Inositol 1,4,5‐trisphosphate receptor regulates replication, differentiation, infectivity and virulence of the parasitic protist <i><scp>T</scp>rypanosoma cruzi</i>

Hashimoto, Muneaki; Enomoto, Masahiro; Morales, Jorge; Kurebayashi, Nagomi; Sakurai, Takashi; Hashimoto, Takeji; Nara, Takeshi; Mikoshiba, Katsuhiko

doi:10.1111/mmi.12155

Cited by 52 publications

(97 citation statements)

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“…Whenever analyses were not restricted to overall nucleotide similarity, similar CRCs could be detected also in some other protozoan species. In the flagellated protozoans, Trypanosoma brucei [135] and Trypanosoma cruzi [136], IP 3 R channels have thus been identified, complemented by thorough experimental work comparable to that with Paramecium. This included activation studies, intracellular localisation, knockdown experiments and functional analysis.…”

Section: Mixed Types Of Crcs and Common Originmentioning

confidence: 97%

Molecular aspects of calcium signalling at the crossroads of unikont and bikont eukaryote evolution – The ciliated protozoan Paramecium in focus

Plattner

2015

Cell Calcium

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The ciliated protozoan, Paramecium tetraurelia has a high basic Ca(2+) leakage rate which is counteracted mainly by export through a contractile vacuole complex, based on its V-type H(+)-ATPase activity. In addition Paramecium cells dispose of P-type Ca(2+)-ATPases, i.e. a plasmamembrane and a sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase (PMCA, SERCA). Antiporter systems are to be expected, as inferred from indirect evidence. Among the best known cytosolic Ca(2+)-binding proteins, calmodulin activates Ca(2+) influx channels in the somatic cell membrane, but inactivates Ca(2+) influx channels in cilia, where it, thus, ends ciliary reversal induced by depolarization via channels in the somatic cell membrane. Centrin inactivates Ca(2+) signals after stimulation by its high capacity/low affinity binding sites, whereas its high affinity sites regulate some other functions. Cortical Ca(2+) stores (alveolar sacs) are activated during stimulated trichocyst exocytosis and thereby mediate store-operated Ca(2+) entry (SOCE). Ca(2+) release channels (CRCs) localised to alveoli and underlying SOCE are considered as Ryanodine receptor-like proteins (RyR-LPs) which are members of a CRC family with 6 subfamilies. These also encompass genuine inositol 1,4,5-trisphosphate receptors (IP3Rs) and intermediates between the two channel types. All IP3R/RyR-type CRCs possess six carboxyterminal transmembrane domains (TMD), with a pore domain between TMD 5 and 6, endowed with a characteristic selectivity filter. There are reasons to assume a common ancestor molecule for such channels and diversification further on in evolution. The distinct distribution of specific CRCs in the different vesicles undergoing intracellular trafficking suggests constitutive formation of very locally restricted Ca(2+) signals during vesicle-vesicle interaction. In summary, essential steps of Ca(2+) signalling already occur at this level of evolution, including an unexpected multitude of CRCs. For dis-/similarities with other bikonts see "Conclusions".

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Section: Mixed Types Of Crcs and Common Originmentioning

confidence: 97%

Molecular aspects of calcium signalling at the crossroads of unikont and bikont eukaryote evolution – The ciliated protozoan Paramecium in focus

Plattner

2015

Cell Calcium

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“…Only very recently has an InsP 3 R been cloned in full and analysed in functional tests within the pathogenic flagellates Trypanosoma brucei (Huang et al, 2013) and Trypanosoma cruzi (Hashimoto et al, 2013); however, the precise domain structure remains to be shown. For other protozoa, no data are available that are comparable with those of the P. tetraurelia CRC types CRC-II and CRC-IV (those that have been analysed most thoroughly) -although CRCs related to CRC-II, which serves for the release of Ca 2+ , must be ubiquitously present in all related organisms according to physiological data ( Table 1).…”

Section: Discussionmentioning

confidence: 99%

“…Characterized by domain and functional analysis Ladenburger and Plattner, 2011); see text for details. Plattner et al, 2012. f For recent analysis see Huang et al, 2013. g For recent analysis see Hashimoto et al, 2013. local increases in [Ca 2+ ] i by approximately two orders of magnitude. These localised changes in [Ca 2+ ] i regulate numerous different functions, from secretion (exocytosis), locomotion (including ciliary beat regulation) to cell division (Berridge et al, 2000;Clapham, 2007).…”

Section: Prokaryotic Foundation Of Camentioning

confidence: 99%

Ca2+ signalling early in evolution – all but primitive

Plattner

Verkhratsky

2013

Journal of Cell Science

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SummaryEarly in evolution, Ca 2+ emerged as the most important second messenger for regulating widely different cellular functions. In eukaryotic cells Ca 2+ signals originate from several sources, i.e. influx from the outside medium, release from internal stores or from both. In mammalian cells, Ca 2+ -release channels represented by inositol 1,4,5-trisphosphate receptors and ryanodine receptors (InsP 3 R and RyR, respectively) are the most important. In unicellular organisms and plants, these channels are characterised with much less precision. In the ciliated protozoan, Paramecium tetraurelia, 34 molecularly distinct Ca 2+ -release channels that can be grouped in six subfamilies, based on criteria such as domain structure, pore, selectivity filter and activation mechanism have been identified. Some of these channels are genuine InsP 3 Rs and some are related to RyRs. Others show some -but not all -features that are characteristic for one or the other type of release channel. Localisation and gene silencing experiments revealed widely different -yet distinctlocalisation, activation and functional engagement of the different Ca 2+ -release channels. Here, we shall discuss early evolutionary routes of Ca 2+ -release machinery in protozoa and demonstrate that detailed domain analyses and scrutinised functional analyses are instrumental for in-depth evolutionary mapping of Ca 2+ -release channels in unicellular organisms.

