A microscopic, transcriptomic, and phylogenetic investigation of the contractile vacuole of Reclinomonas americana (Jakobida, Discoba)
Creators: More, Kiran J
Publisher: name: University of Alberta Library
Year: 2023

<p>The origin and timing of characteristic eukaryotic organelles is one of the key questions in the evolution of life on Earth. For the most part, widespread eukaryotic organelles (e.g., Golgi, mitochondria) have been inferred to be present in the last eukaryotic common ancestor (LECA) by examining the distribution of the organelles and their marker genes across the diversity of eukaryotes (the vast majority of which are unicellular protists). Other common organelles, such as the contractile vacuole (CV), are much more poorly understood. The CV is an osmoregulatory organelle that is found across the diversity of eukaryotes, most prominently in protists from freshwater ecosystems. As freshwater is hypoosmotic to the cell, there is continual movement of water across the plasma membrane, which cells must counteract to prevent lysis. The CV gathers and expels this excess water. Due to its absence from many of the model organisms typically used for cell biological study, there is a paucity of molecular data associated with this organelle. The limited data available from four distantly related protist model organisms show a pattern of repeat involvement of specific proteins known from other organelles. The considerable morphological diversity and patchy distribution (as CVs are generally absent from closely related marine and parasitic species) of these organelles makes their homology uncertain. Are CVs a single, homologous organelle with an origin in the LECA, or have there been multiple more recent evolutionary origins of an osmoregulatory organelle? </p>

<pre><code>Through a combination of microscopy, transcriptomics, and phylogenetics I investigated this question within the eukaryotic supergroup Discoba using the CV model organism Trypanosoma cruzi as a reference point. I microscopically characterized the CV response of the non-model organism Reclinomonas americana to low, control, and high osmolarities. These treatments were then used to induce CV activity, and RNA was harvested for differential expression analysis. Transcripts of the membrane trafficking system that increased as osmolarity decreased were considered putatively CV-associated. The associated proteins were then used as a proxy to investigate the evolutionary history of the organelle. If CVs have a single origin, I would expect the CV-specific paralogs to form clades in phylogenies to the exclusion of versions found elsewhere in the cell. This was never observed. Though several common CV-associated proteins were detected as differentially expressed in R. americana, phylogenetics indicated that these were the result of lineage-specific expansions. These results show no strong support for a single origin of CVs within Discoba, or across eukaryotes. Rather, they support either convergent or parallel evolution of the CVs examined. The reoccurrence of the same proteins in the CVs of diverse eukaryotes raises the question of whether these CVs evolved from the same non-CV organelle with latent osmoregulatory capacity.
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