Differential Regulation of Glyceraldehyde-3-Phosphate Dehydrogenases in the Green Alga Chlorella fusca

Abstract

Plants and eukaryotic microalgae posses three distinct glyceraldehyde-3-phosphate dehydrogenases (GAPDH) which are encoded by different nuclear genes, located in different cell compartments and expressed under different conditions (1). We have studied the regulation of these enzymes in a strain of the green alga Chlorella fusca that can use glucose for heterotrophic or mixotrophic growth. By using RT-PCR with RNA extracted from photoautotrophic cells, we cloned partial DNA fragments from the gapN gene encoding the NADP-dependent non-phosphorylating cytosolic enzyme (EC 1.2.1.9, GAPDHNP), the gapA gene for the chloroplastic NADP-dependent phosphorylating enzyme (EC 1.2.1.13, GAPDHA), and the gapC gene for the cytosolic NAD-dependent enzyme (EC 1.2.1.12, GAPDHC). These probes were used for gene expression studies by Northern blots in cells grown under different conditions and the results compared with activity and Western blot data obtained with cell-free extracts. Although glycolysis has been extensively studied in bacteria and vertebrates, there are marked differences with the structure, compartmentalization and regulation of this degradative pathway in plants and algae (2). Our results strongly suggest that, in photosynthetic eukaryotes, carbohydrate catabolism is carried out both in the chloroplast and the cytosol and that in the latter cell compartment two different degradative pathways may be operative, one involving the phosphorylating GAPDH and another implicating the non-phosphorylating one.