Large-Scale Phylogenetic Analysis of Trypanosomatid Adenylate Cyclases Reveals Associations with Extracellular Lifestyle and Host–Pathogen Interplay

Abstract Receptor adenylate cyclases (RACs) on the surface of trypanosomatids are important players in the host–parasite interface. They detect still unidentified environmental signals that affect the parasites’ responses to host immune challenge, coordination of social motility, and regulation of cell division. A lesser known class of oxygen-sensing adenylate cyclases (OACs) related to RACs has been lost in trypanosomes and expanded mostly in Leishmania species and related insect-dwelling trypanosomatids. In this work, we have undertaken a large-scale phylogenetic analysis of both classes of adenylate cyclases (ACs) in trypanosomatids and the free-living Bodo saltans. We observe that the expanded RAC repertoire in trypanosomatids with a two-host life cycle is not only associated with an extracellular lifestyle within the vertebrate host, but also with a complex path through the insect vector involving several life cycle stages. In Trypanosoma brucei, RACs are split into two major clades, which significantly differ in their expression profiles in the mammalian host and the insect vector. RACs of the closely related Trypanosoma congolense are intermingled within these two clades, supporting early RAC diversification. Subspecies of T. brucei that have lost the capacity to infect insects exhibit high numbers of pseudogenized RACs, suggesting many of these proteins have become redundant upon the acquisition of a single-host life cycle. OACs appear to be an innovation occurring after the expansion of RACs in trypanosomatids. Endosymbiont-harboring trypanosomatids exhibit a diversification of OACs, whereas these proteins are pseudogenized in Leishmania subgenus Viannia. This analysis sheds light on how ACs have evolved to allow diverse trypanosomatids to occupy multifarious niches and assume various lifestyles.

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Metabolic reprogramming during the Trypanosoma brucei life cycle

F1000Research ◽

10.12688/f1000research.10342.2 ◽

2017 ◽

Vol 6 ◽

pp. 683 ◽

Cited By ~ 31

Author(s):

Terry K. Smith ◽

Frédéric Bringaud ◽

Derek P. Nolan ◽

Luisa M. Figueiredo

Keyword(s):

Life Cycle ◽

Trypanosoma Brucei ◽

Model Organism ◽

Protozoan Parasite ◽

Tsetse Fly ◽

Metabolic Reprogramming ◽

Mammalian Host ◽

Insect Vector ◽

Environmental Signals ◽

Cellular Metabolic Activity

Cellular metabolic activity is a highly complex, dynamic, regulated process that is influenced by numerous factors, including extracellular environmental signals, nutrient availability and the physiological and developmental status of the cell. The causative agent of sleeping sickness, Trypanosoma brucei, is an exclusively extracellular protozoan parasite that encounters very different extracellular environments during its life cycle within the mammalian host and tsetse fly insect vector. In order to meet these challenges, there are significant alterations in the major energetic and metabolic pathways of these highly adaptable parasites. This review highlights some of these metabolic changes in this early divergent eukaryotic model organism.

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Metabolic reprogramming during the Trypanosoma brucei life cycle

F1000Research ◽

10.12688/f1000research.10342.1 ◽

2017 ◽

Vol 6 ◽

pp. 683 ◽

Cited By ~ 8

Author(s):

Terry K. Smith ◽

Frédéric Bringaud ◽

Derek P. Nolan ◽

Luisa M. Figueiredo

Keyword(s):

Life Cycle ◽

Trypanosoma Brucei ◽

Model Organism ◽

Protozoan Parasite ◽

Tsetse Fly ◽

Metabolic Reprogramming ◽

Mammalian Host ◽

Insect Vector ◽

Environmental Signals ◽

Cellular Metabolic Activity

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Assessment of reference genes at six different developmental stages of Schistosoma mansoni for quantitative RT-PCR

Scientific Reports ◽

10.1038/s41598-021-96055-7 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Gilbert O. Silveira ◽

Murilo S. Amaral ◽

Helena S. Coelho ◽

Lucas F. Maciel ◽

Adriana S. A. Pereira ◽

...

