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1.
J Cell Sci ; 134(3)2021 02 11.
Artículo en Inglés | MEDLINE | ID: mdl-33495279

RESUMEN

The movement of ciliary membrane proteins is directed by transient interactions with intraflagellar transport (IFT) trains. The green alga Chlamydomonas has adapted this process for gliding motility, using retrograde IFT motors to move adhesive glycoproteins in the flagella membrane. Ca2+ signalling contributes directly to the gliding process, although uncertainty remains over the mechanism through which it acts. Here, we show that flagella Ca2+ elevations initiate the movement of paused retrograde IFT trains, which accumulate at the distal end of adherent flagella, but do not influence other IFT processes. On highly adherent surfaces, flagella exhibit high-frequency Ca2+ elevations that prevent the accumulation of paused retrograde IFT trains. Flagella Ca2+ elevations disrupt the IFT-dependent movement of microspheres along the flagella membrane, suggesting that Ca2+ acts by directly disrupting an interaction between retrograde IFT trains and flagella membrane glycoproteins. By regulating the extent to which glycoproteins on the flagella surface interact with IFT motor proteins on the axoneme, this signalling mechanism allows precise control of traction force and gliding motility in adherent flagella.


Asunto(s)
Chlamydomonas reinhardtii , Chlamydomonas , Transporte Biológico , Chlamydomonas/metabolismo , Chlamydomonas reinhardtii/genética , Chlamydomonas reinhardtii/metabolismo , Cilios/metabolismo , Dineínas/metabolismo , Flagelos/metabolismo , Transporte de Proteínas
2.
PLoS One ; 9(7): e103026, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-25050549

RESUMEN

Classically viewed as a cytosolic pathway, glycolysis is increasingly recognized as a metabolic pathway exhibiting surprisingly wide-ranging variations in compartmentalization within eukaryotic cells. Trypanosomatid parasites provide an extreme view of glycolytic enzyme compartmentalization as several glycolytic enzymes are found exclusively in peroxisomes. Here, we characterize Trypanosoma brucei flagellar proteins resembling glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoglycerate kinase (PGK): we show the latter associates with the axoneme and the former is a novel paraflagellar rod component. The paraflagellar rod is an essential extra-axonemal structure in trypanosomes and related protists, providing a platform into which metabolic activities can be built. Yet, bioinformatics interrogation and structural modelling indicate neither the trypanosome PGK-like nor the GAPDH-like protein is catalytically active. Orthologs are present in a free-living ancestor of the trypanosomatids, Bodo saltans: the PGK-like protein from B. saltans also lacks key catalytic residues, but its GAPDH-like protein is predicted to be catalytically competent. We discuss the likelihood that the trypanosome GAPDH-like and PGK-like proteins constitute molecular evidence for evolutionary loss of a flagellar glycolytic pathway, either as a consequence of niche adaptation or the re-localization of glycolytic enzymes to peroxisomes and the extensive changes to glycolytic flux regulation that accompanied this re-localization. Evidence indicating loss of localized ATP provision via glycolytic enzymes therefore provides a novel contribution to an emerging theme of hidden diversity with respect to compartmentalization of the ubiquitous glycolytic pathway in eukaryotes. A possibility that trypanosome GAPDH-like protein additionally represents a degenerate example of a moonlighting protein is also discussed.


Asunto(s)
Flagelos/enzimología , Glucólisis , Fosfoglicerato Quinasa/metabolismo , Trypanosoma brucei brucei/citología , Trypanosoma brucei brucei/genética , Secuencia de Aminoácidos , Evolución Molecular , Flagelos/genética , Flagelos/metabolismo , Eliminación de Gen , Gliceraldehído-3-Fosfato Deshidrogenasas/análisis , Gliceraldehído-3-Fosfato Deshidrogenasas/genética , Gliceraldehído-3-Fosfato Deshidrogenasas/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Fosfoglicerato Quinasa/análisis , Fosfoglicerato Quinasa/genética , Trypanosoma brucei brucei/enzimología , Trypanosoma brucei brucei/metabolismo
3.
Curr Biol ; 23(22): 2311-2318, 2013 Nov 18.
Artículo en Inglés | MEDLINE | ID: mdl-24210618

RESUMEN

Intraflagellar transport (IFT) underpins many of the important cellular roles of cilia and flagella in signaling and motility. The microtubule motors kinesin-2 and cytoplasmic dynein 1b drive IFT particles (protein complexes carrying ciliary component proteins) along the axoneme to facilitate the assembly and maintenance of cilia. IFT is regulated primarily by cargo loading onto the IFT particles, although evidence suggests that IFT particles also exhibit differential rates of movement. Here we demonstrate that intraflagellar Ca(2+) elevations act to directly regulate the movement of IFT particles. IFT-driven movement of adherent flagella membrane glycoproteins in the model alga Chlamydomonas enables flagella-mediated gliding motility. We find that surface contact promotes the localized accumulation of IFT particles in Chlamydomonas flagella. Highly compartmentalized intraflagellar Ca(2+) elevations initiate retrograde transport of paused IFT particles to modulate their accumulation. Gliding motility induces mechanosensitive intraflagellar Ca(2+) elevations in trailing (dragging) flagella only, acting to specifically clear the accumulated microtubule motors from individual flagella and prevent a futile tug-of-war. Our results demonstrate that compartmentalized intraciliary Ca(2+) signaling can regulate the movement of IFT particles and is therefore likely to play a central role in directing the movement and distribution of many ciliary proteins.


