The regulation of the cGMP-binding cGMP phosphodiesterase by proteins that are immunologically related to gamma subunit of the photoreceptor cGMP phosphodiesterase

A Lochhead, E Nekrasova, V Y Arshavsky, N J Pyne

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

The cGMP phosphodiesterase from retinal rods (PDE-6) is an alphabetagamma2 heterotetramer. The alpha and beta subunits contain catalytic sites for cGMP hydrolysis, whereas the gamma subunits serve as a protein inhibitor of the enzyme. Visual excitation of photoreceptors enables the activated GTP-bound form of the G-protein transducin to remove the inhibitory action of the gamma subunit, thereby triggering PDE-6 activation. The type 5 phosphodiesterase (PDE-5) isoform shares a number of similar characteristics with PDE-6, including binding of cGMP to noncatalytic sites, the cyclic nucleotide specificity, and inhibitor sensitivities. Although the functional role of PDE-5 remains unclear, it has been shown to be activated by protein kinase A (PKA) (Burns, F., Rodger, I. W. & Pyne, N. J. (1992) Biochem. J. 283, 487-491). Here we report that both the recombinant gamma subunit and a peptide corresponding to amino acids 24-46 in this protein inhibited the activation of PDE-5 by PKA. Furthermore, immunoblotting airway smooth muscle membranes with a specific antibody against amino acids 24-46 of the PDE-6 gamma subunit identified two major immunoreactive small molecular mass proteins of 14 and 18 kDa (p14 and p18). These appear to form a complex with PDE-5, because PDE activity was immunoprecipitated using antibody against the PDE-6 gamma subunit. p14 and p18 were also substrates for phosphorylation by a unidentified kinase that was stimulated by a pertussis toxin-sensitive G-protein. Phosphorylation of p14/p18 in membranes treated with guanine nucleotides correlated with a concurrent reduction in the activation of PDE-5 by PKA. We suggest that p14 and p18 share an epitope common to PDE-6 gamma and that this region may interact with PDE-5 to prevent its activation by PKA.
LanguageEnglish
Pages18397-18403
Number of pages7
JournalJournal of Biological Chemistry
Volume272
Issue number29
DOIs
Publication statusPublished - 18 Jul 1997

Fingerprint

Type 6 Cyclic Nucleotide Phosphodiesterases
Phosphoric Diester Hydrolases
Cyclic AMP-Dependent Protein Kinases
Chemical activation
Phosphorylation
GTP-Binding Proteins
Proteins
Transducin
Type 5 Cyclic Nucleotide Phosphodiesterases
Membranes
Retinal Rod Photoreceptor Cells
Amino Acids
Guanine Nucleotides
Antibodies
Cyclic Nucleotides
Pertussis Toxin
Molecular mass
Enzyme Inhibitors
Guanosine Triphosphate
Burns

Keywords

  • 3',5'-cyclic-GMP phosphodiesterases
  • adenosine triphosphate
  • animals
  • antibodies
  • binding sites
  • cell membrane
  • cells, cultured
  • cyclic AMP-dependent protein kinases
  • cyclic GMP
  • cyclic nucleotide phosphodiesterases, type 5
  • GTP-binding proteins
  • guanylyl imidodiphosphate
  • guinea pigs
  • homeostasis
  • kinetics
  • lung
  • macromolecular substances
  • muscle, smooth
  • pertussis toxin
  • phosphoric diester hydrolases
  • recombinant proteins
  • retinal rod photoreceptor cells
  • trachea
  • virulence factors, bordetella

