GTPγS Regulation of a 12-Transmembrane Guanylyl Cyclase Is Retained after Mutation to an Adenylyl Cyclase

Roelofs, Jeroen; Loovers, Harriët M.; Haastert, Peter J.M. van

doi:10.1074/jbc.m105154200

Cited by 12 publications

(9 citation statements)

References 61 publications

(68 reference statements)

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“…First, GTP␥S still stimulates GC activity in lysates from G␤-null cells (37), suggesting that at least the prevailing sGC is still sensitive to GTP␥S. In addition, upon mutation of GCA to an adenylyl cyclase, the enzyme is still stimulated by GTP␥S when expressed in cells with a deletion of G␤ or two G␣ subunits (34). Second, both GCA and sGC show normal Michaelis-Menten kinetics, suggesting that GTP does not stimulate these enzymes.…”

Section: Discussionmentioning

confidence: 93%

“…1B), it would be interesting to determine the threedimensional structure of these unusual cyclases. Mutagenesis of GCA has shown that replacing the glutamate and histidine to the corresponding lysine and aspartate converts GCA into a fully active adenylyl cyclase (34). The notion that GCA and especially sGC contain many unusual amino acids at positions that have been shown to provide substrate specificity suggests that conversion of an adenylyl to a guanylyl cyclase can be achieved in different ways.…”

Section: Discussionmentioning

confidence: 99%

“…GCA belongs to the large group of prevailing cyclases that harbors nearly all vertebrate ACs and GCs; GCA is most closely related to the family of 12-transmembrane ACs (17). Sequence alignment of cyclase domains and site-directed mutagenesis suggest that one amino acid may be crucial in determining substrate specificity of cyclases, which is a lysine in nearly all adenylyl cyclases and a glutamate in guanylyl cyclase (6,(31)(32)(33)(34). In GCA this amino acid is glutamate (Glu 440 ) and in sGC an aspartate (Asp 1332 ).…”

Section: Discussionmentioning

confidence: 99%

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Characterization of Two Unusual Guanylyl Cyclases fromDictyostelium

Roelofs

Haastert

2002

Journal of Biological Chemistry

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Guanylyl cyclase A (GCA) and soluble guanylyl cyclase (sGC) encode GCs in Dictyostelium and have a topology similar to 12-transmembrane and soluble adenylyl cyclase, respectively. We demonstrate that all detectable GC activity is lost in a cell line in which both genes have been inactivated. Cell lines with one gene inactivated were used to characterize the other guanylyl cyclase (i.e. GCA in sgc ؊ null cells and sGC in gca ؊ null cells). Despite the different topologies, the enzymes have many properties in common. In vivo, extracellular cAMP activates both enzymes via a G-protein-coupled receptor. In vitro, both enzymes are activated by GTP␥S (K a ‫؍‬ 11 and 8 M for GCA and sGC, respectively). The addition of GTP␥S leads to a 1.5-fold increase of V max and a 3.5-fold increase of the affinity for GTP. Ca 2؉ inhibits both GCA and sGC with K i of about 50 and 200 nM, respectively. Other biochemical properties are very different; GCA is expressed mainly during growth and multicellular development, whereas sGC is expressed mainly during cell aggregation. Folic acid and cAMP activate GCA maximally about 2.5-fold, whereas sGC is activated about 8-fold. Osmotic stress strongly stimulates sGC but has no effect on GCA activity. Finally, GCA is exclusively membrane-bound and is active mainly with Mg 2؉ , whereas sGC is predominantly soluble and more active with Mn 2؉ .cAMP and cGMP are important signaling molecules. In prokaryotes, cAMP regulates gene expression. Cyanobacteria contain high levels of cGMP relative to other bacteria, but their function as intracellular signaling molecules is not well understood (1). In eukaryotes, cAMP and cGMP regulate enzyme activities, channel activity, and gene expression, mainly via cAMP-and cGMP-dependent protein kinase (2, 3). A large and complex family of adenylyl cyclase (AC) 1 and guanylyl cyclase (GC) is responsible for the synthesis of cAMP and cGMP (4, 5).The crystal structure of mammalian AC (6, 7) suggests that the core of the enzyme consist of two cyclase domains that are associated in an antiparallel manner. In metazoan, four cyclase subgroups are recognized (4, 5, 8): 1) the 12-transmembrane adenylyl cyclase is composed of two different cyclase domains and is regulated by G-proteins; 2) The single-transmembrane guanylyl cyclase contains one cyclase domain and functions as a homodimer, and GC activity is stimulated by extracellular peptides; 3) The nitric oxide-sensitive soluble guanylyl cyclase is a complex of two different proteins with one cyclase domain each; 4) The recently discovered soluble adenylyl cyclase (sAC) from rat and human possesses two cyclase domains, which share the highest degree of identity with bacterial adenylyl cyclases (9).In the social amoeba Dictyostelium, cGMP is implicated as one of the second messengers for chemotaxis (10), although its precise role is not known. On the other hand, cAMP can act as both first and second messenger (11, 12). As first messenger, cAMP induces chemotaxis, cAMP signal relay, and gene expression. Dictyostelium possesse...

