Molecular cloning of the IRE1 gene encoding ER stress sensor Ire1p
It was a new gene when I sequenced it (it took three months) but was found to be identical to the IRE1 gene encoding inositol-requiring enzyme 1 (Ire1p), which was identified by Jun-ichi Nikawa in 1992 as a gene capable of complementing a yeast mutant requiring exogenous inositol for its growth.
If a protein kinase cascade occurred downstream of Ire1p, such screening would likely identify genes existing between IRE1 and my transcription factor gene, such as many protein kinase genes.
I came up with the idea that because yeast cells have only Ire1p as an ER stress sensor, knockout of IRE1 abolished the UPR signaling.
Disruption of the HAC1 gene abolished UPR signaling completely, as in cells lacking Ire1p. This confirmed it--yeast cells have only a single UPR-specific transcription factor!
The reviewers requested that we also reveal how Ire1p activation is connected to Hac1p production.
These changes in Hac1p level and HAC1 mRNA size do not occur in cells deficient in Ire1p. Third, because the 5' end of the intron is located within the HAC1 open reading frame, the Hac1p C-terminus is replaced from 10aa to 18aa without affecting the Hac1p N-terminus of 220aa, as a result of mRNA splicing.
HAC1 intron-mediated translational block might be released under certain circumstances in the absence of Ire1p activation, resulting in the production of Hac1p of 230aa.
Surprisingly, Peter unraveled that the endonuclease is Ire1p itself.
Randy Kaufman and David Ron identified IRE1[alpha] (ubiquitously expressed in mice) and IRE1[beta] (mainly expressed in the gut in mice), respectively, as mammalian homologues of yeast Ire1p in 1998.
(1997 The trans membrane kinase Ire1p is a site-specific endonuclease that initiates mRNA splicing in the unfolded protein response.