Ca2+ signaling proteins and their involvement in cancer homeostasis
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The 2nd Lijiang International Forum on Pharmaceutical sciences
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The Ca2+ selective channel Orai1 mediates Ca2+ signalling for T cell activation, mast cell degranulation and platelet aggregation. Besides these important physiological immune cell functions, growing evidence suggests a key role of Orai channels in cancer cell development.
In this work, me and my team functionally evaluated reported single point mutants of the Orai channel family derived from large-scale cancer genomics data. We systematically screened for gain-of-function and lack-of-function Orai1 mutants using cellular Ca2+ imaging and a gene regulation assay. Specifically, these Orai1 mutants were fluorescence tagged to determine their cellular localization and evaluated for potential constitutive activation of the transcription factor NFAT, the nuclear factor of activated T cells. Ca2+ imaging was used as a complimentary approach to screen for constitutive Orai1 mutants and additionally determined non-functional channels. Identified Orai1 mutants with altered Ca2+/NFAT signalling were further functionally evaluated by whole-cell patch-clamp technique for selectivity and gating. Additionally, we analysed the gating mechanism of constitutively active Orai1 mutants. In a systematic cysteine crosslinking approach, pore residues were revealed that undergo rearrangement from the closed to the open Orai1 channel to relieve hydrophobic or electrostatic barriers.
The investigation of Orai1 cancer mutants in their respective environment determined the pathophysiological role of Orai1 mutants to alter Ca2+ signals and gene regulation and will further resolve functional mechanisms for Ca2+ channel gating and permeation in cancer cells.
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