The regulative capacity of prespore amoebae as demonstrated by fluorescence-activated cell sorting and green fluorescent protein

Abstract

The ability of prespore Dictyostelium discoideum amoebae to undergo redifferentiation so as to reestablish normal spore/stalk proportioning has been demonstrated in various ways over the years, beginning with the classic microdissection work of K. Raper. The discovery of anterior-like cells in the slug posterior, however, cast doubt on that ability, and more recent experiments using a cell-specific toxin suggested that prespore redifferentiation may not in fact occur. To reexamine this question, we performed fluorescence-activated cell sorting (FACS) upon amoebae expressing a mutated green fluorescent protein gene (S65T-GFP) under the control of a prespore-specific (PsA) promoter. FACS produced prespore cell populations with purifies, measured by GFP expression, as high as 99.5%. Sorted GFP+ cells were developmentally competent and produced normally proportioned fruits, indistinguishable from those of 'sham-sorted' (permissively gated, mixed GFP+ and GFP-) amoebae. This result confirms the developmental totipotency of prespore amoebae.