Cite this

@article{60a215ba326c4206a7a252c2ccf3d9a6,
title = "The regulation of the cGMP-binding cGMP phosphodiesterase by proteins that are immunologically related to gamma subunit of the photoreceptor cGMP phosphodiesterase",
abstract = "The cGMP phosphodiesterase from retinal rods (PDE-6) is an alphabetagamma2 heterotetramer. The alpha and beta subunits contain catalytic sites for cGMP hydrolysis, whereas the gamma subunits serve as a protein inhibitor of the enzyme. Visual excitation of photoreceptors enables the activated GTP-bound form of the G-protein transducin to remove the inhibitory action of the gamma subunit, thereby triggering PDE-6 activation. The type 5 phosphodiesterase (PDE-5) isoform shares a number of similar characteristics with PDE-6, including binding of cGMP to noncatalytic sites, the cyclic nucleotide specificity, and inhibitor sensitivities. Although the functional role of PDE-5 remains unclear, it has been shown to be activated by protein kinase A (PKA) (Burns, F., Rodger, I. W. & Pyne, N. J. (1992) Biochem. J. 283, 487-491). Here we report that both the recombinant gamma subunit and a peptide corresponding to amino acids 24-46 in this protein inhibited the activation of PDE-5 by PKA. Furthermore, immunoblotting airway smooth muscle membranes with a specific antibody against amino acids 24-46 of the PDE-6 gamma subunit identified two major immunoreactive small molecular mass proteins of 14 and 18 kDa (p14 and p18). These appear to form a complex with PDE-5, because PDE activity was immunoprecipitated using antibody against the PDE-6 gamma subunit. p14 and p18 were also substrates for phosphorylation by a unidentified kinase that was stimulated by a pertussis toxin-sensitive G-protein. Phosphorylation of p14/p18 in membranes treated with guanine nucleotides correlated with a concurrent reduction in the activation of PDE-5 by PKA. We suggest that p14 and p18 share an epitope common to PDE-6 gamma and that this region may interact with PDE-5 to prevent its activation by PKA.",
keywords = "3',5'-cyclic-GMP phosphodiesterases, adenosine triphosphate, animals, antibodies, binding sites, cell membrane, cells, cultured, cyclic AMP-dependent protein kinases, cyclic GMP, cyclic nucleotide phosphodiesterases, type 5, GTP-binding proteins, guanylyl imidodiphosphate, guinea pigs, homeostasis, kinetics, lung, macromolecular substances, muscle, smooth, pertussis toxin, phosphoric diester hydrolases, recombinant proteins, retinal rod photoreceptor cells, trachea, virulence factors, bordetella",
author = "A Lochhead and E Nekrasova and Arshavsky, {V Y} and Pyne, {N J}",
year = "1997",
month = "7",
day = "18",
doi = "10.1074/jbc.272.29.18397",
language = "English",
volume = "272",
pages = "18397--18403",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
number = "29",

}

The regulation of the cGMP-binding cGMP phosphodiesterase by proteins that are immunologically related to gamma subunit of the photoreceptor cGMP phosphodiesterase. / Lochhead, A; Nekrasova, E; Arshavsky, V Y; Pyne, N J.

In: Journal of Biological Chemistry, Vol. 272, No. 29, 18.07.1997, p. 18397-18403.

Research output: Contribution to journalArticle

TY - JOUR

T1 - The regulation of the cGMP-binding cGMP phosphodiesterase by proteins that are immunologically related to gamma subunit of the photoreceptor cGMP phosphodiesterase

