Date of Award
Doctor of Philosophy
Marvin H. Stromer
This dissertation is divided into two principal parts. The purpose of the first part was to investigate the effects of cations and temperature on the critical concentration of desmin required for filament assembly by using tow independent assays, namely, turbidity and centrifugation. Assembly was done in the presence of 2 mM Ca[superscript]2+, 2 mM Mg[superscript]2+ or 150 mM Na[superscript]+ at 2, 22 and 37°C. Similar values for critical concentration were obtained by both assays. As temperature increased, critical concentration decreased for each cation. Critical concentration was lowest in the presence of Ca[superscript]2+ at 2, 22 and 37°C, but was highest in the presence of 150 mM Na[superscript]+ at 2°C. Negative staining showed that supernatants from the centrifugation assays contained protofilaments, protofibrils and short particles (1 [mu]m) with an average diameter of 10 nm. Thermodynamic analysis indicated that hydrophobic interactions were dominant during desmin assembly but that ionic interactions might also be involved;The objective of the second part was to investigate in vivo changes, especially in cytoplasmic dense bodies (CDBs), in developing smooth muscle cells by transmission electron microscopy and immunoelectron microscopy. The developmental events observed in 6-day embryonic through 7-day post-hatch chicken gizzards were sequentially classified into 7 stages, primarily based on the appearance and the growth of contractile and cytoskeletal elements. These stages are: myoblast proliferation; dense body appearance, thick filament appearance, dense body growth; muscle cell replication; intermediate filament bundle (IFB) appearance; and appearance of adult type cells. These events clearly indicate that smooth muscle myogenesis in vivo is a sequential process in which contractile elements (actin and myosin filaments) and dense bodies are assembled before the axial IFB, a major cytoskeletal element;SDS-PAGE and Western blots together with colloidal gold labeled antibodies and electron microscopy showed that, between 10 days embryonic age and 7-days post-hatch, amounts of myosin heavy chain, [alpha]-actinin, desmin and actin all increase. On the basis of structural changes in the CDB and immunoelectron microscopy labeling, we propose that CDB assembly includes three steps: (1) alignment of actin and intermediate filaments; (2) cross-linking of major longitudinal filaments, probably by [alpha]-actinin; and (3) addition of components that increase the electron density of the CDB.
Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/
Rong-Ghi Robert Chou
Chou, Rong-Ghi Robert, "In vitro desmin assembly and in vivo changes in developing smooth muscle " (1991). Retrospective Theses and Dissertations. 10022.