Identification of a novel phosphorylation site in protein phosphatase inhibitor-1 as a negative regulator of cardiac function.

Abstract

Human and experimental heart failure is characterized by increases in type-1 protein phosphatase activity, which may be partially attributed to inactivation of its endogenous regulator, protein phosphatase inhibitor-1. Inhibitor-1 represents a nodal integrator of two major second messenger pathways, adenosine 3',5'-cyclic monophosphate (cAMP) and calcium, which mediate its phosphorylation at threonine 35 and serine 67, respectively. Here, using recombinant inhibitor-1 wild-type and mutated proteins, we identified a novel phosphorylation site in inhibitor-1, threonine 75. This phosphoamino acid was phosphorylated in vitro by protein kinase Calpha independently and to the same extent as serine 67, the previous protein kinase Calpha-identified site. Generation of specific antibodies for the phosphorylated and dephosphorylated threonine 75 revealed that this site is phosphorylated in rat and dog hearts. Adenoviral-mediated expression of the constitutively phosphorylated threonine 75 inhibitor-1 in isolated myocytes was associated with specific stimulation of type-1 protein phosphatase activity and marked inhibition of the sarcoplasmic calcium pump affinity for calcium, resulting in depressed contractility. Thus, phosphorylation of inhibitor-1 at threonine 75 represents a new mechanism of cardiac contractility regulation, partially through the alteration of sarcoplasmic reticulum calcium transport activity.

• Authors: Patricia Rodriguez, Bryan Mitton, Jason R Waggoner, Evangelia G Kranias
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 281 Issue 50 Pg. 38599-608 (Dec 15 2006) ISSN: 0021-9258 [Print] United States
• PMID: 17046826 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• DNA Primers
• Proteins
• Protein Kinase C-alpha
• PPP1R2C protein, human
• Protein Phosphatase 1
• Calcium

Topics

• Animals
• Base Sequence
• Blotting, Western
• Calcium (metabolism)
• Chromatography, High Pressure Liquid
• DNA Primers
• Electrophoresis, Gel, Two-Dimensional
• Heart (physiology)
• Humans
• Male
• Phosphorylation
• Protein Kinase C-alpha (chemistry, metabolism)
• Protein Phosphatase 1
• Proteins (chemistry, metabolism)
• Rats
• Rats, Sprague-Dawley
• Sarcoplasmic Reticulum (metabolism)
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization