Evidence for alternative splicing of ecto-ATPase associated with termination of purinergic transmission

Vlajkovic, Srdjan M.; Housley, Gary D.; Greenwood, Denise; Thorne, Peter R.

doi:10.1016/s0169-328x(99)00244-2

Cited by 48 publications

(42 citation statements)

References 34 publications

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“…Alternative splice variants also have been described for the rat NTPDase2 (22). Although it is not known whether alternative messages for the rat ecto-ATPase are translated into functional enzyme, the occurrence of alternative splicing in multiple species suggests it to be a common mechanism for regulation of expression of these enzymes.…”

Section: Discussionmentioning

confidence: 99%

“…Thus, the possibility of the existence of splice variants for the human ecto-ATPase was established (14). A novel rat ecto-ATPase mRNA was identified that con-tains an additional 193-bp exon arising from alternative splicing in the 3Ј-end of the gene (22). Splice variants also were reported for the related ecto-nucleotide pyrophosphatase/phosphodiesterase family (E-NPP), where human phosphodiesterase-I␣ (PD-I␣) and autotaxin likely represent splice variants of the NPP2 gene (23)(24)(25)(26) In this study we delineate the genomic organization of the human ecto-ATPase gene and describe three splice variants of the human NTPDase2 gene encoding proteins of 495, 472, and 450 amino acids, respectively.…”

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confidence: 99%

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Requirement of Cys399 for Processing of the Human Ecto-ATPase (NTPDase2) and Its Implications for Determination of the Activities of Splice Variants of the Enzyme

Mateo

Kreda

Henry

et al. 2003

Journal of Biological Chemistry

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Ecto-ATPase (CD39L1) corresponds to the type 2 enzyme of the ecto-nucleoside triphosphate diphosphohydrolase family (E-NTPDase). We have isolated from human ECV304 cells three cDNAs with high homology with members of the E-NTPDase family that encode predicted proteins of 495, 472, and 450 amino acids. Sequencing of a genomic DNA clone confirmed that these three sequences correspond to splice variants of the human ecto-ATPase (NTPDase2␣, -2␤, and -2␥). Although all three enzyme forms were expressed heterologously to similar levels in Chinese hamster ovary cells clone K-1 (CHO-K1) cells, only the 495-amino acid protein (NTPDase2␣ exhibited ecto-ATPase activity. Immunolocalization studies demonstrated that NTPDase2␣ is fully processed and trafficked to the plasma membrane, whereas the NTPDase2␤ and -2␥ splice variants were retained in not fully glycosylated forms in the endoplasmic reticulum. The potential roles of two highly conserved residues, Cys 399 and Asn 443 , in the activity and cellular trafficking of the ecto-ATPase were examined. Mutation of Cys 399 , which is absent in NTPDase2␤ and -2␥, produced a protein completely devoid of nucleotidase activity, while mutation of Asn 443 to Asp resulted in substantial loss of activity. Neither the Cys 399 nor Asn 443 mutants were fully glycosylated, and both were retained in the endoplasmic reticulum. These results indicate that the lack of ecto-nucleotidase activity exhibited by NTPDase2␤ and -2␥ and the C399S mutant, as well as the large reduction of activity in the N443D mutant are due to alterations in the folding/maturation of these proteins.Extracellular nucleotides modulate many physiological responses through interaction with G protein-coupled metabotropic receptors (P2Y) and ligand-gated ionotropic receptors (P2X) (1-3). Termination of nucleotide-promoted signaling is accomplished by rapid degradation and/or interconversion of extracellular nucleotides by ecto-nucleotidases. Although the primary function of ecto-nucleotidases involves the hydrolysis of extracellular nucleotides, the complete physiological significance of these enzymes is not fully understood. For example, ecto-nucleotidases also have been implicated in physiological phenomena as diverse as cell adhesion (4), purine recycling (5), pain transmission (6), immune function (7), blood hemostasis (8), and others (9, 10).The ecto-nucleoside triphosphate diphosphohydrolases (E-NTPDases) 1 are a family of ecto-nucleotidases, including ecto-ATPases and ecto-ATPDases (apyrases), that hydrolyze nucleoside 5Ј-tri-and diphosphates with isozyme-dependent nucleotide selectivities and biochemical properties. The E-NTPDase family includes membrane-associated enzymes (NTPDase1 to NTPDase4) and soluble enzymes (NTPDase5 and NTPDase6) (see Zimmermann (10) for a review). NTPDase2 (also known as ecto-ATPase or CD39L1) exhibits a strong preference for nucleoside triphosphates (NTPs) over diphosphates (nucleoside diphosphates), hydrolyzing nucleoside diphosphates at 3-5% of the rate of NTPs (10 -14). NTPDase2 contai...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Requirement of Cys399 for Processing of the Human Ecto-ATPase (NTPDase2) and Its Implications for Determination of the Activities of Splice Variants of the Enzyme

