Involvement of GFA1, which encodes glutamine-fructose-6-phosphate amidotransferase, in the activation of the chitin synthesis pathway in response to cell-wall defects in Saccharomyces cerevisiae.

Arnaud Lagorce (1) , Véronique Le Berre (2, 3, 4) , Blanca Aguilar-Uscanga , Hélène Martin-Yken (3) , Adilia Dagkessamanskaia (3) , Jean François (5)

1 IGM - Institut de génétique et microbiologie [Orsay]
2 Plateforme Biopuces
3 LISBP - Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés
4 TBI - Toulouse Biotechnology Institute
5 ITAV - Institut des Technologies Avancées en sciences du Vivant

Cell-wall damage caused by mutations of cell-wall-related genes triggers a compensatory mechanism which eventually results in hyperaccumulation of chitin reaching 20% of the cell-wall dry mass. We show that activation of chitin synthesis is accompanied by a rise, from 1.3-fold to 3.5-fold according to the gene mutation, in the expression of most of the genes encoding enzymes of the chitin metabolic pathways. Evidence that GFA1, which encodes glutamine-fructose-6-Phosphate amidotransferase (Gfa1p), the first committed enzyme of this pathway, plays a major role in this process was as follows. Activation of chitin synthesis in the cell-wall mutants correlated with activation of GFA1 and with a proportional increase in Gfa1p activity. Overexpression of GFA1 caused an approximately threefold increase in chitin in the transformed cells, whereas chitin content was barely affected by the joint overexpression of CHS3 and CHS7. Introduction of a gfa1-97 allele mutation in the cell-wall-defective gas1Delta mutant or cultivation of this mutant in a hyperosmotic medium resulted in reduction in chitin synthesis that was proportional to the decrease in Gfa1p activity. Finally, the stimulation of chitin production was also accompanied by an increase in pools of fructose 6-Phosphate, a substrate of Gfa1p. In quantitative terms, we estimated the flux-coefficient control of Gfa1p to be in the range of 0.90, and found that regulation of the chitin metabolic pathway was mainly hierarchical, i.e. dominated by regulation of the amount of newly synthesized GFA1 protein. In the search for the mechanism by which GFA1 is activated in response to cell-wall perturbations, we could only show that neither MCM1 nor RLM1, which encode two transcriptional factors of the MADS box family that are required for expression of cell-cycle and cell-wall-related genes, was involved in this process.

Mots clés: cell wall; chitin; GFA1; metabolic control; Saccharomyces cerevisiae

Domaines: Sciences du Vivant [q-bio]; Biochimie, Biologie Moléculaire; Génomique, Transcriptomique et Protéomique [q-bio.GN]

Arnaud Lagorce, Véronique Le Berre, Blanca Aguilar-Uscanga, Hélène Martin-Yken, Adilia Dagkessamanskaia, et al.. Involvement of GFA1, which encodes glutamine-fructose-6-phosphate amidotransferase, in the activation of the chitin synthesis pathway in response to cell-wall defects in Saccharomyces cerevisiae.. European Journal of Biochemistry, 2002, pp.1697-707. ⟨hal-01849238⟩