Serine/threonine protein kinases have been shown to reversibly regulate cell-cell communication through gap junctions formed by connexin 43 (Cx43) (Godwin et al.7 1993). The Cx43 protein expression levels are unclear and what role phosphorylation plays in the regulation of signal transduction through these gap junctions during heart formation. Reportedly, mutations which replace potentially phosphorylatable serines within the Cx43 protein are associated with heart malformations (Britz-Cunningham et al.. 1995), highlighting the importance of determining the levels of Cx43 and the activities of PKA and PKC during heart development.
Western analysis performed on hearts from developmentally significant time points revealed principally high molecular weight forms of Cx43. Metabolic labeling procedures confirmed that these forms of Cx43 were phosphorylated, suggesting that the development of a functional, four-chambered heart requires gap junctional communication provided by Cx43 during embryogenesis which is differentially regulated by phosphorylation.
PKA expression levels were relatively constant over the developmental time frame except for an increase at 10.5 dpc and minor fluctuations during the completion of heart chamber formation. Thereafter, a steady increase in the amount of enzyme was noted in all time points examined through adulthood.
PKCs was detected below adult concentrations at all time points at levels that were relatively constant except for increases at 10.5 and 15.5 dpc. Protein quantity increased steadily from 16.5 dpc to adulthood, starting from a lower level than that detected at 15.5 dpc and rising to adult concentrations.
PKC5 had the most variable expression of the protein kinases examined, with rapid fluctuations occurring during heart development. Again, detected levels peaked at 15.5 dpc and increasing protein quantities mimicked the rise in PKCs to adult levels starting at 16.5 dpc, except that at birth protein expression declined.
To further understand the importance of these varying levels of expression during organogenesis, in vitro phosphorylation assays for determination of protein kinase activity were completed. PKA activity peaked at days corresponding to the completion of hearttube looping, chamber formation and preparation for birth. Combined nPKC activity levels were relatively high in the early developmental period; thereafter activity continually decreased.
William H. Fletcher
David A. Hessinger
Michael A. Kirby
Cecilia W. Lo
Paul J. McMillan
Kerby C. Oberg
Doctor of Philosophy (Medical Science)
Year Degree Awarded
Date (Title Page)
Library of Congress/MESH Subject Headings
Heart -- anatomy & histology; Heart -- growth & development; Connexin 43; Phosphorylation
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This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.
Duncan, John C., "Regulation of Connexin43 by Phosphorylation in Heart Development" (1998). Loma Linda University Electronic Theses, Dissertations & Projects. 1033.
Loma Linda University Electronic Theses and Dissertations
Loma Linda University. Del E. Webb Memorial Library. University Archives