Most cited article - PubMed ID 31964712
Diversification of CORVET tethers facilitates transport complexity in Tetrahymena thermophila
Genetic variation is the major mechanism behind adaptation and evolutionary change. As most proteins operate through interactions with other proteins, changes in protein complex composition and subunit sequence provide potentially new functions. Comparative genomics can reveal expansions, losses and sequence divergence within protein-coding genes, but in silico analysis cannot detect subunit substitutions or replacements of entire protein complexes. Insights into these fundamental evolutionary processes require broad and extensive comparative analyses, from both in silico and experimental evidence. Here, we combine data from both approaches and consider the gamut of possible protein complex compositional changes that arise during evolution, citing examples of complete conservation to partial and total replacement by functional analogues. We focus in part on complexes in trypanosomes as they represent one of the better studied non-animal/non-fungal lineages, but extend insights across the eukaryotes by extensive comparative genomic analysis. We argue that gene loss plays an important role in diversification of protein complexes and hence enhancement of eukaryotic diversity.
- Keywords
- constructive neutral evolution, evolutionary divergence, evolutionary mechanisms, gene replacement, molecular evolution, protein complexes,
- MeSH
- Eukaryota * genetics MeSH
- Phylogeny MeSH
- Genomics MeSH
- Evolution, Molecular * MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
The contractile vacuole complex (CVC) is a dynamic and morphologically complex membrane organelle, comprising a large vesicle (bladder) linked with a tubular reticulum (spongiome). CVCs provide key osmoregulatory roles across diverse eukaryotic lineages, but probing the mechanisms underlying their structure and function is hampered by the limited tools available for in vivo analysis. In the experimentally tractable ciliate Tetrahymena thermophila, we describe four proteins that, as endogenously tagged constructs, localize specifically to distinct CVC zones. The DOPEY homolog Dop1p and the CORVET subunit Vps8Dp localize both to the bladder and spongiome but with different local distributions that are sensitive to osmotic perturbation, whereas the lipid scramblase Scr7p colocalizes with Vps8Dp. The H+-ATPase subunit Vma4 is spongiome specific. The live imaging permitted by these probes revealed dynamics at multiple scales including rapid exchange of CVC-localized and soluble protein pools versus lateral diffusion in the spongiome, spongiome extension and branching, and CVC formation during mitosis. Although the association with DOP1 and VPS8D implicate the CVC in endosomal trafficking, both the bladder and spongiome might be isolated from bulk endocytic input.
- Keywords
- Tetrahymena, Contractile vacuole complex, Organelle biogenesis, Organelle dynamics, Osmotic regulation,
- MeSH
- Endosomes MeSH
- Mitosis MeSH
- Proteins metabolism MeSH
- Tetrahymena thermophila * MeSH
- Vacuoles * metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Research Support, N.I.H., Extramural MeSH
- Names of Substances
- Proteins MeSH
In the ciliate Tetrahymena thermophila, lysosome-related organelles called mucocysts accumulate at the cell periphery where they secrete their contents in response to extracellular events, a phenomenon called regulated exocytosis. The molecular bases underlying regulated exocytosis have been extensively described in animals but it is not clear whether similar mechanisms exist in ciliates or their sister lineage, the Apicomplexan parasites, which together belong to the ecologically and medically important superphylum Alveolata. Beginning with a T. thermophila mutant in mucocyst exocytosis, we used a forward genetic approach to uncover MDL1 (Mucocyst Discharge with a LamG domain), a novel gene that is essential for regulated exocytosis of mucocysts. Mdl1p is a 40 kDa membrane glycoprotein that localizes to mucocysts, and specifically to a tip domain that contacts the plasma membrane when the mucocyst is docked. This sub-localization of Mdl1p, which occurs prior to docking, underscores a functional asymmetry in mucocysts that is strikingly similar to that of highly polarized secretory organelles in other Alveolates. A mis-sense mutation in the LamG domain results in mucocysts that dock but only undergo inefficient exocytosis. In contrast, complete knockout of MDL1 largely prevents mucocyst docking itself. Mdl1p is physically associated with 9 other proteins, all of them novel and largely restricted to Alveolates, and sedimentation analysis supports the idea that they form a large complex. Analysis of three other members of this putative complex, called MDD (for Mucocyst Docking and Discharge), shows that they also localize to mucocysts. Negative staining of purified MDD complexes revealed distinct particles with a central channel. Our results uncover a novel macromolecular complex whose subunits are conserved within alveolates but not in other lineages, that is essential for regulated exocytosis in T. thermophila.
- MeSH
- Exocytosis genetics MeSH
- Lysosomes metabolism MeSH
- Organelles metabolism MeSH
- Secretory Vesicles genetics metabolism MeSH
- Tetrahymena thermophila * genetics MeSH
- Tetrahymena * MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Research Support, N.I.H., Extramural MeSH
Eukaryotic cells arose ~1.5 billion years ago, with the endomembrane system a central feature, facilitating evolution of intracellular compartments. Endomembranes include the nuclear envelope (NE) dividing the cytoplasm and nucleoplasm. The NE possesses universal features: a double lipid bilayer membrane, nuclear pore complexes (NPCs), and continuity with the endoplasmic reticulum, indicating common evolutionary origin. However, levels of specialization between lineages remains unclear, despite distinct mechanisms underpinning various nuclear activities. Several distinct modes of molecular evolution facilitate organellar diversification and to understand which apply to the NE, we exploited proteomic datasets of purified nuclear envelopes from model systems for comparative analysis. We find enrichment of core nuclear functions amongst the widely conserved proteins to be less numerous than lineage-specific cohorts, but enriched in core nuclear functions. This, together with consideration of additional evidence, suggests that, despite a common origin, the NE has evolved as a highly diverse organelle with significant lineage-specific functionality.
