Water in peripheral TM-interfaces of Orai1-channels triggers pore opening.

The activation of the Ca-channel Orai1 via the physiological activator stromal interaction molecule 1 (STIM1) requires structural rearrangements within the entire channel complex involving a series of gating checkpoints. Focusing on the gating mechanism operating along the peripheral transmembrane domain (TM) 3/TM4-interface, we report here that some charged substitutions close to the center of TM3 or TM4 lead to constitutively active Orai1 variants triggering nuclear factor of activated T-cell (NFAT) translocation into the nucleus. Molecular dynamics simulations unveil that this gain-of-function correlates with enhanced hydration at peripheral TM-interfaces, leading to increased local structural flexibility of the channel periphery and global conformational changes permitting pore opening.

Synthetic Biology Meets Ca Release-Activated Ca Channel-Dependent Immunomodulation.

Many essential biological processes are triggered by the proximity of molecules. Meanwhile, diverse approaches in synthetic biology, such as new biological parts or engineered cells, have opened up avenues to precisely control the proximity of molecules and eventually downstream signaling processes. This also applies to a main Ca entry pathway into the cell, the so-called Ca release-activated Ca (CRAC) channel.

Photocrosslinking-induced CRAC channel-like Orai1 activation independent of STIM1.

Ca release-activated Ca (CRAC) channels, indispensable for the immune system and various other human body functions, consist of two transmembrane (TM) proteins, the Ca-sensor STIM1 in the ER membrane and the Ca ion channel Orai1 in the plasma membrane. Here we employ genetic code expansion in mammalian cell lines to incorporate the photocrosslinking unnatural amino acids (UAA), p-benzoyl-L-phenylalanine (Bpa) and p-azido-L-phenylalanine (Azi), into the Orai1 TM domains at different sites. Characterization of the respective UAA-containing Orai1 mutants using Ca imaging and electrophysiology reveal that exposure to UV light triggers a range of effects depending on the UAA and its site of incorporation.

CRAC and SK Channels: Their Molecular Mechanisms Associated with Cancer Cell Development.

Cancer represents a major health burden worldwide. Several molecular targets have been discovered alongside treatments with positive clinical outcomes. However, the reoccurrence of cancer due to therapy resistance remains the primary cause of mortality.

Orai1 Boosts SK3 Channel Activation.

The interplay of SK3, a Ca sensitive K ion channel, with Orai1, a Ca ion channel, has been reported to increase cytosolic Ca levels, thereby triggering proliferation of breast and colon cancer cells, although a molecular mechanism has remained elusive to date. We show in the current study, via heterologous protein expression, that Orai1 can enhance SK3 K currents, in addition to constitutively bound calmodulin (CaM). At low cytosolic Ca levels that decrease SK3 K permeation, co-expressed Orai1 potentiates SK3 currents.