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Section: Discussionmentioning

confidence: 93%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Characterization of Two Unusual Guanylyl Cyclases fromDictyostelium

Roelofs

Haastert

2002

Journal of Biological Chemistry

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“…It has been proposed, that H504 serves as a hydrogen-bond donor to O 6 of the guanine moiety like the cysteine and serine in mammalian and Paramecium GCs, respectively. This is reasonable, because a change of glutamate-histidine to lysine-aspartate generated a functional AC, i.e., the unusual H504 is in the correct position for substrate-binding (17). In conclusion the GTP-selecting glutamate is functionally conserved among all class IIIaderived GCs investigated to date, but the second selector residue appears to be a variable H-bond donor or may be absent as in the case of Plasmodium GCs.…”

Section: Protozoan Gcs Derived From Class Iiiamentioning

confidence: 91%

Substrate selection by class III adenylyl cyclases and guanylyl cyclases

Linder

2005

IUBMB Life (International Union of Biochemistry and Molecular B

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SummaryThe second messengers cAMP and cGMP are of central importance in signal transduction pathways. To assure pathway specificity adenylyl and guanylyl cyclases are highly selective for their substrates, ATP and GTP, respectively. The universal class III cyclases are equipped with a variety of purine-binding modes, which have been identified by structure determination and mutagenesis. Most selection mechanisms rely on a pair of residues which form hydrogen bonds to N1 and the N 6 -amino or O 6 -keto group of adenine and guanine, respectively. Furthermore, selection is supported by hydrogen bonds involving the peptide backbone and by constraints imposed by hydrophobic side-chains. IUBMB Life, 57: 797 -803, 2005

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“…Ras-interacting protein 3 (RIP3) is required for normal chemotaxis and for receptor activation of both ACA and GC (Lee et al 1999). Small G proteins have been implicated in the regulation of both the soluble and membrane-associated GCs of Dictyostelium (Roelofs et al 2001). In some respects, certain ras-null cells have chemotactic defects that are similar to that of G␣9 Q196L cells, suggesting a potential genetic or biochemical interaction (Wilkins and Insall 2001).…”

Section: Discussionmentioning

confidence: 99%

A Gα-dependent pathway that antagonizes multiple chemoattractant responses that regulate directional cell movement

Brzostowski¹,

Parent²,

Kimmel³

2004

Genes Dev.

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Chemotactic cells, including neutrophils and Dictyostelium discoideum, orient and move directionally in very shallow chemical gradients. As cells polarize, distinct structural and signaling components become spatially constrained to the leading edge or rear of the cell. It has been suggested that complex feedback loops that function downstream of receptor signaling integrate activating and inhibiting pathways to establish cell polarity within such gradients. Much effort has focused on defining activating pathways, whereas inhibitory networks have remained largely unexplored. We have identified a novel signaling function in Dictyostelium involving a G␣ subunit (G␣9) that antagonizes broad chemotactic response. Mechanistically, G␣9 functions rapidly following receptor stimulation to negatively regulate PI3K/PTEN, adenylyl cyclase, and guanylyl cyclase pathways. The coordinated activation of these pathways is required to establish the asymmetric mobilization of actin and myosin that typifies polarity and ultimately directs chemotaxis. Most dramatically, cells lacking G␣9 have extended PI(3,4,5)P 3 , cAMP, and cGMP responses and are hyperpolarized. In contrast, cells expressing constitutively activated G␣9 exhibit a reciprocal phenotype. Their second message pathways are attenuated, and they have lost the ability to suppress lateral pseudopod formation. Potentially, functionally similar G␣-mediated inhibitory signaling may exist in other eukaryotic cells to regulate chemoattractant response.[Keywords: cAMP; cGMP; PI(3,4,5)P 3 ; actin; myosin; Dictyostelium] Supplemental material is available at http://www.genesdev.org.

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GTPγS Regulation of a 12-Transmembrane Guanylyl Cyclase Is Retained after Mutation to an Adenylyl Cyclase

Cited by 12 publications

References 61 publications

Characterization of Two Unusual Guanylyl Cyclases fromDictyostelium

Characterization of Two Unusual Guanylyl Cyclases fromDictyostelium

Substrate selection by class III adenylyl cyclases and guanylyl cyclases

A Gα-dependent pathway that antagonizes multiple chemoattractant responses that regulate directional cell movement

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