Heather J. Esson

Morphology of the Trypanosome Bilobe, a Novel Cytoskeletal Structure

ABSTRACT The trypanosome bilobe is a cytoskeletal structure of unclear function. To date, four proteins have been shown to localize stably to it: TbMORN1, TbLRRP1, TbCentrin2, and TbCentrin4. In this study, a combination of immunofluorescence microscopy and electron microscopy was used to explore the morphology of the bilobe and its relationship to other nearby cytoskeletal structures in the African trypanosome procyclic trypomastigote. The use of detergent/salt-extracted flagellum preparations was found to be an effective way of discerning features of the cytoskeletal ultrastructure that are normally obscured. TbMORN1 and TbCentrin4 together define a hairpin structure comprising an arm of TbCentrin4 and a fishhook of TbMORN1. The two arms flank a specialized microtubule quartet and the flagellum attachment zone filament, with TbMORN1 running alongside the former and TbCentrin4 alongside the latter. The hooked part of TbMORN1 sits atop the flagellar pocket collar marked by TbBILBO1. The TbMORN1 bilobe occasionally exhibits tendrillar extensions that seem to be connected to the basal and probasal bodies. The TbMORN1 molecules present on these tendrils undergo higher rates of turnover than those for molecules on the main bilobe structure. These observations have been integrated with previous detailed descriptions of the cytoskeletal elements in trypanosome cells.

Morphology and molecular phylogeny of a marine interstitial tetraflagellate with putative endosymbionts: Auranticordis quadriverberis n. gen. et sp. (Cercozoa)

BackgroundComparative morphological studies and environmental sequencing surveys indicate that marine benthic environments contain a diverse assortment of microorganisms that are just beginning to be explored and characterized. The most conspicuous predatory flagellates in these habitats range from about 20–150 μm in size and fall into three major groups of eukaryotes that are very distantly related to one another: dinoflagellates, euglenids and cercozoans. The Cercozoa is a diverse group of amoeboflagellates that cluster together in molecular phylogenies inferred mainly from ribosomal gene sequences. These molecular phylogenetic studies have demonstrated that several enigmatic taxa, previously treated as Eukaryota insertae sedis, fall within the Cercozoa, and suggest that the actual diversity of this group is largely unknown. Improved knowledge of cercozoan diversity is expected to help resolve major branches in the tree of eukaryotes and demonstrate important cellular innovations for understanding eukaryote evolution.ResultsA rare tetraflagellate, Auranticordis quadriverberis n. gen. et sp., was isolated from marine sand samples. Uncultured cells were in low abundance and were individually prepared for electron microscopy and DNA sequencing. These flagellates possessed several novel features, such as (1) gliding motility associated with four bundled recurrent flagella, (2) heart-shaped cells about 35–75 μm in diam., and (3) bright orange coloration caused by linear arrays of muciferous bodies. Each cell also possessed about 2–30 pale orange bodies (usually 4–5 μm in diam.) that were enveloped by two membranes and sac-like vesicles. The innermost membrane invaginated to form unstacked thylakoids that extended towards a central pyrenoid containing tailed viral particles. Although to our knowledge, these bodies have never been described in any other eukaryote, the ultrastructure was most consistent with photosynthetic endosymbionts of cyanobacterial origin. This combination of morphological features did not allow us to assign A. quadriverberis to any known eukaryotic supergroup. Thus, we sequenced the small subunit rDNA sequence from two different isolates and demonstrated that this lineage evolved from within the Cercozoa.ConclusionOur discovery and characterization of A. quadriverberis underscores how poorly we understand the diversity of cercozoans and, potentially, represents one of the few independent cases of primary endosymbiosis within the Cercozoa and beyond.

A model for the morphogenesis of strip reduction patterns in phototrophic euglenids: evidence for heterochrony in pellicle evolution

SUMMARY We propose a general developmental model that explains the evolutionary origin, diversification, and inheritance of pellicle strip patterns in phototrophic euglenids. Dividing cells of Euglena gracilis, E. viridis, and Phacus similis were observed with scanning electron microscopy in order to study the morphogenesis of posterior whorls of strip reduction. We found evidence that constant whorl numbers are maintained through cell division because of organized strip growth before and during cytokinesis. Alternating nascent strips form a new whorl of strip reduction at each of the anterior and posterior ends of daughter cells. Strips that terminated to form posterior whorls in the mother cell change in length during the development of daughter cells. In the mother cells of E. gracilis, the strips forming whorls I and II grow to become whorls II and III, respectively, in the daughter cells; the strips forming whorl III in the mother cell lengthen and meet with other strips already present at the posterior tip of daughter cells. This process of whorl morphogenesis during asexual reproduction is consistent with known variation in pellicle strip patterns and suggests that heterochrony played a major role in the ultrastructural evolution of phototrophic euglenids.

