E-GEOD-51602 - Systems-level analysis of N-starvation induced modifications of carbon metabolism in a Chlamydomonas starchless mutant

Status
Released on 24 October 2013, last updated on 3 May 2014
Organism
Chlamydomonas reinhardtii
Samples (31)
Protocols (4)
Description
To understand the molecular basis underlying increased triacylglycerol accumulation in starchless Chlamydomonas mutants, we undertook comparative time course transcriptomics of sta6 (CC-4348), a cw strain purported to represent the parental STA6 strain (CC-4349), and 3 independent STA6 strains (STA6-C2, STA6-C4 and STA6-C6, generated by complementation of sta6, CC-4565, –4566 and –4567) in the context of nitrogen deprivation. Despite the N starvation induced dramatic remodeling of the transcriptome, there are relatively few differences (5 x 102) between sta6 and STA6, the most dramatic of which are in the abundance of transcripts encoding key regulated or rate-limiting steps in central carbon metabolism, specifically ICL1, MAS1, TAL1, FBP1 and PCK1, suggestive of increased carbon movement towards hexosephosphate in sta6 by up-regulation of the glyoxylate pathway and gluconeogenesis. Enzyme assays validated the increase in isocitrate lyase and malate synthase activities. Targeted metabolite analysis indicates increased succinate and malate, increased Glc-6-P and decreased fructose-1,6-bisphosphate, which documents the impact of these changes. Comparisons of the independent datasets in multiple strains allowed the delineation of a sequence of events in a global N-starvation response in Chlamydomonas, starting within minutes with up-regulation of alternative N assimilation routes and carbohydrate synthesis and subsequently a more gradual up-regulation of genes encoding enzymes of TAG synthesis. Finally, genome re-sequencing analysis indicated that i) the deletion in sta6 extends into the neighboring gene, RBO1, and ii) a commonly-used STA6 strain (CC-4349) as well as the sequenced reference (CC-503) are not congenic with respect to sta6 (CC-4348), underscoring the importance of using complemented strains for more rigorous assignment of phenotype to genotype. The transcriptomes of five different Chlamydomonas strains (wild type CC-4349, sta6 mutant (CC-4348),and three indepdent STA6 strains (STA6-C2, STA6-C4 and STA6-C6) are profiled by RNA-Seq in timecourse experiments after the transition to minus nitrogen
Experiment type
RNA-seq of coding RNA 
Contacts
Sabeeha Merchant <merchant@chem.ucla.edu>, Anne Glaesener, Bensheng Liu, Christoph Benning, David Casero, Ian Blaby, Janette Kropat, Mark Stitt, Matteo Pellegrini, Nanette Boyle, Sean Gallaher, Shannon Johnson, Sorel Fitz-Gibbon, Tabea Mettler
MINSEQE
Exp. designProtocolsVariablesProcessedSeq. reads
Files
Investigation descriptionE-GEOD-51602.idf.txt
Sample and data relationshipE-GEOD-51602.sdrf.txt
Additional data (1)E-GEOD-51602.additional.1.zip
Links