Keyword(s):

Gene Expression ◽

Life Cycle ◽

Schistosoma Mansoni ◽

Reference Genes ◽

Large Scale ◽

Developmental Stages ◽

Adult Males ◽

Histone H4 ◽

Expression Levels ◽

Life Cycle Stages

AbstractReverse-transcription quantitative real-time polymerase chain reaction (RT-qPCR) is the most used, fast, and reproducible method to confirm large-scale gene expression data. The use of stable reference genes for the normalization of RT-qPCR assays is recognized worldwide. No systematic study for selecting appropriate reference genes for usage in RT-qPCR experiments comparing gene expression levels at different Schistosoma mansoni life-cycle stages has been performed. Most studies rely on genes commonly used in other organisms, such as actin, tubulin, and GAPDH. Therefore, the present study focused on identifying reference genes suitable for RT-qPCR assays across six S. mansoni developmental stages. The expression levels of 25 novel candidates that we selected based on the analysis of public RNA-Seq datasets, along with eight commonly used reference genes, were systematically tested by RT-qPCR across six developmental stages of S. mansoni (eggs, miracidia, cercariae, schistosomula, adult males and adult females). The stability of genes was evaluated with geNorm, NormFinder and RefFinder algorithms. The least stable candidate reference genes tested were actin, tubulin and GAPDH. The two most stable reference genes suitable for RT-qPCR normalization were Smp_101310 (Histone H4 transcription factor) and Smp_196510 (Ubiquitin recognition factor in ER-associated degradation protein 1). Performance of these two genes as normalizers was successfully evaluated with females maintained unpaired or paired to males in culture for 8 days, or with worm pairs exposed for 16 days to double-stranded RNAs to silence a protein-coding gene. This study provides reliable reference genes for RT-qPCR analysis using samples from six different S. mansoni life-cycle stages.

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Inhibitor of Cysteine Proteases Is Critical for Motility and Infectivity of Plasmodium Sporozoites

mBio ◽

10.1128/mbio.00874-13 ◽

2013 ◽

Vol 4 (6) ◽

Cited By ~ 12

Author(s):

Katja E. Boysen ◽

Kai Matuschewski

Keyword(s):

Life Cycle ◽

Salivary Glands ◽

Cysteine Proteases ◽

Gliding Motility ◽

Parasitic Infections ◽

Mammalian Host ◽

Insect Vector ◽

Mosquito Vector ◽

Infected Female ◽

Wide Range

ABSTRACT Malaria is transmitted when motile sporozoites are injected into the dermis by an infected female Anopheles mosquito. Inside the mosquito vector, sporozoites egress from midgut-associated oocysts and eventually penetrate the acinar cells of salivary glands. Parasite-encoded factors with exclusive vital roles in the insect vector can be studied by classical reverse genetics. Here, we characterized the in vivo roles of Plasmodium berghei falstatin/ICP (inhibitor of cysteine proteases). This protein was previously suggested to act as a protease inhibitor during erythrocyte invasion. We show by targeted gene disruption that loss of ICP function does not affect growth inside the mammalian host but causes a complete defect in sporozoite transmission. Sporogony occurred normally in icp(−) parasites, but hemocoel sporozoites showed a defect in continuous gliding motility and infectivity for salivary glands, which are prerequisites for sporozoite transmission to the mammalian host. Absence of ICP correlates with enhanced cleavage of circumsporozoite protein, in agreement with a role as a protease regulator. We conclude that ICP is essential for only the final stages of sporozoite maturation inside the mosquito vector. This study is the first genetic evidence that an ICP is necessary for the productive motility of a eukaryotic parasitic cell. IMPORTANCE Cysteine proteases and their inhibitors are considered ideal drug targets for the treatment of a wide range of diseases, including cancer and parasitic infections. In protozoan parasites, including Leishmania, Trypanosoma, and Plasmodium, cysteine proteases play important roles in life cycle progression. A mouse malaria model provides an unprecedented opportunity to study the roles of a parasite-encoded inhibitor of cysteine proteases (ICP) over the entire parasite life cycle. By precise gene deletion, we found no evidence that ICP influences disease progression or parasite virulence. Instead, we discovered that this factor is necessary for parasite movement and malaria transmission from mosquitoes to mammals. This finding in a fast-moving unicellular protozoan has important implications for malaria intervention strategies and the roles of ICPs in the regulation of eukaryotic cell migration.

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Amastigotes of Trypanosoma cruzi escape destruction by the terminal complement components.

