Regulation of plant cytosolic glyceraldehyde 3‐phosphate dehydrogenase isoforms by thiol modifications

Abstract

Cytosolic NAD-dependent glyceraldehyde 3-P dehydrogenase (GAPDH; GapC; EC 1.2.1.12) catalyzes the oxidation of triose phosphates during glycolysis in all organisms, but additional functions of the protein has been put forward. Because of its reactive cysteine residue in the active site, it is susceptible to protein modification and oxidation. The addition of GSSG, and much more efficiently of S-nitrosoglutathione, was shown to inactivate the enzymes from Arabidopsis thaliana (isoforms GapC1 and 2), spinach, yeast and rabbit muscle. Inactivation was fully or at least partially reversible upon addition of DTT. The incorporation of glutathione upon formation of a mixed disulfide could be shown using biotinylated glutathione ethyl ester. Furthermore, using the biotin-switch assay, nitrosylated thiol groups could be shown to occur after treatment with nitric oxide donors. Using mass spectrometry and mutant proteins with one cysteine lacking, both cysteines (Cys-155 and Cys-159) were found to occur as glutathionylated and as nitrosylated forms. In preliminary experiments, it was shown that both GapC1 and GapC2 can bind to a partial gene sequence of the NADP-dependent malate dehydrogenase (EC 1.2.1.37; At5g58330). Transiently expressed GapC-green fluorescent protein fusion proteins were localized to the nucleus in A. thaliana protoplasts. As nuclear localization and DNA binding of GAPDH had been shown in numerous systems to occur upon stress, we assume that such mechanism might be part of the signaling pathway to induce increased malate-valve capacity and possibly other protective systems upon overreduction and initial formation of reactive oxygen and nitrogen species as well as to decrease and protect metabolism at the same time by modification of essential cysteine residues.

Phosphatidic Acid Binds to Cytosolic Glyceraldehyde-3-phosphate Dehydrogenase and Promotes Its Cleavage in Arabidopsis

“…6B). The nuclear localization of GAPC is consistent with previous reports (42,43). Semiquantification by densitometric analysis suggests that ϳ98% of total GAPC2 is in the cytosol and 2% in the nucleus.…”

Nuclear Accumulation of Cytosolic Glyceraldehyde-3-Phosphate Dehydrogenase in Cadmium-Stressed Arabidopsis Roots

“…Although complex formation in the nucleus was not unaffected by stress conditions or NO donors, the interaction was abolished when both active-site cysteines of GAPDH were mutated into serines (Wawer et al, 2010). These studies were performed with Arabidopsis protoplasts expressing GAPDH under the control of the p35S promoter (Holtgrefe et al, 2008;Wawer et al, 2010), and in similar experiments, we could nicely confirm the nuclear localization of GAPC1-YFP expressed under the control of p35S (data not shown). However, replacing this constitutive strong promoter with the GAPC1 endogenous promoter largely prevented, in control conditions, the nuclear accumulation of GAPC1 (Supplemental Fig.…”

“…Anderson et al (2004) have shown by immunogold localization the presence of GAPC both in the cytosol and the nucleus of pea leaf cells. Holtgrefe et al (2008) have proposed that, in Arabidopsis protoplasts, both GAPC1 and GAPC2 could localize also in the nucleus, in addition to the cytosol, and bind DNA. In the same Arabidopsis system, tobacco (Nicotiana tabacum) cytosolic GAPDH was shown to interact, both in the cytosol and in the nucleus, with an osmotic stress-activated protein kinase (NtOSAK) from tobacco too (Wawer et al, 2010).…”

“…Because of the high reactivity of its catalytic Cys, cytosolic glyceraldehyde 3-P dehydrogenase (GAPDH) has been identified as a target of different types of redox modifications (Hancock et al, 2005(Hancock et al, , 2006Lindermayr et al, 2005;Holtgrefe et al, 2008;Bedhomme et al, 2012).…”

Pollen tube growth: where does the energy come from?

“…Therefore, GapCs possibly function as sensor molecules for the detection of H 2 O 2 diffusing out of the chloroplast or mitochondria upon imbalances of the electron transport chains and are part of a redox-dependent retrograde signal transduction network. [81][82][83][84] Due to fact that gapc1gapc2 double knock-out mutants are fertile, 85 whereas gapcp1gapcp2 double mutants are male sterile, 64 it can be assumed that plastidic glycolysis is more important for pollen maturation and development than is cytosolic glycolysis.…”