Acid Prohormone Sequence Determines Size, Shape, and Docking of Secretory Vesicles in Atrial Myocytes

• Published in: Circulation research Vol. 89; no. 3; pp. e23 - e29
• Main Authors: Baertschi, Alex J, Monnier, Dominique, Schmidt, Uta, Levitan, Edwin S, Fakan, Stanislas, Roatti, Angela
• Format: Journal Article
• Language: English
• Published: United States American Heart Association, Inc 03.08.2001; Lippincott Williams & Wilkins Ovid Technologies
• Subjects: Animals; Animals, Newborn; Atrial Natriuretic Factor - genetics; Binding Sites - physiology; Biological Transport - physiology; Calcium - metabolism; Cell Membrane - metabolism; Cell Membrane - ultrastructure; Cells, Cultured; Green Fluorescent Proteins; Heart Atria - metabolism; Heart Atria - ultrastructure; Heart Ventricles - cytology; Heart Ventricles - metabolism; Luminescent Proteins - genetics; Mice; Microscopy, Immunoelectron; Microspheres; Mutagenesis, Site-Directed; Myocardium - metabolism; Myocardium - ultrastructure; Particle Size; Protein Precursors - genetics; Protein Sorting Signals - physiology; Rats; Rats, Sprague-Dawley; Recombinant Fusion Proteins - analysis; Recombinant Fusion Proteins - biosynthesis; Recombinant Fusion Proteins - genetics; Secretory Vesicles - metabolism; Secretory Vesicles - ultrastructure; Signal Transduction - physiology; Structure-Activity Relationship; trans-Golgi Network - metabolism
• ISSN: 0009-7330; 1524-4571
• DOI: 10.1161/hh1501.095715

ABSTRACT—How vesicles are born in the trans-Golgi network and reach their docking sites at the plasma membrane is still largely unknown and is investigated in the present study on live, primary cultured atrial cardiomyocytes. Secretory vesicles (n=422) are visualized by expressing fusion proteins of proatrial natriuretic peptide (proANP) and green fluorescent protein. Myocytes expressing fusion proteins with intact proANP display two populations of fluorescent vesicles with apparent diameters of 120 and 175 nm, moving at a top velocity of 0.3 μm/s. The number of docked vesicles is significantly correlated with the number of mobile vesicles (r =0.71, P <0.0005). The deletion of the acidic N-terminal proANP[1-44] or point mutations (glu23,24→gln23,24) change size and shape—but not velocity—of the vesicles, and, strikingly, abolish their docking at the plasma membrane. The shapes thus change from spheres to larger, irregular floppy bags or vesicle trains. Deletion of the C-terminal proANP[45-127], where the ANP and its disulfide bond reside, does not change size, shape, docking, or velocity of the mobile vesicles. The N-terminal acid calcium-binding sequence of proANP is known to cause protein aggregation at the high calcium concentration prevailing in the trans-Golgi network. Therefore, these results indicate that amino acid residues favoring cargo aggregation are critically important in shaping the secretory vesicles and determining their fate—docking or not docking—at the plasma membrane. The full text of this article is available at http://www.circresaha.org.