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“…The channel is localized to the inner mitochondrial membrane of a variety of cells, including Trypanosoma brucei (10). TcIP 3 R was reported to have ER localization in T. cruzi (11). However, the immunofluorescence evidence reported was disputed (12), because there was no clear reticular pattern or co-localization with a T. brucei ER marker, TbBiP, in the figures published (11).…”

mentioning

confidence: 99%

“…TcIP 3 R was reported to have ER localization in T. cruzi (11). However, the immunofluorescence evidence reported was disputed (12), because there was no clear reticular pattern or co-localization with a T. brucei ER marker, TbBiP, in the figures published (11). In addition, the T. brucei IP 3 R localized to the acidocalcisomes as demonstrated using antibodies against the endogenous tagged protein (13) and specific antibodies against the protein (14), as well as proteomic and functional analyses (13,14).…”

mentioning

confidence: 99%

CRISPR/Cas9-mediated endogenous C-terminal Tagging of Trypanosoma cruzi Genes Reveals the Acidocalcisome Localization of the Inositol 1,4,5-Trisphosphate Receptor

Lander

Chiurillo

Storey

et al. 2016

Journal of Biological Chemistry

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Edited by Dennis VoelkerMethods for genetic manipulation of Trypanosoma cruzi, the etiologic agent of Chagas disease, have been highly inefficient, and no endogenous tagging of genes has been reported to date. We report here the use of the CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated gene 9) system for endogenously tagging genes in this parasite. The utility of the method was established by tagging genes encoding proteins of known localization such as TcFCaBP (flagellar calcium binding protein) and TcVP1 (vacuolar proton pyrophosphatase), and two proteins of undefined or disputed localization, the TcMCU (mitochondrial calcium uniporter) and TcIP 3 R (inositol 1,4,5-trisphosphate receptor). We confirmed the flagellar and acidocalcisome localization of TcFCaBP and TcVP1 by co-localization with antibodies to the flagellum and acidocalcisomes, respectively. As expected, TcMCU was colocalized with the voltage-dependent anion channel to the mitochondria. However, in contrast to previous reports and our own results using overexpressed TcIP 3 R, endogenously tagged TcIP 3 R showed co-localization with antibodies against VP1 to acidocalcisomes. These results are also in agreement with our previous reports on the localization of this channel to acidocalcisomes of Trypanosoma brucei and suggest that caution should be exercised when overexpression of tagged genes is done to localize proteins in T. cruzi.The application of the CRISPR/Cas9 technology to the study of protist parasites has dramatically increased the tools available for their genetic manipulation (1). Trypanosoma cruzi, the etiologic agent of Chagas disease, which is a significant cause of morbidity and mortality from the South of the United States to the South of Argentina and Chile, has been particularly refractory to genetic manipulation. However, the recent use of the CRISPR/Cas9 technology to knock down or knock out genes (2, 3) has revolutionized their study.The localization of proteins is important to determine their cellular function, and previous studies in T. cruzi have used either antibodies or gene tagging methods with vectors that overexpressed the proteins (4). Although specific antibodies are useful to detect the endogenous proteins, it is not always possible to obtain them because either the proteins have low antigenicity or the antibodies cross-react with other proteins. Plasmids that enable the tagging of genes at their endogenous loci are not available for T. cruzi, and a major drawback of the overexpression of tagged proteins is that the proteins of interest sometimes are retained in the endoplasmic reticulum (ER) 4 or localize to other compartments.Here we have adapted the CRISPR/Cas9 system to tag genes of T. cruzi at their endogenous loci and tested this system with two genes encoding proteins of well recognized localization (flagellar calcium binding protein or TcFCaBP and the acidocalcisome vacuolar proton pyrophosphatase or TcVP1) and two encoding proteins of undefined or disputed localizati...

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Inositol 1,4,5‐trisphosphate receptor regulates replication, differentiation, infectivity and virulence of the parasitic protist Trypanosoma cruzi

Cited by 52 publications

References 64 publications

Molecular aspects of calcium signalling at the crossroads of unikont and bikont eukaryote evolution – The ciliated protozoan Paramecium in focus

Molecular aspects of calcium signalling at the crossroads of unikont and bikont eukaryote evolution – The ciliated protozoan Paramecium in focus

Ca2+ signalling early in evolution – all but primitive

CRISPR/Cas9-mediated endogenous C-terminal Tagging of Trypanosoma cruzi Genes Reveals the Acidocalcisome Localization of the Inositol 1,4,5-Trisphosphate Receptor

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