Self-recognition in social amoebae is mediated by allelic pairs of tiger genes

Abstract

Free-living cells of the social amoebae Dictyostelium discoideum can aggregate and develop into multicellular fruiting bodies in which many die altruistically as they become stalk cells that support the surviving spores. Dictyostelium cells exhibit kin discrimination--a potential defense against cheaters, which sporulate without contributing to the stalk. Kin discrimination depends on strain relatedness, and the polymorphic genes tgrB1 and tgrC1 are potential components of that mechanism. Here, we demonstrate a direct role for these genes in kin discrimination. We show that a matching pair of tgrB1 and tgrC1 alleles is necessary and sufficient for attractive self-recognition, which is mediated by differential cell-cell adhesion. We propose that TgrB1 and TgrC1 proteins mediate this adhesion through direct binding. This system is a genetically tractable ancient model of eukaryotic self-recognition.

Fig. 1

Cells (all in the AX4 background) labeled with GFP (green) or RFP (red) were grown separately, mixed at equal proportions and allowed to develop together (A–E). Multicellular structures were photographed with epifluorescence microscopy during streaming aggregation (8–12 hours, main pictures) and at the finger stage (14–18 hours, inserts). Bar = 0.5mm. The cartoons are models of interactions between cells. Cells are illustrated as elongated amoebae. TgrB1 and TgrC1 are illustrated as membrane proteins; the shading indicates their origins. Interactions between matching pairs are highlighted. (A) The parental strain (AX4, red) mixed with a tgrB1-null tgrC1-null strain (green). (B) AX4 (red) mixed with a strain in which tgrB1-tgrC1 were replaced with matching AX4 alleles. (C,D) Strains (green) in which tgrB1-tgrC1 were replaced with alleles from wild strains QS4 (C) or QS38 (D), mixed with AX4 (red). (E) Double gene-replacement cells carrying tgrB1-tgrC1 from QS4 (red) mixed with double gene-replacement cells carrying tgrB1-tgrC1 from QS38 (green). Pure populations were also developed into fruiting bodies (48h; F – bar = 1mm, G–I – bar = 0.5mm). (F) tgrB1-null tgrC1-null cells. (G–I) Double gene-replacement cells carrying tgrB1-tgrC1 alleles from AX4 (G), QS4 (H), or QS38 (I). (J) Sporulation efficiencies of these and two additional double gene-replacement strains harboring tgrB1-tgrC1 from strains QS31 or QS45 as indicated under the x-axis. The graph shows the fraction (%) of cells that became spores. The means and standard errors of 5–6 replicates are given.

Fig. 2

We labeled the wild strains QS4 (A–C) and QS31 (D–F) with GFP (green text) and the double gene-replacement strains (in AX4 background) with RFP (red text): tgrB1-tgrC1 were from AX4 (A,D), QS4 (B,F) or QS31 (C,E). We grew the cells separately, mixed them at equal proportions as indicated and allowed them to develop together. We photographed the structures as in Fig. 1. Bar = 0.5mm. Cartoons are as in Fig. 1. The shadings indicate genetic differences between cells and the respective origins of the alleles.

Fig. 3

We developed pure populations of RFP (red) and GFP (green) labeled cells, all in the AX4 background, disaggregated and mixed them at equal proportions, allowed them to adhere in suspension and photographed with confocal fluorescence microscopy (bar = 0.1mm). (A) AX4 cells (red) mixed with a strain in which tgrB1-tgrC1 were replaced with matching alleles from AX4 (green). (B) AX4 (red) mixed with a strain in which tgrB1-tgrC1 were replaced with matching alleles from QS4 (green). (C) Strains (both red and green) in which tgrB1-tgrC1 were replaced with matching alleles from QS4. Cartoons are as in Fig. 1.

Fig. 4

We made merodiploid strains by introducing extra pairs of matching alleles into AX4: tgrB1-tgrC1 from QS4 (A–C), QS31 (D–F) and AX4 (G–I). We labeled these cells with RFP (red) and mixed them with GFP-labeled haploid cells: AX4 (A,D,G), or double gene replacement strains with tgrB1-tgrC1 from QS4 (B,E,H), or QS31 (C,F,I). Cell growth, mixing, photography and cartoons are as in Fig. 1. Bar = 0.5mm.