A Conserved Structural Determinant Located at the Interdomain Region of Mammalian IRE1α

Abstract

The UPR sensor inositol‐requiring enzyme 1α (IRE1α) is a bifunctional enzyme containing both kinase and RNase domains that are important for trans‐autophosphorylation and Xbp1 mRNA splicing, respectively, in response to ER stress. However, the amino acid residues important for structural integrity remain largely unknown. Here, through analysis of IRE1α mutants associated with human somatic cancers, we have identified a highly conserved proline residue at position 830 (P830) that is critical for its structural integrity, hence the activation of both kinase and RNase domains. Structural analysis revealed that P830 may form a highly conserved structural linker with adjacent tryptophan and tyrosine residues at positions 833 and 945 (W833 and Y945), thereby bridging the kinase and RNase domains. Indeed, mutation of P830 to leucine (P830L) completely abolished the kinase and RNase activities, significantly decreased protein stability and prevented oligomerization of IRE1α upon ER stress; similar observations were made for mutations of W833 to alanine (W833A) and to a lesser extent for Y945A. Our finding may facilitate the identification of small molecules to specifically compromise IRE1α function.