Toward a Synthetic Yeast Endosymbiont with a Minimal Genome

Abstract

Based on the endosymbiotic theory, one of the key events that occurred during mitochondrial evolution was an extensive loss of nonessential genes from the protomitochondrial endosymbiont genome and transfer of some of the essential endosymbiont genes to the host nucleus. We have developed an approach to recapitulate various aspects of endosymbiont genome minimization using a synthetic system consisting of Escherichia coli endosymbionts within host yeast cells. As a first step, we identified a number of E. coli auxotrophs of central metabolites that can form viable endosymbionts within yeast cells. These studies provide a platform to identify nonessential biosynthetic pathways that can be deleted in the E. coli endosymbionts to investigate the evolutionary adaptations in the host and endosymbiont during the evolution of mitochondria.

Figure 1:. Strategy to test permissive E. coli auxotrophs that could establish endosymbiosis with S. cerevisiaecox2–60.

X- key central metabolite provided by the yeast cytosol, M-mitochondria, N-nucleus, ER-endoplasmic reticulum, G-Golgi apparatus, V-vacuole.

Figure 2:. Characterization of Saccharomyces cerevisiae cox2–60 - E. coli DH10B ΔthiC:gfp ΔnadA ΔmetA ΔilvD, ΔargA, ΔproA – pAM136 chimeras

(A) and (B), Growth observed for chimeras on Selection Medium I and Selection Medium II respectively, thereby demonstrating a respiration competent phenotype. (C) and (D), Detection of NB97-specific MATa mating type by PCR analysis of total DNA samples isolated from chimeras shown in Figure 2A and 2B respectively. Std indicates DNA molecular weight standards (E) and (F) Detection of gfp gene by PCR analysis of DNAs isolated from chimeras shown in Figure 2A and 2B respectively. (G) Detection of GFP expressing E. coli endosymbionts within Saccharomyces cerevisiae cox2–60. No GFP signals are detected in control Saccharomyces cerevisiae cox2–60 cells. Scale bars represent10 μm.