Bypass of Activation Loop Phosphorylation by Aspartate 836 in Activation of the Endoribonuclease Activity of Ire1.

The bifunctional protein kinase-endoribonuclease Ire1 initiates splicing of the mRNA for the transcription factor Hac1 when unfolded proteins accumulate in the endoplasmic reticulum. Activation of Ire1 coincides with autophosphorylation of its activation loop at S840, S841, T844, and S850. Mass spectrometric analysis of Ire1 expressed in identified S837 as another potential phosphorylation site Mutation of all five potential phosphorylation sites in the activation loop decreased, but did not completely abolish, splicing of mRNA, induction of and mRNAs, and expression of a β-galactosidase reporter activated by Hac1 Phosphorylation site mutants survive low levels of endoplasmic reticulum stress better than deletions strains.