Journal of Experimental Medicine ◽

10.1084/jem.169.3.881 ◽

1989 ◽

Vol 169 (3) ◽

pp. 881-891 ◽

Cited By ~ 36

Author(s):

K Iida ◽

M B Whitlow ◽

V Nussenzweig

Keyword(s):

Trypanosoma Cruzi ◽

Alternative Pathway ◽

Surface Protein ◽

Protozoan Parasite ◽

Mammalian Host ◽

Insect Vector ◽

Complement Components ◽

Life Cycle Stages ◽

Sds Page ◽

Terminal Complement

We studied the effect of complement on two life cycle stages of the protozoan parasite Trypanosoma cruzi: epimastigotes, found in the insect vector, and amastigotes, found in the mammalian host. We found that while both stages activate vigorously the alternative pathway, only epimastigotes are destroyed. The amounts of C3 and C5b-7 deposited on the amastigotes were similar to those bound to the much larger epimastigotes. Binding of C9 to amastigotes was four to six times less than binding to epimastigotes, resulting in a lower C9/C5b-7 ratio. Although a fairly large amount of C9 bound stably to amastigotes, no functional channels were formed as measured by release of incorporated 86Rb. The bound C9 had the characteristic properties of poly-C9, that is, it expressed a neo-antigen unique to poly-C9, and migrated in SDS-PAGE with an apparent Mr greater than 10(5). The poly-C9 was removed from the surface of amastigotes by treatment with trypsin, indicating that it was not inserted in the lipid bilayer. Modification of amastigote surface by pronase treatment rendered the parasites susceptible to complement attack. These results suggest that amastigotes have a surface protein that binds to the C5b-9 complex and inhibits membrane insertion, thus protecting the parasites from complement-mediated lysis.

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The ecology and evolution of microsporidian parasites

Parasitology ◽

10.1017/s0031182009991818 ◽

2009 ◽

Vol 136 (14) ◽

pp. 1901-1914 ◽

Cited By ~ 43

Author(s):

J. E. SMITH

Keyword(s):

Phylogenetic Analysis ◽

Life Cycle ◽

Sex Ratio ◽

Vertical Transmission ◽

Environmental Risk ◽

Disease Phenotype ◽

Life Cycle Stages ◽

Wide Range ◽

Ratio Distortion ◽

Sex Ratio Distortion

SUMMARYThe phylum Microspora is ancient and diverse and affects a wide range of hosts. There is unusually high use of vertical transmission and this has significant consequences for transmission and pathogenicity. Vertical transmission is associated with low pathogenesis but nevertheless can have significant impact through associated traits such as sex ratio distortion. The majority of microsporidia have mixed transmission cycles and it is not clear whether they are able to modify their phenotype according to environmental circumstances. There is a great need to understand the mechanisms controlling transmission and one of the first challenges for the genomics era is to find genes associated with life cycle stages. Similarly we cannot currently predict the ease with which these parasites might switch between host groups. Phylogenetic analysis suggests that there are strong relationships between Microsporidia and their hosts. However closer typing of parasite isolates, in relation to host range and disease phenotype, is required to assess future environmental risk from these pathogens.

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Functional Analyses of Cytokinesis Regulators in Bloodstream Stage Trypanosoma brucei Parasites Identify Functions and Regulations Specific to the Life Cycle Stage

mSphere ◽

10.1128/msphere.00199-19 ◽

2019 ◽

Vol 4 (3) ◽

Cited By ~ 5

Author(s):

Xuan Zhang ◽

Tai An ◽

Kieu T. M. Pham ◽

Zhao-Rong Lun ◽

Ziyin Li

Keyword(s):

