Differential expression of a family of putative adenylate/guanylate cyclase genes in Trypanosoma brucei

Alexandre, S; Paindavoine, Pascale; Tebabi, Patricia; Pays, Annette; Halleux, S de; Steinert, Maurice; Pays, Étienne

doi:10.1016/0166-6851(90)90152-c

Cited by 79 publications

(59 citation statements)

References 28 publications

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“…It has been pointed out that the structures of these adenylate cyclases are similar to that of the transmembrane receptor protein (38). It has also been noted that in T. brucei, adenylate cyclase pierces the membrane twice and that the region between these two transmembrane spans is supposed to function as a sensor of a certain exogenous signal (2). Although it is unknown whether the transmembrane region near the N terminus is processed, the domain lying between the two transmembrane spans seems to be exposed to the periplasmic space or thylakoid lumen.…”

Section: Resultsmentioning

confidence: 97%

“…This adenylate cyclase is similar to those found in the lower eukaryotes Dictyostelium and Trypanosoma. In these organisms, adenylate cyclase is considered to function as a receptor or a sensor (2,38). Recently, the adenylate cyclase of Dictyostelium that is expressed in spores was shown to be involved in sensing extracellular osmolarity (48).…”

Section: Discussionmentioning

confidence: 99%

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Isolation and characterization of multiple adenylate cyclase genes from the cyanobacterium Anabaena sp. strain PCC 7120

Katayama

Ohmori

1997

J Bacteriol

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Adenylate cyclase genes, designated cyaA, cyaB1, cyaB2, cyaC, and cyaD, were isolated from the filamentous cyanobacterium Anabaena sp. strain PCC 7120 by complementation of a strain of Escherichia coli defective for the presence of cya. These genes encoded polypeptides consisting of 735, 859, 860, 1,155, and 546 amino acid residues, respectively. Deduced amino acid sequences of the regions near the C-terminal ends of these cya genes were similar to those of catalytic domains of eukaryotic adenylate cyclases. The remaining part of each cya gene towards its N-terminal end showed a characteristic structure. CyaA had two putative membrane-spanning regions. Both CyaB1 and CyaB2 had regions that were very similar to the cyclic GMP (cGMP)-binding domain of cGMP-stimulated cGMP phosphodiesterase. CyaC consisted of four distinct domains forming sequentially from the N terminus: a response regulator-like domain, a histidine kinase-like domain, a response regulatorlike domain, and the catalytic domain of adenylate cyclase. CyaD contained the forkhead-associated domain in its N-terminal region. Expression of these genes was examined by reverse transcription-PCR. The transcript of cyaC was shown to be predominant in this cyanobacterium. The cellular cyclic AMP level in the disruptant of the cyaC mutant was much lower than that in the wild type.Cyclic AMP (cAMP) regulates various biological activities in both prokaryotes and eukaryotes. In prokaryotes, cAMP and cAMP receptor protein regulate gene expression (7). In eukaryotes, cAMP regulates enzyme activity (11), channel activity (42), and gene expression (34) via the cAMP-dependent protein kinase. In cyanobacteria, it has been reported that cellular cAMP content is affected by growth conditions (17,22), that cAMP concentration changes rapidly in response to light-off and light-on signals (37) or to a pH shift (36), and that cAMP interferes with the pattern formation of heterocysts (43). However, it is still unknown whether cAMP functions as an intracellular signal molecule in cyanobacteria.Recently, we isolated adenylate cyclase (cya) genes from the filamentous cyanobacteria Anabaena cylindrica (26) and Spirulina platensis (52). Regions near the C-terminal ends of the adenylate cyclases of both cyanobacteria showed significant structural similarity to the catalytic domain of the adenylate cyclase of eukaryotes but showed no homology to that of prokaryotes such as Escherichia coli. Furthermore, cyanobacterial adenylate cyclases appeared to be membrane oriented.To study the physiological functions of these adenylate cyclases, disruption or overexpression of cya genes is indispensable. Unfortunately, both A. cylindrica and S. platensis are not transformable, so they are not suitable for this purpose. In this study, we isolated cya genes from a transformable cyanobacterium, Anabaena sp. strain PCC 7120, and constructed mutants defective in cya genes. Some physiological properties of these mutants were determined. MATERIALS AND METHODSStrains and growth conditions. Anabaena sp. s...

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Section: Resultsmentioning

confidence: 97%

Section: Discussionmentioning

confidence: 99%

Isolation and characterization of multiple adenylate cyclase genes from the cyanobacterium Anabaena sp. strain PCC 7120

Katayama

Ohmori

1997

J Bacteriol

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“…More than 30 different adenylyl cyclases and guanylyl cyclases share the common homologous cytoplasmic domains, designated as the cyclase domains (1). For the functional enzyme, the cyclase domains can work as the homo-oligomer, such as adenylyl cyclases from yeast, Trypanosome, Dictyostelium (ACG), Leishmania, Brevibacterium, and Rhizobium (23)(24)(25)(26)(27)(28)(29)(30). The cyclase domains can also exist as an obligatory heterodimer for enzymatic activity such as the C 1 and C 2 domains from type I to type IX adenylyl cyclases, Drosophila rutabaga, and Dictyostelium ACA, and the ␣ and ␤ subunits of soluble guanylyl cyclases (22,23,(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46).…”