AU - Lochhead, A

AU - Nekrasova, E

AU - Arshavsky, V Y

AU - Pyne, N J

PY - 1997/7/18

Y1 - 1997/7/18

N2 - The cGMP phosphodiesterase from retinal rods (PDE-6) is an alphabetagamma2 heterotetramer. The alpha and beta subunits contain catalytic sites for cGMP hydrolysis, whereas the gamma subunits serve as a protein inhibitor of the enzyme. Visual excitation of photoreceptors enables the activated GTP-bound form of the G-protein transducin to remove the inhibitory action of the gamma subunit, thereby triggering PDE-6 activation. The type 5 phosphodiesterase (PDE-5) isoform shares a number of similar characteristics with PDE-6, including binding of cGMP to noncatalytic sites, the cyclic nucleotide specificity, and inhibitor sensitivities. Although the functional role of PDE-5 remains unclear, it has been shown to be activated by protein kinase A (PKA) (Burns, F., Rodger, I. W. & Pyne, N. J. (1992) Biochem. J. 283, 487-491). Here we report that both the recombinant gamma subunit and a peptide corresponding to amino acids 24-46 in this protein inhibited the activation of PDE-5 by PKA. Furthermore, immunoblotting airway smooth muscle membranes with a specific antibody against amino acids 24-46 of the PDE-6 gamma subunit identified two major immunoreactive small molecular mass proteins of 14 and 18 kDa (p14 and p18). These appear to form a complex with PDE-5, because PDE activity was immunoprecipitated using antibody against the PDE-6 gamma subunit. p14 and p18 were also substrates for phosphorylation by a unidentified kinase that was stimulated by a pertussis toxin-sensitive G-protein. Phosphorylation of p14/p18 in membranes treated with guanine nucleotides correlated with a concurrent reduction in the activation of PDE-5 by PKA. We suggest that p14 and p18 share an epitope common to PDE-6 gamma and that this region may interact with PDE-5 to prevent its activation by PKA.

AB - The cGMP phosphodiesterase from retinal rods (PDE-6) is an alphabetagamma2 heterotetramer. The alpha and beta subunits contain catalytic sites for cGMP hydrolysis, whereas the gamma subunits serve as a protein inhibitor of the enzyme. Visual excitation of photoreceptors enables the activated GTP-bound form of the G-protein transducin to remove the inhibitory action of the gamma subunit, thereby triggering PDE-6 activation. The type 5 phosphodiesterase (PDE-5) isoform shares a number of similar characteristics with PDE-6, including binding of cGMP to noncatalytic sites, the cyclic nucleotide specificity, and inhibitor sensitivities. Although the functional role of PDE-5 remains unclear, it has been shown to be activated by protein kinase A (PKA) (Burns, F., Rodger, I. W. & Pyne, N. J. (1992) Biochem. J. 283, 487-491). Here we report that both the recombinant gamma subunit and a peptide corresponding to amino acids 24-46 in this protein inhibited the activation of PDE-5 by PKA. Furthermore, immunoblotting airway smooth muscle membranes with a specific antibody against amino acids 24-46 of the PDE-6 gamma subunit identified two major immunoreactive small molecular mass proteins of 14 and 18 kDa (p14 and p18). These appear to form a complex with PDE-5, because PDE activity was immunoprecipitated using antibody against the PDE-6 gamma subunit. p14 and p18 were also substrates for phosphorylation by a unidentified kinase that was stimulated by a pertussis toxin-sensitive G-protein. Phosphorylation of p14/p18 in membranes treated with guanine nucleotides correlated with a concurrent reduction in the activation of PDE-5 by PKA. We suggest that p14 and p18 share an epitope common to PDE-6 gamma and that this region may interact with PDE-5 to prevent its activation by PKA.

KW - 3',5'-cyclic-GMP phosphodiesterases

KW - adenosine triphosphate

KW - animals

KW - antibodies

KW - binding sites

KW - cell membrane

KW - cells, cultured

KW - cyclic AMP-dependent protein kinases

KW - cyclic GMP

KW - cyclic nucleotide phosphodiesterases, type 5

KW - GTP-binding proteins

KW - guanylyl imidodiphosphate

KW - guinea pigs

KW - homeostasis

KW - kinetics

KW - lung

KW - macromolecular substances

KW - muscle, smooth

KW - pertussis toxin

KW - phosphoric diester hydrolases

KW - recombinant proteins

KW - retinal rod photoreceptor cells

KW - trachea

KW - virulence factors, bordetella

U2 - 10.1074/jbc.272.29.18397

DO - 10.1074/jbc.272.29.18397

M3 - Article

VL - 272

SP - 18397

EP - 18403

JO - Journal of Biological Chemistry

T2 - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 29

ER -