NOVEL PELLICLE SURFACE PATTERNS ON EUGLENA OBTUSA (EUGLENOPHYTA) FROM THE MARINE BENTHIC ENVIRONMENT: IMPLICATIONS FOR PELLICLE DEVELOPMENT AND EVOLUTION 1

Euglena obtusa F. Schmitz possesses novel pellicle surface patterns, including the greatest number of strips (120) and the most posterior subwhorls of strip reduction in any euglenid described so far. Although the subwhorls form a mathematically linear pattern of strip reduction, the pattern observed here differs from the linear pattern described for Euglena mutabilis F. Schmitz in that it contains seven linear subwhorls, rather than three, and is developmentally equivalent to three whorls of exponential reduction, rather than two. These properties imply that the seven‐subwhorled linear pattern observed in E. obtusa is evolutionarily derived from an ancestral bilinear pattern, rather than from a linear pattern, of strip reduction. Furthermore, analysis of the relative lateral positions of the strips forming the subwhorls in E. obtusa indicates that (1) the identity (relative length, lateral position, and maturity) of each strip in any mother cell specifies that strip’s identity in one of the daughter cells following pellicle duplication and cell division, (2) the relative length of any given pellicle strip regulates the length of the nascent strip it will produce during pellicle duplication, and (3) pellicle pores develop within the heels of the most mature pellicle strips. These observations suggest that continued research on pellicle development could eventually establish an ideal system for understanding mechanisms associated with the morphogenesis and evolution of related eukaryotic cells.

Evolution of Distorted Pellicle Patterns in Rigid Photosynthetic Euglenids (Phacus Dujardin)

ABSTRACT. Members of the euglenid genus Phacus are morphologically differentiated from other photosynthetic species by the presence of a rigid cytoskeleton (pellicle) and predominantly dorsoventrally flattened, leaf‐shaped cells. In order to better understand the evolutionary history of this lineage, we used scanning electron microscopy to examine patterns of pellicle strips in Phacus acuminatus, Phacus longicauda var. tortus, Phacus triqueter, Phacus segretii, Phacus pleuronectes, Phacus similis, Phacus pusillus, Phacus orbicularis, Phacus warszewiczii, and Discoplastis spathirhyncha, a putative close relative of Phacus and Lepocinclis. Our observations showed that while the earliest diverging species in our analyses, namely P. warszewiczii, has three whorls of exponential reduction, most members of Phacus have clustered patterns of posterior strip reduction that are bilaterally symmetrical distortions of the radially symmetrical “whorled” patterns found in other photosynthetic euglenids. Comparative morphology, interpreted within the context of molecular phylogenetic analyses of combined nuclear small subunit rDNA and partial nuclear large subunit rDNA sequences, demonstrates that clustered patterns of posterior strip reduction arose after the divergence of Phacus from other photosynthetic euglenids and are the result of developmental processes that govern individual strip length. Clustered patterns of pellicle strips in Phacus do not appear to be adaptively significant themselves; they evolved in association with the origin of cell flattening and cell rigidity, which may be adaptations to a planktonic lifestyle.

Visualizing the complex substructure of euglenid pellicle strips with SEM

H.J. Esson and B.S. Leander. 2008. Visualizing the complex substructure of euglenid pellicle strips with SEM. Phycologia 47: 529–532. DOI: 10.2216/08-26.1 Comparative analysis of cytoskeletal diversity within the Euglenophyceae has provided important context for understanding the phylogenetic relationships and major evolutionary transitions within the group (e.g. switches in modes of nutrition and motility). Some ultrastructural characters used in earlier cladistic analyses of euglenids involved different states for the lateral projections that extend from the frame of each pellicle strip in photosynthetic lineages. Previously, the overall structure of ‘strip projections’ in different lineages was (arduously) reconstructed from a series of ultra-thin sections viewed with transmission electron microscopy (TEM). In this study, we were able to determine the structure of strip projections with greater precision, and without the laborious protocols associated with TEM (e.g. ultramicrotomy), by examining disrupted pellicles from three photosynthetic euglenids (Lepocinclis fusiformis, Phacus longicauda var. tortus, and P. segretii) using scanning electron microscopy (SEM). The structure of the strip projections observed here demonstrated that either (1) previous TEM studies of the pellicle overlooked certain ultrastructural features in some taxa or (2) the (prearticular) strip projections in L. fusiformis, P. segretii, and P. longicauda var. tortus represent a novel character state that could be phylogenetically informative.

Extensive gain and loss of photosystem I subunits in chromerid algae, photosynthetic relatives of apicomplexans

In oxygenic photosynthesis the initial photochemical processes are carried out by photosystem I (PSI) and II (PSII). Although subunit composition varies between cyanobacterial and plastid photosystems, the core structures of PSI and PSII are conserved throughout photosynthetic eukaryotes. So far, the photosynthetic complexes have been characterised in only a small number of organisms. We performed in silico and biochemical studies to explore the organization and evolution of the photosynthetic apparatus in the chromerids Chromera velia and Vitrella brassicaformis, autotrophic relatives of apicomplexans. We catalogued the presence and location of genes coding for conserved subunits of the photosystems as well as cytochrome b6f and ATP synthase in chromerids and other phototrophs and performed a phylogenetic analysis. We then characterised the photosynthetic complexes of Chromera and Vitrella using 2D gels combined with mass-spectrometry and further analysed the purified Chromera PSI. Our data suggest that the photosynthetic apparatus of chromerids underwent unique structural changes. Both photosystems (as well as cytochrome b6f and ATP synthase) lost several canonical subunits, while PSI gained one superoxide dismutase (Vitrella) or two superoxide dismutases and several unknown proteins (Chromera) as new regular subunits. We discuss these results in light of the extraordinarily efficient photosynthetic processes described in Chromera.