Life Cycle ◽

Trypanosoma Brucei ◽

Protozoan Parasite ◽

Bloodstream Form ◽

Signaling Cascade ◽

Insect Vector ◽

Content Type ◽

Procyclic Form ◽

Life Cycle Stages ◽

Evolutionarily Conserved

ABSTRACT The early divergent protozoan parasite Trypanosoma brucei alternates between the insect vector and the mammalian hosts during its life cycle and proliferates through binary cell fission. The cell cycle control system in T. brucei differs substantially from that in its mammalian hosts and possesses distinct mitosis-cytokinesis checkpoint controls between two life cycle stages, the procyclic form and the bloodstream form. T. brucei undergoes an unusual mode of cytokinesis, which is controlled by a novel signaling cascade consisting of evolutionarily conserved protein kinases and trypanosome-specific regulatory proteins in the procyclic form. However, given the distinct mitosis-cytokinesis checkpoints between the two forms, it is unclear whether the cytokinesis regulatory pathway discovered in the procyclic form also operates in a similar manner in the bloodstream form. Here, we showed that the three regulators of cytokinesis initiation, cytokinesis initiation factor 1 (CIF1), CIF2, and CIF3, are interdependent for subcellular localization but not for protein stability as in the procyclic form. Further, we demonstrated that KLIF, a regulator of cytokinesis completion in the procyclic form, plays limited roles in cytokinesis in the bloodstream form. Finally, we showed that the cleavage furrow-localizing protein FRW1 is required for cytokinesis initiation in the bloodstream form but is nonessential for cytokinesis in the procyclic form. Together, these results identify conserved and life cycle-specific functions of cytokinesis regulators, highlighting the distinction in the regulation of cytokinesis between different life cycle stages of T. brucei. IMPORTANCE The early divergent protozoan parasite Trypanosoma brucei is the causative agent of sleeping sickness in humans and nagana in cattle in sub-Saharan Africa. This parasite has a complex life cycle by alternating between the insect vector and the mammalian hosts and proliferates by binary cell fission. The control of cell division in trypanosomes appears to be distinct from that in its human host and differs substantially between two life cycle stages, the procyclic (insect) form and the bloodstream form. Cytokinesis, the final step of binary cell fission, is regulated by a novel signaling cascade consisting of two evolutionarily conserved protein kinases and a cohort of trypanosome-specific regulators in the procyclic form, but whether this signaling pathway operates in a similar manner in the bloodstream form is unclear. In this report, we performed a functional analysis of multiple cytokinesis regulators and discovered their distinct functions and regulations in the bloodstream form.

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Gene expression profiles of dicyemid life-cycle stages may explain how dispersing larvae locate new hosts

Zoological Letters ◽

10.1186/s40851-019-0146-y ◽

2019 ◽

Vol 5 (1) ◽

Author(s):

Tsai-Ming Lu ◽

Hidetaka Furuya ◽

Noriyuki Satoh

Keyword(s):

Gene Expression ◽

Life Cycle ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Molecular Transport ◽

Complex Life Cycle ◽

Sensory Response ◽

Specific Gene ◽

Life Cycle Stages ◽

New Hosts

Abstract Metazoans have evolved a great variety of life histories in response to environmental conditions. A unique example is encountered in dicyemid mesozoans. In addition to a highly simplified adult body comprising only ~ 30 cells, dicyemids exhibit a parasitic lifestyle that includes nematogens (asexual reproductive adults), rhombogens (sexual reproductive adults), vermiform larvae generated by nematogens, and infusoriform larvae generated by rhombogens. However, due to the difficulties of observing microscopic endoparasites, the complex life cycle and biological functions of life-cycle stages of dicyemids have remained mysterious. Taking advantage of the recently decoded genome of Dicyema japonicum, we examined genes that undergird this lifestyle. Using stage-specific gene expression profiles, we found that biological processes associated with molecular transport, developmental regulation, and sensory response are specified at different stages. Together with the expression of potential neurotransmitters, we further suggest that apical cells in infusoriform larva probably serve sensory functions, although dicyemids have no nervous system. Gene expression profiles show that more genes are expressed in free-living infusoriform larvae than in the other three stages, and that some of these genes are likely involved in locating new hosts. These data provide molecular information about the unique lifestyle of dicyemids and illustrate how an extremely simplified endoparasite adapted and retained gene sets and morphological characters to complete its life cycle.

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The role of membrane transporters in Leishmania virulence

Emerging Topics in Life Sciences ◽

10.1042/etls20170119 ◽

2017 ◽

Vol 1 (6) ◽

pp. 601-611 ◽

Cited By ~ 1

Author(s):

Snezhana Akpunarlieva ◽

Richard Burchmore

Keyword(s):

Life Cycle ◽

Amino Acid ◽

Membrane Transporters ◽

Mammalian Host ◽

Insect Vector ◽

Parasite Life Cycle ◽

Severe Morbidity ◽

Intracellular Amastigotes ◽

Genomic Analyses

Leishmania are parasitic protozoa which infect humans and cause severe morbidity and mortality. Leishmania parasitise as extracellular promastigotes in the insect vector and as intracellular amastigotes in the mammalian host. Cycling between hosts involves implementation of stringent and co-ordinated responses to shifting environmental conditions. One of the key dynamic aspects of Leishmania biology is substrate acquisition and metabolism. Genomic analyses have revealed that Leishmania encode many putative membrane transporters, many of which are differentially expressed during the parasite life cycle. Only a small fraction of these transporters, however, have been functionally characterised. Currently, most information is available about nutrient transporters, mainly involved in carbohydrate, amino acid, nucleobase and nucleoside, cofactor, and ion acquisition. Several have apparent roles in Leishmania virulence and will be discussed in this perspective.

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