title: Profiling Chlamydomonas metabolism under dark, anoxic H2-producing conditions using a combined proteomic, transcriptomic, and metabolomic approach.
abstract: Chlamydomonas reinhardtii is well adapted to survive under different environmental conditions due to the unique flexibility of its metabolism. Here we report metabolic pathways that are active during acclimation to anoxia, but were previously not thoroughly studied under dark, anoxic H2-producing conditions in this model green alga. Proteomic analyses, using 2D-differential in-gel electrophoresis in combination with shotgun mass fingerprinting, revealed increased levels of proteins involved in the glycolytic pathway downstream of 3-phosphoglycerate, the glyoxylate pathway, and steps of the tricarboxylic acid (TCA) reactions. Upregulation of the enzyme, isocitrate lyase (ICL), was observed, which was accompanied by increased intracellular succinate levels, suggesting the functioning of glyoxylate pathway reactions. The ICL-inhibitor study revealed presence of reverse TCA reactions under these conditions. Contributions of the serine-isocitrate lyase pathway, glycine cleavage system, and c1-THF/serine hydroxymethyltransferase pathway in the acclimation to dark anoxia were found. We also observed increased levels of amino acids (AAs) suggesting nitrogen reorganization in the form of de novo AA biosynthesis during anoxia. Overall, novel routes for reductant utilization, in combination with redistribution of carbon and nitrogen, are used by this alga during acclimation to O2 deprivation in the dark.
authors.affiliation: National Renewable Energy Laboratory , Golden, Colorado 80401, United States.
authors.forename: Venkataramanan
authors.initials: V
authors.lastname: Subramanian
authors.forename: Alexandra
authors.initials: A
authors.lastname: Dubini
authors.forename: David P
authors.initials: DP
authors.lastname: Astling
authors.forename: Lieve M L
authors.initials: LM
authors.lastname: Laurens
authors.forename: William M
authors.initials: WM
authors.lastname: Old
authors.forename: Arthur R
authors.initials: AR
authors.lastname: Grossman
authors.forename: Matthew C
authors.initials: MC
authors.lastname: Posewitz
authors.forename: Michael
authors.initials: M
authors.lastname: Seibert
mesh_terms: D020418:Algal Proteins / Q000235:genetics / Q000378:metabolism; D000596:Amino Acids / Q000378:metabolism; D000693:Anaerobiosis; D015153:Blotting, Western; D002244:Carbon / Q000378:metabolism; D016825:Chlamydomonas reinhardtii / Q000235:genetics* / Q000378:metabolism*; D003624:Darkness; D015180:Electrophoresis, Gel, Two-Dimensional; D008401:Gas Chromatography-Mass Spectrometry; D020869:Gene Expression Profiling / Q000379:methods*; D006859:Hydrogen / Q000378:metabolism*; D013058:Mass Spectrometry; D053858:Metabolic Networks and Pathways / Q000235:genetics; D055442:Metabolome; D055432:Metabolomics / Q000379:methods*; D009584:Nitrogen / Q000378:metabolism; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D020418:Algal Proteins; D000596:Amino Acids; D020543:Proteome; D002244:Carbon; D006859:Hydrogen; D009584:Nitrogen
keywords: Chlamydomonas reinhardtii; TCA reactions; amino acid synthesis; glyoxylate pathway; hydrogen production; isocitrate lyase; proteomics