Calcium in ciliated protozoa: Sources, regulation, and calcium-regulated cell functions

Plattner, Helmut; Klauke, Norbert

doi:10.1016/s0074-7696(01)01003-8

Cited by 105 publications

(142 citation statements)

References 397 publications

(288 reference statements)

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“…Therefore, these IP 3 Rs may here mediate a latent, graded reflux of Ca 2ϩ for fine-tuning of [Ca 2ϩ ] i and thus serve [Ca 2ϩ ] homeostasis (53). Besides [Ca 2ϩ ] homeostasis, the Paramecium cell has to regulate a variety of well-characterized processes (75). This includes exocytosis of dense-core secretory vesicles (trichocysts) (71,74,99).…”

Section: Camentioning

confidence: 99%

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Ladenburger¹,

Sehring²,

Korn³

et al. 2009

Molecular and Cellular Biology

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A database search of the Paramecium genome reveals 34 genes related to Ca 2؉ -release channels of the inositol-1,4,5-trisphosphate (IP 3 ) or ryanodine receptor type (IP 3 R, RyR). Phylogenetic analyses show that these Ca 2؉ release channels (CRCs) can be subdivided into six groups (Paramecium tetraurelia CRC-I to CRC-VI), each one with features in part reminiscent of IP 3 Rs and RyRs. We characterize here the P. tetraurelia CRC-IV-1 gene family, whose relationship to IP 3 Rs and RyRs is restricted to their C-terminal channel domain. CRC-IV-1 channels localize to cortical Ca 2؉ stores (alveolar sacs) and also to the endoplasmic reticulum. This is in contrast to a recently described true IP 3 channel, a group II member (P. tetraurelia IP 3 R N -1), found associated with the contractile vacuole system. Silencing of either one of these CRCs results in reduced exocytosis of dense core vesicles (trichocysts), although for different reasons. Knockdown of P. tetraurelia IP 3 R N affects trichocyst biogenesis, while CRC-IV-1 channels are involved in signal transduction since silenced cells show an impaired release of Ca 2؉ from cortical stores in response to exocytotic stimuli. Our discovery of a range of CRCs in Paramecium indicates that protozoans already have evolved multiple ways for the use of Ca 2؉ as signaling molecule.Ca 2ϩ is an important component of cell activity in all organisms, from protozoa to mammals. Thereby Ca 2ϩ may originate from the outside medium and/or from internal stores (7, 18). Ca 2ϩ release from internal stores is mediated by various Ca 2ϩ release channels (CRCs), of which the inositol-1,4,5-trisphosphate receptor (IP 3 R) and ryanodine receptor (RyR) families have been studied most extensively (8,9,29,63 (14). IP 3 generates and maintains a Ca 2ϩ gradient in the hyphal tip of Neurospora crassa and the IP 3 -sensitive channels have been reconstituted and characterized with the planar bilayer method (87). In summary, these publications suggest that IP 3 -dependent signaling pathways are conserved among unicellular organisms, including protozoa.Despite these data, the molecular characterization of IP 3 or ryanodine receptors in low eukaryotes is currently a challenge since the identification of orthologues has not been possible thus far, probably because of evolutionary sequence divergence (66). Traynor et al. (96) identified an IP 3 receptor-like protein, IplA, in Dictyostelium discoideum, which possesses regions related to IP 3 R sequences, but thus far no evidence for IP 3 interaction exists. We have recently described an IP 3 R in the ciliated protozoa Paramecium tetraurelia (referred to here as P. tetraurelia IP 3 R N ) (53), with features characteristic of mammalian IP 3 Rs in terms of topology and ability for IP 3 binding. The expression level of P. tetraurelia IP 3 R N is modulated by extracellular Ca 2ϩ concentrations ([Ca 2ϩ ] o ) and immunofluorescence studies reveal an unexpected localization to the contractile vacuole complex (CVC), the major organelle involved in osmoregulation...

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Section: Camentioning

confidence: 99%

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Ladenburger¹,

Sehring²,

Korn³

et al. 2009

Molecular and Cellular Biology

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121

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“…This made us confident to see any potential effect on Ca 2 +-signaling, if it would exist, without any side-effects. Using routine ultrathin section electron microscopy, we first analyzed the effect of CytB on the positioning not only of ASs but also of mitochondria-another organelle of relevance for Ca2+ -signaling [15]. Yet, no effect on their positioning was found in quantitative evaluations (data not shown).…”

