A putative flagellar Ca<sup>2+</sup>-binding protein of the flagellum of trypanosomatid protozoan parasites

Lee, Mary Gwo-Shu; Chen, Jifan; Ho, Ann-Shu; D'Alesandro, Philip A.; Ploeg, Lex H.T. Van der

doi:10.1093/nar/18.14.4252

Cited by 20 publications

(10 citation statements)

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“…13) The two trypanosome proteins 1 F8 (Gonzalez et al 1985) or p24 (Engman et al 1989) and TB17 (Lee et al 1990) are quite similar to one another by all of our analyses. They are in the same subfamily.…”

Section: Xyz29~mentioning

confidence: 56%

Evolution of EF-hand calcium-modulated proteins. II. Domains of several subfamilies have diverse evolutionary histories

1992

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In the first report in this series we described the relationships and evolution of 152 individual proteins of the EF-hand subfamilies. Here we add 66 additional proteins and define eight (CDC, TPNV, CLNB, LPS, DGK, 1F8, VIS, TCBP) new subfamilies and seven (CAL, SQUD, CDPK, EFH5, TPP, LAV, CRGP) new unique proteins, which we assume represent new subfamilies. The main focus of this study is the classification of individual EF-hand domains. Five subfamilies--calmodulin, troponin C, essential light chain, regulatory light chain, CDC31/caltractin--and three uniques--call, squidulin, and calcium-dependent protein kinase--are congruent in that all evolved from a common four-domain precursor. In contrast calpain and sarcoplasmic calcium-binding protein (SARC) each evolved from its own one-domain precursor. The remaining 19 subfamilies and uniques appear to have evolved by translocation and splicing of genes encoding the EF-hand domains that were precursors to the congruent eight and to calpain and to SARC. The rates of evolution of the EF-hand domains are slower following formation of the subfamilies and establishment of their functions. Subfamilies are not readily classified by patterns of calcium coordination, interdomain linker stability, and glycine and proline distribution. There are many homoplasies indicating that similar variants of the EF-hand evolved by independent pathways.

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Section: Xyz29~mentioning

confidence: 56%

Evolution of EF-hand calcium-modulated proteins. II. Domains of several subfamilies have diverse evolutionary histories

1992

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“…Equally important, FCaBP is a prototype of a growing group of flagellar calcium-binding proteins of parasites. Similar proteins have been identified in T. lewisi (13), T. brucei, the agent of African sleeping sickness (36,37), T. rangeli (38), and Tritrichomonas suis (39). Some of these proteins have up to eight potential EF-hand calcium-binding motifs, sites for N-terminal myristoylation and palmitoylation, and even C-terminal acylation.…”

Section: Discussionmentioning

confidence: 69%

A Flagellum-specific Calcium Sensor

Buchanan

Ames

Asfaw

et al. 2005

Journal of Biological Chemistry

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The flagellar calcium-binding protein (FCaBP) of the flagellated protozoan Trypanosoma cruzi associates with the flagellar membrane via its N-terminal myristate and palmitate moieties in a calcium-modulated, conformation-dependent manner. This mechanism of localization is similar to that described for neuronal calcium sensors, which undergo calcium-dependent changes in conformation, which modulate the availability of the acyl groups for membrane interaction and partner association. To test whether FCaBP undergoes a calcium-dependent conformational change and to explore the role of such a change in flagellar targeting, we first introduced point mutations into each of the two EF-hand calcium-binding sites of FCaBP to define their affinities. Analysis of recombinant EF-3 mutant (E151Q), EF-4 mutant (E188Q), and double mutant proteins showed EF-3 to be the high affinity site (K d ϳ9 M) and EF-4 the low affinity site (K d ϳ120 M). These assignments also correlated with partial (E188Q), nearly complete (E151Q), and complete (E151Q,E188Q) disruption of calcium-induced conformational changes determined by NMR spectrometry. We next expressed the FCaBP E151Q mutant and the double mutant in T. cruzi epimastigotes. These transproteins localized to the flagellum, suggesting the existence of a calcium-dependent interaction of FCaBP that is independent of its intrinsic calcium binding capacity. Several proteins were identified by FCaBP affinity chromatography that interact with FCaBP in a calcium-dependent manner, but with differential dependence on calcium-binding by FCaBP. These findings may have broader implications for the calcium acyl switch mechanism of protein regulation. Flagellar calcium-binding protein (FCaBP)2 is a 24-kDa highly immunogenic protein found in the flagellum of the protozoan parasite Trypanosoma cruzi (1). FCaBP contains four EF-hand calcium-binding domains, the third and fourth (EF-3 and EF-4) of which bind calcium (2). The protein is modified at the N terminus by the addition of myristate and palmitate, both of which are required for association with the inner leaflet of the flagellar membrane (3). Calcium is required for stable flagellar localization as well, since FCaBP can be washed out of detergent-permeabilized trypanosomes if calcium chelators are included in the wash solutions. Because of the size, acylation, calcium binding, membrane association, and calcium-dependent localization of FCaBP, we concluded that the protein is a calcium acyl switch protein. These proteins undergo calcium-dependent membrane association by virtue of calcium-regulated extrusion or sequestration of a myristate moiety that mediates membrane binding (4).The best studied calcium acyl switch protein is recoverin (Rv), a myristoylated calcium-binding protein of the mammalian retina (5) that inhibits rhodopsin kinase (RK) (6) and regulates recovery of the photoreceptor cell after photoexcitation (7). Rv regulates RK via the calcium myristoyl switch mechanism (4). In the resting state of the cell, Rv binds two calcium ions a...

