Divergent Functions of Late ESCRT Components in Giardia lamblia: Insights from Subcellular Distributions and Protein Interactions

Abstract

Giardia lamblia, a human gut pathogen, possesses a minimal ESCRT (Endosomal Sorting Complex Required for Transport) machinery. Paradoxically, there are multiple paralogs of some late-ESCRT components. There are three paralogs for Vps4, GlVps4a, GlVps4b, and GlVps4c, and two for Vps46, GlVps46a, and GlVps46b. This study addressed whether these paralogs discharge overlapping and/or distinct cellular functions by determining the sub-cellular distribution of the paralogs in trophozoites and during encysting. Consistent with the distribution of orthologs in model organisms, most of these components were found to be associated with various cellular membranes, particularly in regions of acute membrane bending. Some of these paralogs are also associated with microtubule structures, such as cytoplasmic axonemes and the median body. Considering their diverse sub-cellular distributions, it is likely that they perform non-overlapping functions within the cell. Furthermore, their redistribution during encystation indicates that they may play a role in the morphological and functional changes accompanying this transition. The study also characterized GlIst1, an ESCRT-III accessory protein that undergoes unique post-translational myristoylation at lysine 43, potentially aiding its membrane recruitment. GlIst1 selectively interacts with GlVps4b through non-canonical MIT-MIM interactions. GlIst1 also exhibits selective interaction with GlVps46b. Such selective interaction of GlIst1 with only specific paralogs of GlVps4 and GlVps46 further underscores the distinct cellular roles of these paralogs. Author SummaryGiardia lamblia, a unicellular protozoan parasite, is the causative agent of giardiasis, a water-transmitted disease affecting millions globally. This disease poses a substantial threat to public health, especially in less developed countries, where clean water and proper sanitation are scarce. The parasite manifests in two morphologically distinct forms, trophozoites and cysts. Transformation between these forms is essential for the organisms survival, spread, and infection processes. Trophozoites, the active and motile form of Giardia, inhabit the small intestine of the host and trigger infections. These trophozoites can transform into cysts through encystation, enabling the parasite to endure harsh external environments and spread between hosts through contaminated water or food sources. The transition between these states necessitates extensive membrane restructuring. Such changes are likely to involve the Endosomal Sorting Complex Required for Transport (ESCRT) machinery, as it has been shown to participate in both prokaryotic and eukaryotic membrane remodeling events. Our research sheds light on the ESCRT machinery in G. lamblia, a crucial membrane-shaping system that may facilitate the transition between trophozoites and cysts. The ESCRT machinery in G. lamblia is distinct from that in yeast and humans, representing one of the most basic ESCRT systems. Our investigation provides valuable information about the intracellular distribution of various late-ESCRT components under different conditions, their potential functions, and their interactions with other late-ESCRT components. These findings may contribute significantly to our understanding of the basic operation of the ESCRT machinery in this parasite.