its potential participation in TLA1 gene expression, thus in the regulation of the chlorophyll antenna size, genetic and biochemical analyses were undertaken. The coding and UTR regions of the ORF-P were defined and delineated from those of the adjacent TLA1 gene. ORF-P is shown to encode a protein with a distinct RING-like zinc finger domain that is present in numerous eukaryotic proteins, believed to play a role in cellular ubiquitination, leading to regulation of cellular processes like signaling, growth, transcription, and DNA repair. It is further shown that the two genes share a 74-bp overlap between the 3' UTR region of ORF-P and the 5' UTR region of TLA1. However, they possess distinct start and stop codons and separate coding sequences, and transcribed as separate mRNAs without any trans-splicing between them. Complementation experiments showed that the TLA1 gene alone is sufficient to rescue the truncated chlorophyll antenna size phenotype of the tla1 mutant. Protein sequence alignments in C. reinhardtii and the colorless microalga Polytomella parva suggested that TLA1 defines the relationship between nucleus and organelle in microalgae, indirectly affecting the development of the chlorophyll antenna size.

SUBMITTER: Mitra M

PROVIDER: S-EPMC2806527 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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Genetic and biochemical analysis of the TLA1 gene in Chlamydomonas reinhardtii.

Mitra Mautusi M Melis Anastasios A

Planta 20091215 3

The Chlamydomonas reinhardtii genomic DNA database contains a predicted open reading frame (ORF-P) without an apparent stop-codon and unknown coding sequence, located in close proximity and immediately upstream of the TLA1 gene (GenBank Accession No. AF534570). The latter was implicated in the regulation of the light-harvesting chlorophyll antenna size of photosynthesis (Tetali et al. Planta 225:813-829, 2007). To provide currently lacking information on ORF-P and its potential participation in ...[more]

PMID: 20012986

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Project description:BackgroundPhotosynthetic organisms convert atmospheric carbon dioxide into numerous metabolites along the pathways to make new biomass. Aquatic photosynthetic organisms, which fix almost half of global inorganic carbon, have great potential: as a carbon dioxide fixation method, for the economical production of chemicals, or as a source for lipids and starch which can then be converted to biofuels. To harness this potential through metabolic engineering and to maximize production, a more thorough understanding of photosynthetic metabolism must first be achieved. A model algal species, C. reinhardtii, was chosen and the metabolic network reconstructed. Intracellular fluxes were then calculated using flux balance analysis (FBA).ResultsThe metabolic network of primary metabolism for a green alga, C. reinhardtii, was reconstructed using genomic and biochemical information. The reconstructed network accounts for the intracellular localization of enzymes to three compartments and includes 484 metabolic reactions and 458 intracellular metabolites. Based on BLAST searches, one newly annotated enzyme (fructose-1,6-bisphosphatase) was added to the Chlamydomonas reinhardtii database. FBA was used to predict metabolic fluxes under three growth conditions, autotrophic, heterotrophic and mixotrophic growth. Biomass yields ranged from 28.9 g per mole C for autotrophic growth to 15 g per mole C for heterotrophic growth.ConclusionThe flux balance analysis model of central and intermediary metabolism in C. reinhardtii is the first such model for algae and the first model to include three metabolically active compartments. In addition to providing estimates of intracellular fluxes, metabolic reconstruction and modelling efforts also provide a comprehensive method for annotation of genome databases. As a result of our reconstruction, one new enzyme was annotated in the database and several others were found to be missing; implying new pathways or non-conserved enzymes. The use of FBA to estimate intracellular fluxes also provides flux values that can be used as a starting point for rational engineering of C. reinhardtii. From these initial estimates, it is clear that aerobic heterotrophic growth on acetate has a low yield on carbon, while mixotrophically and autotrophically grown cells are significantly more carbon efficient.

Dataset Information

Genetic and biochemical analysis of the TLA1 gene in Chlamydomonas reinhardtii.

Publications

Genetic and biochemical analysis of the TLA1 gene in Chlamydomonas reinhardtii.

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