Asunto(s)
Señalización del Calcio , Chlamydomonas/citología , Chlamydomonas/metabolismo , Flagelos/metabolismo , Transporte Biológico/fisiología , Cilios/metabolismo , Potenciales de la Membrana
4.
Protist ; 164(4): 528-40, 2013 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-23787017

RESUMEN

In the flagellum of the African sleeping sickness parasite Trypanosoma brucei calmodulin (CaM) is found within the paraflagellar rod (PFR), an elaborate extra-axonemal structure, and the axoneme. In dissecting mechanisms of motility regulation we analysed CaM function using RNAi. Unexpectedly CaM depletion resulted in total and catastrophic failure in PFR assembly; even connections linking axoneme to PFR failed to form following CaM depletion. This provides an intriguing parallel with the role in the green alga Chlamydomonas of a CaM-related protein in docking outer-dynein arms to axoneme outer-doublet microtubules. Absence of CaM had no discernible effect on axoneme assembly, but the failure in PFR assembly was further compounded by loss of the normal linkage between PFR and axoneme to the flagellum attachment zone of the cell body. Thus, flagellum detachment was a secondary, time-dependent consequence of CaM RNAi, and coincided with the loss of normal trypomastigote morphology, thereby linking the presence of PFR architecture with maintenance of cell form, as well as cell motility. Finally, wider comparison between the flagellum detachment phenotypes of RNAi mutants for CaM and the FLA1 glycoprotein potentially provides new perspective into the function of the latter into establishing and maintaining flagellum-cell body attachment.


Asunto(s)
Calmodulina/metabolismo , Flagelos/metabolismo , Proteínas Protozoarias/metabolismo , Trypanosoma brucei brucei/metabolismo , Axonema/genética , Axonema/metabolismo , Calmodulina/genética , Movimiento Celular , Flagelos/genética , Humanos , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Protozoarias/genética , Trypanosoma brucei brucei/citología , Trypanosoma brucei brucei/genética , Tripanosomiasis Africana/parasitología
5.
BMC Bioinformatics ; 13: 117, 2012 May 30.
Artículo en Inglés | MEDLINE | ID: mdl-22646090

RESUMEN

BACKGROUND: The generation of multiple sequence alignments (MSAs) is a crucial step for many bioinformatic analyses. Thus improving MSA accuracy and identifying potential errors in MSAs is important for a wide range of post-genomic research. We present a novel method called MergeAlign which constructs consensus MSAs from multiple independent MSAs and assigns an alignment precision score to each column. RESULTS: Using conventional benchmark tests we demonstrate that on average MergeAlign MSAs are more accurate than MSAs generated using any single matrix of sequence substitution. We show that MergeAlign column scores are related to alignment precision and hence provide an ab initio method of estimating alignment precision in the absence of curated reference MSAs. Using two novel and independent alignment performance tests that utilise a large set of orthologous gene families we demonstrate that increasing MSA performance leads to an increase in the performance of downstream phylogenetic analyses. CONCLUSION: Using multiple tests of alignment performance we demonstrate that this novel method has broad general application in biological research.


Asunto(s)
Alineación de Secuencia/métodos , Programas Informáticos , Sustitución de Aminoácidos , Secuencia de Consenso , Procesamiento de Imagen Asistido por Computador , Filogenia
6.
Parasitology ; 137(9): 1467-75, 2010 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-20233494

RESUMEN

Enzymes moonlight in a non-enzymatic capacity in a diverse variety of cellular processes. The discovery of these non-enzymatic functions is generally unexpected, and moonlighting enzymes are known in both prokaryotes and eukaryotes. Importantly, this unexpected multi-functionality indicates that caution might be needed on some occasions in interpreting phenotypes that result from the deletion or gene-silencing of some enzymes, including some of the best known enzymes from classic intermediary metabolism. Here, we provide an overview of enzyme moonlighting in parasitic protists. Unequivocal and putative examples of moonlighting are discussed, together with the possibility that the unusual biological characteristics of some parasites either limit opportunities for moonlighting to arise or perhaps contribute to the evolution of novel proteins with clear metabolic ancestry.


Asunto(s)
Enzimas/metabolismo , Eucariontes/enzimología , Proteínas Protozoarias/metabolismo , Animales , Apicomplexa/enzimología , Glucólisis , Humanos , Parásitos/enzimología , Trichomonas/enzimología , Trypanosomatina/enzimología
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