A hexahistidine-Zn
            <sup>2+</sup>
            -dye label reveals STIM1 surface exposure

Hauser, Christina; Tsien, Roger Y.

doi:10.1073/pnas.0611713104

Cited by 148 publications

(96 citation statements)

References 30 publications

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“…It is of note that large N-terminal fusion proteins in contrast to C-terminal ones interfere with STIM1 plasma-membrane insertion. 54 Functional evidence for plasmamembrane STIM1 is presented by antibodies against an N-terminal sequence of STIM1 that results in partial suppression of SOCs, 16,55 but could not be confirmed in another study. 52 Reports that observe STIM1 distribution underneath but not in the plasma-membrane have been widely based on TIRF microscopy using YFP-tagged STIM1 as well as other, even larger tags.…”

Section: Store-operated Activation Of Stim/oraimentioning

confidence: 99%

The STIM/Orai coupling machinery

Frischauf¹,

Schindl²,

Derler³

et al. 2008

Channels

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Section: Store-operated Activation Of Stim/oraimentioning

confidence: 99%

The STIM/Orai coupling machinery

Frischauf¹,

Schindl²,

Derler³

et al. 2008

Channels

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“…The unusually long coiled-coil domain of STIM1 likely provides the structural features needed for connection between the two membranes to transfer the signal from the ER to the PM that activates CRAC channel. A likely candidate for such interactions is PM-STIM1 (6,11,13,14). However, direct interactions with Orai1 are also possible.…”

Section: Analysis Of the Crac Channel Selectivity By Co-expression Ofmentioning

confidence: 99%

“…STIM1 aggregates into puncta and translocates toward the PM, when the stores are depleted (5, 16). Physical incorporation of STIM1 molecules into the PM after Ca 2ϩ store depletion has been clearly demonstrated (11,13). The time course of the ER Ca 2ϩ depletion followed by CRAC activation correlates with the time course of STIM1 translocation that is ϳ52 s (5, 17).…”

mentioning

confidence: 99%

“…STIM1 functions as the Ca 2ϩ sensor in the ER that triggers the CRAC channel activation (5,6,11). However, its translocation to the PM upon activation (11,13) and functional role in the PM have also been demonstrated (6,14). Even though a report by Mignen et al (14) revealed that PM-STIM1 regulates another Ca 2ϩ channel (arachidonate-regulated Ca 2ϩ channels), the same group later revealed that arachidonate-regulated Ca 2ϩ channels have the same molecular identity as CRAC channels (Ref.…”

mentioning

confidence: 99%

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Voltage Gating at the Selectivity Filter of the Ca2+ Release-activated Ca2+ Channel Induced by Mutation of the Orai1 Protein

Spassova

Hewavitharana

Fandino

et al. 2008

Journal of Biological Chemistry

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The Ca 2؉ release-activated Ca 2؉ (CRAC) channel is a plasma membrane (PM) channel that is uniquely activated when free Ca 2؉ level in the endoplasmic reticulum (ER) is substantially reduced. Several small interfering RNA screens identified two membrane proteins, Orai1 and STIM1, to be essential for the CRAC channel function. STIM1 appears to function in the PM and as the Ca 2؉ sensor in the ER. Orai1 is forming the pore of the CRAC channel. Despite the recent breakthroughs, a mechanistic understanding of the CRAC channel gating is still lacking. Here we reveal new insights on the structure-function relationship of STIM1 and Orai1. Our data suggest that the cytoplasmic coiled-coil region of STIM1 provides structural means for coupling of the ER membrane to the PM to activate the CRAC channel. We mutated two hydrophobic residues in this region to proline (L286P/L292P) to introduce a kink in the first ␣-helix of the coiled-coil domain. This STIM1 mutant caused a dramatic inhibition of the CRAC channel gating compared with the wild type. Structure-function analysis of the Orai1 protein revealed the presence of intrinsic voltage gating of the CRAC channel. A mutation of Orai1 (V102I) close to the selectivity filter modified CRAC channel voltage sensitivity. Expression of the Orai1 V102I mutant resulted in slow voltage gating of the CRAC channel by negative potentials. The results revealed that the alteration of Val 102 develops voltage gating in the CRAC channel. Our data strongly suggest the presence of a novel voltage gating mechanism at the selectivity filter of the CRAC channel.The CRAC 2 channel is activated by depletion of the intracellular Ca 2ϩ stores and mediates sustained Ca 2ϩ entry in hematopoietic cells (1-3) and some other cell types (1, 3). Until recently, the molecular identity of the machinery linking store depletion with Ca 2ϩ entry remained elusive. However, recent studies identified two proteins, STIM1 and Orai1, as being essential for CRAC function (4 -11). Although STIM1 resides predominantly in the ER and the plasma membrane (PM) (11), Orai1 is almost exclusively expressed in the PM (12). STIM1 functions as the Ca 2ϩ sensor in the ER that triggers the CRAC channel activation (5, 6, 11). However, its translocation to the PM upon activation (11, 13) and functional role in the PM have also been demonstrated (6,14). Even though a report by Mignen et al. (14) revealed that PM-STIM1 regulates another Ca 2ϩ channel (arachidonate-regulated Ca 2ϩ channels), the same group later revealed that arachidonate-regulated Ca 2ϩ channels have the same molecular identity as CRAC channels (Ref. 41; see also Ref. 15). STIM1 aggregates into puncta and translocates toward the PM, when the stores are depleted (5, 16). Physical incorporation of STIM1 molecules into the PM after Ca 2ϩ store depletion has been clearly demonstrated (11,13). The time course of the ER Ca 2ϩ depletion followed by CRAC activation correlates with the time course of STIM1 translocation that is ϳ52 s (5, 17).How STIM1 relays signals about ER C...