Section: Isolation and Analysis Of Loss Of Function Ic 1 Iic 2 Mutants-mentioning

confidence: 99%

“…Second are the integral membrane proteins that have one transmembrane helix. This class is found in the parasites, Trypanosoma brucei (Tb ESAG, expression site-associated gene), and Leishmania donovani (Ld-RAC, receptoradenylyl cyclase), and in the slime mold (Dictyostelium discoideum, Dd ACG) (23)(24)(25). Third are the peripheral membrane proteins from yeast (Schizosaccharomyces pombe, Saccharomyces cerevisiae, and Saccharomyces klyuyveri) (26 -28).…”

mentioning

confidence: 99%

The Conserved Asparagine and Arginine Are Essential for Catalysis of Mammalian Adenylyl Cyclase

Yan

Huang

Shaw

et al. 1997

Journal of Biological Chemistry

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Mammalian adenylyl cyclases have two homologous cytoplasmic domains (C 1 and C 2 ), and both domains are required for the high enzymatic activity. Mutational and genetic analyses of type I and soluble adenylyl cyclases suggest that the C 2 domain is catalytically active and the C 1 domain is not; the role of the C 1 domain is to promote the catalytic activity of the C 2 domain. Two amino acid residues, Asn-1025 and Arg-1029 of type II adenylyl cyclase, are conserved among the C 2 domains, but not among the C 1 domains, of adenylyl cyclases with 12 putative transmembrane helices. Mutations at each amino acid residue alone result in a 30 -100-fold reduction in K cat of adenylyl cyclase. However, the same mutations do not affect the K m for ATP, the half-maximal concentration (EC 50 ) for the C 2 domain of type II adenylyl cyclase to associate with the C 1 domain of type I adenylyl cyclase and achieve maximal enzyme activity, or the EC 50 for forskolin to maximally activate enzyme activity with or without G s␣ . This indicates that the mutations at these two residues do not cause gross structural alteration. Thus, these two conserved amino acid residues appear to be crucial for catalysis, and their absence from the C 1 domains may account for its lack of catalytic activity. Mutations at both amino acid residues together result in a 3,000-fold reduction in K cat of adenylyl cyclase, suggesting that these two residues have additive effects in catalysis. A second site suppressor of the Asn-1025 to Ser mutant protein has been isolated. This suppressor has 17-fold higher activity than the mutant and has a Pro-1015 to Ser mutation.The activity of mammalian adenylyl cyclase, the enzyme that converts ATP to cAMP, is the key step in modulating intracellular cAMP concentration in response to extracellular stimulation by hormones, neurotransmitters, and odorants. Nine isoforms of mammalian adenylyl cyclases have been cloned to date, and they belong to a rapid expanding cyclase family that includes class III adenylyl cyclases 1 and guanylyl cyclases (1).All nine isoforms have a similar structure, including two intensely hydrophobic domains (M 1 and M 2 ) and two 40-kDa cytoplasmic domains (C 1 and C 2 ). The two cytoplasmic domains contain sequences (C 1a and C 2a ) that are homologous to each other and to class III adenylyl cyclases and guanylyl cyclases. Each isoform of adenylyl cyclase not only has distinct patterns of tissue expression but also has unique responses to extracellular and intracellular stimuli (1-3). In common, each adenylyl cyclase can be activated by the G protein 2 ␣ subunit, designated as G s␣ , and each can be inhibited by certain adenosine analogues (P-site inhibitors). However, there are several subtype-specific regulators, including forskolin, G protein ␤␥ subunits, the ␣ subunit of G i , G o , and G z , Ca 2ϩ ion, Ca 2ϩ -calmodulin, Ca 2ϩ -calcineurin, cAMP-dependent protein kinase, and PKC (for review, see Refs. 1-3). Certain ones of these regulators (e.g. ␤␥) can be either stimulatory or inhibitory...

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“…cAMP is involved in the regulation of differentiation of bloodstream form trypanosomes from the proliferative "long slender" forms to the insect-preadapted, nonproliferative "short stumpy" forms (26). Several gene families for adenylyl cyclases have been identified in T. brucei (24,27,28). 2 Even less is known about trypanosomal phosphodiesterases.…”

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confidence: 99%

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

Zoraghi¹,

Kunz

Gong³

et al. 2001

Journal of Biological Chemistry

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This study reports the identification and characterization of a cAMP-specific phosphodiesterase from the parasitic hemoflagellate Trypanosoma brucei. TbPDE2A is a class I phosphodiesterase. Its catalytic domain exhibits 30 -40% sequence identity with those of all 11 mammalian phosphodiesterase (PDE) families, as well as with PDE2 from Saccharomyces cerevisiae, dunce from Drosophila melanogaster, and regA from Dictyostelium discoideum. The overall structure of TbPDE2A resembles that of human PDE11A in that its N-terminal region contains a single GAF domain. This domain is very similar to those of the mammalian PDE2, -5, -6, -10, and -11, where it constitutes a potential cGMP binding site. TbPDE2A can be expressed in S. cerevisiae, and it complements an S. cerevisiae PDE deletion strain. Recombinant TbPDE2A is specific for cAMP, with a K m of ϳ2 M. It is entirely resistant to the nonselective PDE inhibitor 3-isobutyl-1-methylxanthine, but it is sensitive to trequinsin, dipyridamole, sildenafil, and ethaverine with IC 50 values of 5.4, 5.9, 9.4, and 14.2 M, respectively. All four compounds inhibit proliferation of bloodstream form trypanosomes in culture, indicating that TbPDE2A is an essential enzyme.

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Differential expression of a family of putative adenylate/guanylate cyclase genes in Trypanosoma brucei

Cited by 79 publications

References 28 publications

Isolation and characterization of multiple adenylate cyclase genes from the cyanobacterium Anabaena sp. strain PCC 7120

Isolation and characterization of multiple adenylate cyclase genes from the cyanobacterium Anabaena sp. strain PCC 7120

The Conserved Asparagine and Arginine Are Essential for Catalysis of Mammalian Adenylyl Cyclase

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

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