NO-mediated apoptosis in yeast

Abstract

Nitric oxide (NO) is a small molecule with distinct roles in diverse physiological functions in biological systems, among them the control of the apoptotic signalling cascade. By combining proteomic, genetic and biochemical approaches we demonstrate that NO and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) are crucial mediators of yeast apoptosis. Using indirect methodologies and a NO-selective electrode, we present results showing that H2O2-induced apoptotic cells synthesize NO that is associated to a nitric oxide synthase (NOS)-like activity as demonstrated by the use of a classical NOS kit assay. Additionally, our results show that yeast GAPDH is a target of extensive proteolysis upon H2O2-induced apoptosis and undergoes S-nitrosation. Blockage of NO synthesis with Nω-nitro-L-arginine methyl ester leads to a decrease of GAPDH S-nitrosation and of intracellular reactive oxygen species (ROS) accumulation, increasing survival. These results indicate that NO signalling and GAPDH S-nitrosation are linked with H2O2-induced apoptotic cell death. Evidence is presented showing that NO and GAPDH S-nitrosation also mediate cell death during chronological life span pointing to a physiological role of NO in yeast apoptosis.

Different carbon sources affect lifespan and protein redox state during Saccharomyces cerevisiae chronological ageing

“…Recent studies have revealed that Gapdh is a redox-sensitive glycolytic enzyme and it is involved in neuronal cell death which is triggered by oxidative stress. In fact this stress induces Glyceraldehyde-3-phosphate dehydrogenase 3 oxidation: [1,2] carbonylation: [3,4], S-thiolation: [5] oxidative stress, apoptosis, ageing Protein MET17 includes: O-acetylhomoserine sulfhydrylase oxidation: [1], this work oxidative stress…”

“…A great number of recent reports confirm the Harman "free radical theory of ageing" proposed some fifty years ago although many unknown details still remain. For example, although many proteins are found in an oxidized state during cell senescence [1,2,3,4,5], the identity of the more relevant targets of oxidation and how their modification can affect cell lifespan remains unknown [6]. In the budding yeast S. cerevisiae two types of lifespan can be measured: replicative and chronological.…”

“…Otherwise, partial oxidation (disulfide bond or sulfenic acid form) is reversible and is known to play an important role in the regulation of transcription factors, for example Yap1 in S. cerevisiae [19], and several phosphotyrosine phosphatases in mammals [20]. Recently S-nitrosylation was found to take place in S. cerevisiae although the mechanism of NO production is still a matter for debate because of the lack of mammalian nitric oxide synthase (NOS) orthologues in the yeast genome [2]. However, Castello et al found that yeast cells are capable of producing NO in mitochondria under hypoxic conditions [21].…”

Apoptosis in yeast: triggers, pathways, subroutines

“…Inhibition of NO synthesis promotes survival by decreasing both GAPDH S-nitrosation and intracellular ROS accumulation. 63 During chronological aging, scavenging of NO by oxyhemoglobin delays the onset of death and reduces the levels of superoxide anion. 63 Interestingly, the NO-target GAPDH has been implicated both in H 2 O 2 -induced yeast apoptosis 64 and in the regulation of apoptosis in human cells.…”

“…63 During chronological aging, scavenging of NO by oxyhemoglobin delays the onset of death and reduces the levels of superoxide anion. 63 Interestingly, the NO-target GAPDH has been implicated both in H 2 O 2 -induced yeast apoptosis 64 and in the regulation of apoptosis in human cells. 65 Finally, during development of multicellular yeast colonies, ammonia accumulates in the center of the colonies and triggers the death of older cells, allowing young cells on the rim to exploit the released nutrients.…”