Section: Effects Of Actin-reactive Drugs On Sacmentioning

confidence: 99%

Refilling of cortical calcium stores in Paramecium cells: in situ analysis in correlation with store-operated calcium influx

et al. 2003

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This is the first thorough study of refilling of a cortical calcium store in a secretory cell after stimulation in which we combined widely different methodologies. Stimulation of dense-core vesicle ("trichocysts") exocytosis in Paramecium involves a Ca2+ -influx" superimposed to Ca2+ -release from cortical stores ("alveolar sacs" (ASs). In quenched-flow experiments, membrane fusion frequency rose with increasing [Ca2+]o in the medium, from~20-25% at [Ca 2 +]0 :::0.25 j.LM to 100% at [Ca 2 +]0 between 2 and 10 j.LM, i.e. close to the range of estimated local intracellular [Ca2+] during membrane fusion. Next, we analyzed Ca2+ -specific fluorochrome signals during stimulation under different conditions. Treatment with actin-reactive drugs had no effect on Ca2+ -signaling. In double trigger experiments, with BAPTA in the second secretagogue application (BAPTA only for stimulation and analysis), the cortical Ca2+ -signal (due solely toCa2+ released from cortical stores) recovered with 11/2~65 min. When ASs were analyzed in situ by X-ray microanalysis after different trigger times (+Ca2+0),11/2 for store refilling was similar,~60 min. These values are similar to previously measured 45Ca2+ -uptake by isolated ASs. In sum we find, (i) exogenous Ca2+ increases exocytosis/membrane fusion performance with EC50 = 0.7 j.LM, (ii) Ca 2 +-signaling in this system is not sensitive to actin-reactive drugs, and (iii) refilling of these cortical calcium stores goes on over hours and thus is much slower than expected.

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“…Additional information comes from injection of calcium buffers of different affinity (K d ) immediately act followed by AED stimulation. Thus, we determined that a local [Ca 2 1 L of ~5 uM is required to induce exocytosis (Plattner and Klauke, 2001). This is ~ 10 x higher than registered by fluorochromes in the cell cortex during AED stimulation (Klauke and Plattner, 1997)-a phenomenon which is well known from other cells (Marengo and Monck, 2000;Neher, 1998).…”

Section: A Correlation Of Quenched-flow/em Analysis With Supplementamentioning

confidence: 74%

“…1995(Neher. , 1998; see also Plattner and Klauke, 2001) to be exploited by the available improved instrumentation. In the end, it would be desirable to determine calcium concentrations, [Ca] and respectively, with the highest possible spatial and temporal resolution.…”

Section: A Correlation Of Quenched-flow/em Analysis With Supplementamentioning

confidence: 99%

“…are easy to perform with rapidly frozen cells. this is much more difficult with some other phenomena, (e.g.. calcium signaling) (Plattner and Klauke, 2001) for which we discuss useful preparation protocols in conjunction with subsequent analysis by EM-based x-ray microanalysis. The method described here is different from that successfully used with muscle cells in previous work (Sornlyo et al, 1981;Wendt-Gallitelli and Isenberg, 1991).…”

Section: Introductionmentioning

confidence: 99%

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Sub-Second Cellular Dynamics: Time-Resolved Electron Microscopy and Functional Correlation

Plattner¹,

Hentschel²

2006

International Review of Cytology

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Subcellular processes, from molecular events to organellar responses and cell movement, cover a broad scale in time and space. Clearly the extremes, such as ion channel activation are accessible only by electrophysiology, whereas numerous routine methods exist for relatively slow processes. However, many other processes, from a millisecond time scale on, can be "caught" only by methods providing appropriate time resolution. Fast freezing (cryofixation) is the method of choice in that case. In combination with follow-up methodologies appropriate for electron microscopic (EM) analysis, with all its variations, such technologies can also provide high spatial resolution. Such analyses may include, for example, freeze-fracturing for analyzino restructuring of membrane components, scanning EM and other standard EM techniques, as well as analytical EM analyses. The latter encompass energy-dispersive x-ray microanalysis and electron spectroscopic imaging, all applicable, for instance, to the second messenger, calcium. Most importantly, when conducted in parallel, such analyses can provide a structural background to the functional analyses, such as cyclic nucleotide formation or protein de-or rephosphorylation during cell stimulation. In sum, we discuss many examples of how it is practically possible to achieve strict function-structure correlations in the sub-second time range. We complement this review by discussing alternative methods currently available to analyze fast cellular phenomena occurring in the sub-second time range.

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Calcium in ciliated protozoa: Sources, regulation, and calcium-regulated cell functions

Cited by 105 publications

References 397 publications

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Refilling of cortical calcium stores in Paramecium cells: in situ analysis in correlation with store-operated calcium influx

Sub-Second Cellular Dynamics: Time-Resolved Electron Microscopy and Functional Correlation

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Calcium in ciliated protozoa: Sources, regulation, and calcium-regulated cell functions

Cited by 105 publications

References 397 publications

Novel Types of Ca2+ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Novel Types of Ca2+ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Refilling of cortical calcium stores in Paramecium cells: in situ analysis in correlation with store-operated calcium influx

Sub-Second Cellular Dynamics: Time-Resolved Electron Microscopy and Functional Correlation

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Product

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Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells

Novel Types of Ca²⁺ Release Channels Participate in the Secretory Cycle of Paramecium Cells