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“…Some STIM1 resides in the PM in unstimulated cells (Manji et al, 2000;Williams et al, 2002), and externally applied anti-STIM1 antibodies have been reported to inhibit channel activation , but the relationship between PM STIM1 and the STIM1 that moves to puncta after store depletion is not clear. Many studies on STIM1 localization were performed with YFP-tagged fusion proteins that could not insert into the PM because of the YFP moiety (Hauser and Tsien, 2007) yet these fusion proteins were clearly capable of activating CRAC channels (Liou et al, 2005;Baba et al, 2006;Mercer et al, 2006;Soboloff et al, 2006;Wu et al, 2006). Recent work has shown that STIM1 and Orai1 are in membrane contact points between the ER and PM and that movement of a complex containing the two proteins requires at least 10 nm between the two membranes (Varnai et al, 2007).…”

mentioning

confidence: 99%

Dynamic Movement of the Calcium Sensor STIM1 and the Calcium Channel Orai1 in Activated T-Cells: Puncta and Distal Caps

Barr

Bernot

Srikanth

et al. 2008

MBoC

129

142

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The proteins STIM1 and Orai1 are the long sought components of the store-operated channels required in T-cell activation. However, little is known about the interaction of these proteins in T-cells after engagement of the T-cell receptor. We found that T-cell receptor engagement caused STIM1 and Orai1 to colocalize in puncta near the site of stimulation and accumulate in a dense structure on the opposite side of the T-cell. FRET measurements showed a close interaction between STIM1 and Orai1 both in the puncta and in the dense cap-like structure. The formation of cap-like structures did not entail rearrangement of the entire endoplasmic reticulum. Cap formation depended on TCR engagement and tyrosine phosphorylation, but not on channel activity or Ca 2؉ influx. These caps were very dynamic in T-cells activated by contact with superantigen pulsed B-cells and could move from the distal pole to an existing or a newly forming immunological synapse. One function of this cap may be to provide preassembled Ca 2؉ channel components to existing and newly forming immunological synapses. INTRODUCTIONT-cell activation in response to antigen induces and requires the elevation of intracellular Ca 2ϩ (Hogan et al., 2003;Quintana et al., 2005;Feske, 2007). T-cell receptor (TCR) engagement leads to activation of well-documented signal transduction pathways that cause a rapid release of Ca 2ϩ from the endoplasmic reticulum (ER; Samelson, 2002;Panyi et al., 2004;Gwack et al., 2007a). The depletion of Ca 2ϩ from this internal store then leads to the opening of ion channels in the plasma membrane (PM) allowing an influx of Ca 2ϩ from outside the cell. The store-operated channel responsible for Ca 2ϩ influx in T-cells is known as the Ca 2ϩ release-activated Ca 2ϩ (CRAC) channel, which has been studied by electrophysiological techniques for many years (Lewis, 2001;Prakriya and Lewis, 2003;Cahalan et al., 2007;Hewavitharana et al., 2007;Hogan and Rao, 2007;Putney, 2007a).Sustained elevation of cytosolic Ca 2ϩ is required for complete T-cell activation (Iezzi et al., 1998;Lewis, 2001;Hogan et al., 2003;Feske, 2007). High levels of intracellular Ca 2ϩ are necessary to maintain the interaction between a T-cell and antigen-presenting cell (APC) that leads to formation of the specialized contact surface known as the immunological synapse (IS). Increased Ca 2ϩ levels are also required for the activation of transcription factors. In particular, elevated Ca 2ϩ maintains prolonged nuclear accumulation of nuclear factor of activated T-cells (NFAT) by activating the phosphatase calcineurin that dephosphorylates NFAT, allowing it to translocate to the nucleus and activate genes such as IL2 (Hogan et al., 2003;Macian, 2005). Several hours of Ca 2ϩ influx are required to complete the T-cell activation program, which involves expression of a large number of activation-associated genes (Lewis, 2001;Macian et al., 2002;Hogan et al., 2003).Although sustained Ca 2ϩ influx in T-cells has been studied for decades, the proteins involved have only been ident...

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A hexahistidine-Zn ²⁺ -dye label reveals STIM1 surface exposure

Cited by 148 publications

References 30 publications

The STIM/Orai coupling machinery

The STIM/Orai coupling machinery

Voltage Gating at the Selectivity Filter of the Ca2+ Release-activated Ca2+ Channel Induced by Mutation of the Orai1 Protein

Dynamic Movement of the Calcium Sensor STIM1 and the Calcium Channel Orai1 in Activated T-Cells: Puncta and Distal Caps

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A hexahistidine-Zn 2+ -dye label reveals STIM1 surface exposure

Cited by 148 publications

References 30 publications

The STIM/Orai coupling machinery

The STIM/Orai coupling machinery

Voltage Gating at the Selectivity Filter of the Ca2+ Release-activated Ca2+ Channel Induced by Mutation of the Orai1 Protein

Dynamic Movement of the Calcium Sensor STIM1 and the Calcium Channel Orai1 in Activated T-Cells: Puncta and Distal Caps

Contact Info

Product

Resources

About

A hexahistidine-Zn ²⁺ -dye label reveals STIM1 surface exposure