Signal transduction from the endoplasmic reticulum to the nucleus in mammalian cells.

The unfolded protein response (UPR) is an intercompartmental signaling pathway between the endoplasmic reticulum (ER) and the nucleus. This signaling pathway protects eukaryotic cells from stress caused by accumulation of unfolded or misfolded protein in the ER by upregulating the transcription of genes encoding ER protein folding machinery, such as the protein chaperone BiP. In Saccharomyces cerevisiae, the signaling is mediated through an ER receptor kinase, Ire1p. Ire1p exhibits bifunctional catalytic activities: protein Serine/Threonine kinase and endoribonuclease. The dual activity is essential for UPR-dependent splicing of HAC1 mRNA encoding a transcriptional factor that transactivates the UPR responsive genes. In mammalian cells, two related genes encoding proteins homologous to yeast Ire1p have been identified and are referred to as IRE1alpha and IRE1beta. IRE1alpha was first identified from a human complementary DNA library. Human Ire1alphap (hIre1alphap) displayed intrinsic autophosphorylation activity and an endoribonuclease activity that cleaved the 5' splice site of yeast HAC1 mRNA, a substrate for the endoribonuclease activity of yeast Ire1p. Over-expression of wild-type hIre1alphap constitutively activated a reporter gene under transcriptional control of the rat BiP promoter. In contrast, expression of a catalytically inactive hIre1alphap acted in a trans-dominant negative manner to prevent transcriptional activation of the BiP promoter in response to ER stress. The endoribonuclease activity of hIre1alphap is critical for both the UPR and autoregulation by of limiting the hIRE1alpha mRNA abundance. The endoribonuclease activity of hIre1alphap is required to degrade hIRE1alpha mRNA, thus limiting hIre1alphap synthesis. To investigate the physiological significance of the UPR signaling pathway in a multicellular organism, a mouse model deficient in IRE1alpha was generated by gene targeting. In comparison to wild-type and heterozygous mIRE1alpha+/- mouse embryonic fibroblast (MEF), MEFs from mIREalpha-/- mice had only a slight reduction in BiP mRNA induction in response to tunicamycin treatment, indicating a redundancy in this signaling pathway. While heterozygous mIREalpha+/- mice appear normal, the homozygous mIRE1alpha-/- mouse embryos are significantly smaller and frequently found dead at 12.5 days p.c. indicating that this signaling pathway plays a distinct essential role for mammalian development. These results indicate the presence of at least two UPR signaling pathways that play distinct roles in embryonic development.Ph.D.BiochemistryBiological SciencesMolecular biologyPure SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/132261/2/9959875.pd