Store-operated calcium entry (SOCE) is the major Ca2+ entry pathway in most non-excitable cells maintaining prolonged elevation of cytosolic Ca2+ levels required for gene transcription. SOCE is activated by the loss of endoplasmic reticulum (ER) Ca2+ through stromal interaction molecules (STIM), ER-membrane associated Ca2+ sensors. In humans, STIM1 and STIM2 share 65% sequence similarity but differentially regulate SOCE. Biophysical studies on the luminal Ca2+-binding region suggests that STIM2 EF-SAM is more stable than STIM1. The NMR structure of Ca2+-loaded STIM2 EF-SAM determined in this work suggests a more stable SAM and a tighter EF-hand and SAM interaction in STIM2 may be account for its higher stability. Chimeric swapping of the EF-hand and SAM domains generates an unstable ES211. Introducing ES211 into cherryFP-STIM1 shows constitutive puncta which activate SOCE independent of ER depletion. The current work demonstrates that the instability of the EF-SAM plays an important role in regulating SOCE initiation.

Store-operated calcium entry (SOCE) is the major Ca2+ entry pathway in most non-excitable cells maintaining prolonged elevation of cytosolic Ca2+ levels required for gene transcription. SOCE is activated by the loss of endoplasmic reticulum (ER) Ca2+ through stromal interaction molecules (STIM), ER-membrane associated Ca2+ sensors. In humans, STIM1 and STIM2 share 65% sequence similarity but differentially regulate SOCE. Biophysical studies on the luminal Ca2+-binding region suggests that STIM2 EF-SAM is more stable than STIM1. The NMR structure of Ca2+-loaded STIM2 EF-SAM determined in this work suggests a more stable SAM and a tighter EF-hand and SAM interaction in STIM2 may be account for its higher stability. Chimeric swapping of the EF-hand and SAM domains generates an unstable ES211. Introducing ES211 into cherryFP-STIM1 shows constitutive puncta which activate SOCE independent of ER depletion. The current work demonstrates that the instability of the EF-SAM plays an important role in regulating SOCE initiation.

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