The regulation of actin polymerization during amoeboid chemotaxis in Dictyostelium discoideum

Abstract

The studies reported in this thesis describe responses of Dictyostelium discoideum amoebae stimulated with chemoattractants. Changes in cell shape and pseudopod extension and their relationship to actin polymerization are described in Chapter I and its Addendum. An intracellular activity that nucleates actin filament formation and is regulated by chemotactic stimulation was discovered in cell lysates of amoebae that had been stimulated prior to lysis. The description of this activity and the discovery of an inhibitor of nucleation activity, also regulated by chemotactic stimulation, are presented in Chapter II. An investigation of the actin nucleation and polymerization responses in a number of different contexts in which these cells chemotax is described in Chapter III. A study of the transduction pathways involved in control of the actin nucleation and actin polymerization responses to chemoattractant stimulation is contained in Chapter IV.;The success of studies on cell physiology requires careful attention to the methods used to handle cells and gather data. This is particularly true in the case of Dictyostelium amoebae because a number of strains are in use, the growth conditions and composition of nutrient media varies between laboratories, and the buffers used for incubations of living cells vary as well. In order to obtain reproducible results during the studies reported here, these factors were standardized and are presented in exhaustive detail in the Materials and Methods section of Chapter I with attention given to the differing requirements of various strains of D. discoideum. Close reading of the Materials and Methods section of Chapter I is not necessary for an understanding of the remainder and is of interest principally to those who wish to reproduce our results or compare our methods with those of others.;Additional findings that grew out of the work reported in Chapter I and which confirm our interpretations of that work are reported briefly in the Addendum to Chapter I. The remaining text presents the background, findings and interpretations of our analyses.