Mateo

Kreda

Henry

et al. 2003

Journal of Biological Chemistry

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“…The presence of ectonucleotidase was shown experimentally both in the endolymphatic (Vlajkovic et al, 1998a) and perilymphatic compartments (Vlajkovic et al, 1998b;Vlajkovic et al, 1996). Of six members of the E-NTPDase family, NTPDase 1, NTPDase 2, and NTPDase 3 are likely the key elements for termination of purinergic signaling in the cochlea (Vlajkovic et al, 1999;Vlajkovic et al, 2006). NTPDase 1 degrades both nucleoside triphosphates and diphosphates.…”

Section: Terminationmentioning

confidence: 97%

Purinergic signaling in the inner ear

Lee

Marcus

2008

Hearing Research

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Epithelial cells of the inner ear coordinate their ion transport activity through a number of mechanisms. One important mechanism is the autocrine and paracrine signaling among neighboring cells in the ear via nucleotides, such as adenosine, ATP and UTP. This review summarizes observations on the release, detection and degradation of nucleotides by epithelial cells of the inner ear. Purinergic signaling is thought to be important for endolymph ion homeostasis and for protection from over stimulation.

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“…An additional splice variant also exists for rat NTPDase2 (NTPDase2α and β) [100]. NTPDase2β (545 aa) has an extended cytosolic C-terminus and was found to be localized both at the plasma membrane and at intracellular membranes.…”

Section: General Molecular Propertiesmentioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

854

922

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Ecto-nucleotidases play a pivotal role in purinergic signal transmission. They hydrolyze extracellular nucleotides and thus can control their availability at purinergic P2 receptors. They generate extracellular nucleosides for cellular reuptake and salvage via nucleoside transporters of the plasma membrane. The extracellular adenosine formed acts as an agonist of purinergic P1 receptors. They also can produce and hydrolyze extracellular inorganic pyrophosphate that is of major relevance in the control of bone mineralization. This review discusses and compares four major groups of ectonucleotidases: the ecto-nucleoside triphosphate diphosphohydrolases, ecto-5′-nucleotidase, ecto-nucleotide pyrophosphatase/phosphodiesterases, and alkaline phosphatases. Only recently and based on crystal structures, detailed information regarding the spatial structures and catalytic mechanisms has become available for members of these four ecto-nucleotidase families. This permits detailed predictions of their catalytic mechanisms and a comparison between the individual enzyme groups. The review focuses on the principal biochemical, cell biological, catalytic, and structural properties of the enzymes and provides brief reference to tissue distribution, and physiological and pathophysiological functions.

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Evidence for alternative splicing of ecto-ATPase associated with termination of purinergic transmission

Cited by 48 publications

References 34 publications

Requirement of Cys399 for Processing of the Human Ecto-ATPase (NTPDase2) and Its Implications for Determination of the Activities of Splice Variants of the Enzyme

Requirement of Cys399 for Processing of the Human Ecto-ATPase (NTPDase2) and Its Implications for Determination of the Activities of Splice Variants of the Enzyme

Purinergic signaling in the inner ear

Cellular function and molecular structure of ecto-nucleotidases

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