Functional analysis of Paramecium genes responsiveness to Holospora infection

Abstract

Paramecium caudatum is a single-celled ciliate that sweeps food into its oral groove to acquire nutrients. The cells become infected through this feeding apparatus by Holospora undulata, an obligate endosymbiont that occupies the micronucleus of P. caudatum in two morphologically distinct forms, the infectious and reproductive forms. Once the P. caudatum cells begin to starve, the reproductive forms of the parasitic bacteria consume most of the food in the micronuclei and differentiate into infectious forms. The bacteria eventually break out of the cell and seek other hosts to infect. To better understand the mechanism by which H. undulata successfully infiltrates their host’s defense systems, P. caudatum strains were infected and varying levels of susceptibility to infection were identified. Whole genome RNA sequencing found that certain proteins are significantly upregulated upon uptake of the H. undulata. By cloning the significantly up-regulated genes into E. coli, feeding this silencing media to naïve stocks of P. caudatum, and analyzing the infection phenotype of each, we hoped to characterize the genes that function in the infiltration of H. undulata. Varying levels of infection prevalence were seen amongst the gene knockdowns and across replicates of each infection. However, five knockdowns show significant resistance compared to their respective controls, revealing a possible involvement of 5 genes in H. undulata susceptibility. After finding which genes are capable of inducing resistance, we plan to identify the associated proteins and track the gene’s evolutionary divergence in P. caudatum strains with varying infectability.