Dataset Information

Reduction of Phosphoribulokinase Amount and Re-Routing Metabolism in Chlamydomonas reinhardtii CP12 Mutants

ABSTRACT: The chloroplast protein CP12 is involved in the dark/light regulation of the Calvin-Benson-Bassham cycle, and in particular in the dark inhibition of two enzymes: glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK), but other functions related to stress have been proposed. We knocked out the unique CP12 gene to prevent its expression in Chlamydomonas reinhardtii (ΔCP12). The growth rates of both wild-type and ΔCP12 cells were nearly identical, as was the GAPDH protein abundance and activity in both cell lines. On the contrary, the abundance of PRK and its specific activity were significantly reduced in ΔCP12, as revealed by relative quantitative proteomics. Isolated PRK lost irreversibly its activity over-time in vitro, that was prevented in the presence of recombinant CP12 in a redox-independent manner. We have identified amino acid residues in the CP12 protein that are required for this new function preserving PRK activity. Numerous proteins involved in redox homeostasis and stress responses were more abundant and the expressions of various metabolic pathways were also increased or decreased in the absence of CP12. These results highlight CP12 as a moonlighting protein with additional functions beyond its well-known regulatory role in carbon metabolism.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Chlamydomonas Reinhardtii

SUBMITTER: Regine Lebrun

LAB HEAD: Lebrun Regine

PROVIDER: PXD031803 | Pride | 2022-04-04

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	2021118_03_029_BMG_GdtL_Wt1.raw	Raw
	2021118_06__029_BMG_GdtL_Wt2.raw	Raw
	2021118_09__029_BMG_GdtL_Wt3.raw	Raw
	2021118_15__029_BMG_GdtL_dCP12_2.raw	Raw
	2021118_18__029_BMG_GdtL_dCP12_3.raw	Raw

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Publications

Reduction in Phosphoribulokinase Amount and Re-Routing Metabolism in Chlamydomonas reinhardtii CP12 Mutants.

Gérard Cassy C Lebrun Régine R Lemesle Erwan E Avilan Luisana L Chang Kwang Suk KS Jin EonSeon E Carrière Frédéric F Gontero Brigitte B Launay Hélène H

International journal of molecular sciences 20220228 5

The chloroplast protein CP12 is involved in the dark/light regulation of the Calvin-Benson-Bassham cycle, in particular, in the dark inhibition of two enzymes: glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK), but other functions related to stress have been proposed. We knocked out the unique CP12 gene to prevent its expression in Chlamydomonas reinhardtii (ΔCP12). The growth rates of both wild-type and ΔCP12 cells were nearly identical, as was the GAP ...[more]

PMID: 35269851

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Project description:Background: Calorie restriction (CR) is the only intervention known to extend lifespan in a wide range of organisms, including mammals. However, the mechanisms by which it regulates mammalian aging remain largely unknown and the involvement of the TOR and Sirtuin pathways (which regulate aging in lower organisms) remain controversial. Femaleness is a second phenotype generally associated with longevity but the relationship between sex-biased and CR-induced gene expression remains undetermined. Methodology/Principal Findings: We generated microarray gene expression data from livers of male mice fed high calorie or CR diets, and we find that CR significantly changes the expression of over 3,000 genes, many between 10- and 50-fold. We compare our data to the GenAge database of known aging-related genes and to prior microarray expression data of genes expressed differently between male and female mice. CR generally feminizes gene expression and many of the most significantly changed individual genes are involved in aging, hormone signaling, and p53-associated regulation of the cell cycle and apoptosis. Among the genes showing the largest and most statistically significant CR-induced expression differences are Ddit4, a key regulator of the TOR pathway, and Nnmt, a regulator of lifespan linked to the Sirtuin pathway. Using Western analyses, we confirmed post-translational inhibition of the TOR pathway. Conclusions: Our data show that CR induces widespread gene expression changes and acts through highly evolutionarily conserved pathways, from microorganisms to mammals, and that its life-extension effects might arise partly from a shift toward a gene expression profile more typical of the longer-lived female sex. Keywords: Two-class gene expression comparison. Calorie restriction (CR) versus HIGH CALORIE feeding. Design of the experiment is a two-class paired design in which the two classes are HIGH CALORIE and CALORIE RESTRICTION (CR) dietary regimens fed to mice, resulting in 15 HIGH CALORIE microarrays and 8 CR microarrays; 23 total microarrays. Four HIGH CALORIE and 2 CR microarrays were produced using pools of 3 RNA samples for each microarray (6 pools of 3, plus 17 individual samples = 35 individual mouse liver samples, 23 microarrays). Total liver RNA was labeled directly. Reference RNA: Stratagene Universal Mouse Reference RNA.

Dataset Information

Reduction of Phosphoribulokinase Amount and Re-Routing Metabolism in Chlamydomonas reinhardtii CP12 Mutants

Dataset's files

Publications

Reduction in Phosphoribulokinase Amount and Re-Routing Metabolism in <i>Chlamydomonas reinhardtii</i> CP12 Mutants.

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