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“…In T. brucei , another trypanosomatid, the down-regulation of this protein results in growth arrest and loss of cell viability [50]. Also in T. brucei , calmodulin is located in the flagellum and possibly takes part in the movement of the microtubules from the axoneme during the flagellar beat cycle [51]–[53]. Recycling of the transferrin receptor to the cell surface and exocytosis of transferrin [54] are calmodulin-dependent events, demonstrating a direct relationship between iron homeostasis and the expression of this protein.…”

Section: Discussionmentioning

confidence: 99%

Cellular Growth and Mitochondrial Ultrastructure of Leishmania (Viannia) braziliensis Promastigotes Are Affected by the Iron Chelator 2,2-Dipyridyl

et al. 2013

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BackgroundIron is an essential element for the survival of microorganisms in vitro and in vivo, acting as a cofactor of several enzymes and playing a critical role in host-parasite relationships. Leishmania (Viannia) braziliensis is a parasite that is widespread in the new world and considered the major etiological agent of American tegumentary leishmaniasis. Although iron depletion leads to promastigote and amastigote growth inhibition, little is known about the role of iron in the biology of Leishmania. Furthermore, there are no reports regarding the importance of iron for L. (V.) braziliensis.Methodology/Principal FindingsIn this study, the effect of iron on the growth, ultrastructure and protein expression of L. (V.) braziliensis was analyzed by the use of the chelator 2,2-dipyridyl. Treatment with 2,2-dipyridyl affected parasites' growth in a dose- and time-dependent manner. Multiplication of the parasites was recovered after reinoculation in fresh culture medium. Ultrastructural analysis of treated promastigotes revealed marked mitochondrial swelling with loss of cristae and matrix and the presence of concentric membranar structures inside the organelle. Iron depletion also induced Golgi disruption and intense cytoplasmic vacuolization. Fluorescence-activated cell sorting analysis of tetramethylrhodamine ester-stained parasites showed that 2,2-dipyridyl collapsed the mitochondrial membrane potential. The incubation of parasites with propidium iodide demonstrated that disruption of mitochondrial membrane potential was not associated with plasma membrane permeabilization. TUNEL assays indicated no DNA fragmentation in chelator-treated promastigotes. In addition, two-dimensional electrophoresis showed that treatment with the iron chelator induced up- or down-regulation of proteins involved in metabolism of nucleic acids and coordination of post-translational modifications, without altering their mRNA levels.ConclusionsIron chelation leads to a multifactorial response that results in cellular collapse, starting with the interruption of cell proliferation and culminating in marked mitochondrial impairment in some parasites and their subsequent cell death, whereas others may survive and resume proliferating.

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A putative flagellar Ca²⁺-binding protein of the flagellum of trypanosomatid protozoan parasites

Cited by 20 publications

References 3 publications

Evolution of EF-hand calcium-modulated proteins. II. Domains of several subfamilies have diverse evolutionary histories

Evolution of EF-hand calcium-modulated proteins. II. Domains of several subfamilies have diverse evolutionary histories

A Flagellum-specific Calcium Sensor

Cellular Growth and Mitochondrial Ultrastructure of Leishmania (Viannia) braziliensis Promastigotes Are Affected by the Iron Chelator 2,2-Dipyridyl

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A putative flagellar Ca2+-binding protein of the flagellum of trypanosomatid protozoan parasites

Cited by 20 publications

References 3 publications

Evolution of EF-hand calcium-modulated proteins. II. Domains of several subfamilies have diverse evolutionary histories

Evolution of EF-hand calcium-modulated proteins. II. Domains of several subfamilies have diverse evolutionary histories

A Flagellum-specific Calcium Sensor

Cellular Growth and Mitochondrial Ultrastructure of Leishmania (Viannia) braziliensis Promastigotes Are Affected by the Iron Chelator 2,2-Dipyridyl

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A putative flagellar Ca²⁺-binding protein of the flagellum of trypanosomatid protozoan parasites