Structure-function analysis and MD-simulations of STIM-mediated Ca2+ influx
Sprache des Titels:
Englisch
Original Kurzfassung:
The Stromal Interaction Molecule (STIM) is a distinct single-pass transmembrane protein located in the
endoplasmic reticulum (ER) membrane. Together with the plasma membrane resident, highly selective Ca2+
channel Orai, STIM initiates Ca2+ influx, depending on the ER calcium content. Structural destabilization of
the compactly folded N-terminal EF-SAM domain of STIM, triggered by the loss of an EF-hand bound Ca2+
ion and followed by di/heteromerization, is only partially resolved. Mutations within this luminal domain are
linked to pathological phenotypes and are associated with various forms of cancer. In our current study we
analysed several amino acid residues located within the SAM domain of STIM1, potentially important in the
luminal aggregation process via inter- and intramolecular EF-SAM interactions. We therefore used molecular
dynamics (MD) simulations together with live-cell imaging methods to gain detailed insight into the EF-SAM
di/heteromerization and activation mechanism. Structural unfolding of the EF-SAM domain, already
described for constitutively active mutants, induces store independent SOCE initiation via puncta formation,
while our SAM mutants lack the ability to form inducible puncta. On the basis of our results we present
essential key residues in the EF-SAM domain, crucial for the initiation of the SOCE activation cascade, as
they show reduced puncta formation, lower intermolecular FRET levels and are incapable to fully activate
Orai channels upon mutation.