Abstract
Adenosine 3′, 5′-cyclic monophosphate (cAMP) and guanosine 3′, 5′-cyclic monophosphate (cGMP) are well-studied second messengers that transmit extracellular signals into mammalian cells, with conserved functions in various other species such as Caenorhabditis elegans (C. elegans). cAMP is generated by adenylyl cyclases, and cGMP is generated by guanylyl cyclases, respectively. Studies using C. elegans have revealed additional roles for cGMP signaling in lifespan extension. For example, mutants lacking the function of a specific receptor-bound guanylyl cyclase, DAF-11, have an increased life expectancy. While the daf-11 phenotype has been attributed to reductions in intracellular cGMP concentrations, the actual content of cyclic nucleotides has not been biochemically determined in this system. Similar assumptions were made in studies using phosphodiesterase loss-of-function mutants or using adenylyl cyclase overexpressing mutants. In the present study, cyclic nucleotide regulation in C. elegans was studied by establishing a special nematode protocol for the simultaneous detection and quantitation of cyclic nucleotides. We also examined the influence of reactive oxygen species (ROS) on cyclic nucleotide metabolism and lifespan in C. elegans using highly specific HPLC-coupled tandem mass-spectrometry and behavioral assays. Here, we show that the relation between cGMP and survival is more complex than previously appreciated.
Highlights
The first description of guanosine 39, 59-cyclic monophosphate as a biological substance can be dated back to 1963 [1]. cGMP is a well-established second messenger, like the earlier identified adenosine 39, 59-cyclic monophosphate [2,3]
An HPLC-MS/MS Assay for cAMP and cGMP in C. elegans Wild-type animals were synchronized by bleaching and 72 h old adult hermaphrodites were freeze-cracked to break their cuticle in order to prepare the extracts
No change in cAMP levels were observed in the pde1,2,3,5 quadruple mutant compared to wild-type, while the animals displayed a strong increase in cGMP concentration (Figure 1 C, D)
Summary
The first description of guanosine 39, 59-cyclic monophosphate (cGMP) as a biological substance can be dated back to 1963 [1]. cGMP is a well-established second messenger, like the earlier identified adenosine 39, 59-cyclic monophosphate (cAMP) [2,3]. CGMP is a well-established second messenger, like the earlier identified adenosine 39, 59-cyclic monophosphate (cAMP) [2,3]. These cyclic nucleotides (cNMPs) are generated by the ATP- and GTP-converting adenylyl- and guanylyl cyclase, respectively [1,2,3,4,5]. The genome of C. elegans harbors 32 genes with similarity to guanylyl cyclases These include 25 membrane-bound, receptor-like guanylyl cyclases and 7 cytosolic guanylyl cyclases (gcy-31–gcy-37) [13]. Two of the receptor-bound guanylyl cyclase genes, daf-11 and odr, are expressed in olfactory and pheromone-sensing neurons and act downstream of G-protein-coupled receptors. Two of the receptor-bound guanylyl cyclase genes, daf-11 and odr, are expressed in olfactory and pheromone-sensing neurons and act downstream of G-protein-coupled receptors. daf-11 is expressed in the ciliated ASI, ASJ, AWC, AWB, ASK neurons where it is involved in olfactory and pheromone sensing and behavior [14,15]. daf-11 plays a non-autonomous role in chemotaxis in ciliated ASE neurons [14], and daf-11 regulates dauer-formation and recovery
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