Characterization of a novel regulator of the unfolded protein response in Ustilago maydis and mammals
Domenica Martorana
Doctoral thesis
Date of Examination:
2019-06-05
Date of issue:
2019-07-16
Advisor:
Heimel, Kai apl. Prof. Dr.
Referee:
Heimel, Kai apl. Prof. Dr.
Referee:
Braus, Gerhard Prof. Dr.
Persistent Address: http://hdl.handle.net/21.11130/00-1735-0000-0003-C168-E
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Abstract
English
The endoplasmic reticulum (ER) is an eukaryotic organelle which is the entry point into the secretory pathway and responsible for protein synthesis and processing. The amount of proteins to be folded in the ER lumen is highly variable and depends on different factors, such as the physiology and the environment of a cell. Accumulation of unfolded proteins in the ER activates the unfolded protein response (UPR) which functions to counter ER stress. Once activated, the UPR restores ER homeostasis or, if ER stress remains unresolved, induces apoptosis. In higher eukaryotes, the UPR is a dynamic signaling network regulated by three main transducers, ATF6, PERK and IRE1α. Activated IRE1α unconventionally splices the constitutively expressed XBP1 mRNA encoding the bZIP transcription factor XBP1s which, in turn, transcriptionally regulates the expression of UPR target genes. Under unstressed conditions, as well as during the recovery phase of ER stress, the unspliced XBP1 mRNA is translated into XBP1u, a highly unstable protein containing a bZIP domain. XBP1u has previously been demonstrated to function as a negative regulator of XBP1s. However, an increasing number of recent studies indicate that XBP1u plays a more important role in UPR regulation than assumed so far. A comprehensive understanding of the regulatory role of XBP1u in the ER stress response as well as the molecular details are still missing. In the course of this work XBP1u was shown to restore cell survival and promote cell proliferation under unstressed and partly under ER stress conditions. Moreover, ER stress resistance was increased in the presence of XBP1u without affecting the expression of common UPR target genes and fully independent of a functional DNA-binding domain or transcriptional activity. Additionally, the deletion of XBP1 resulted in morphological aberrations of the ER that were suppressed by XBP1u. Similar to the situation in higher eukaryotes, fungi activate the UPR to resolve ER stress but only rely on the conserved IRE1 signaling pathway. In the fungus U. maydis, the bZIP transcription factor Cib1s represents the main UPR regulator. The Cib1u protein, which is derived from the unspliced cib1 mRNA, is homologous to XBP1u and functions in the ER stress response independent of Cib1s. Analogous to XBP1u, Cib1u mediates increased ER stress resistance and this does not require a functional DNA-binding domain. Consistently, genome-wide analysis revealed that expression of common UPR target genes is not affected. Finally, it was demonstrated that Cib1u and XBP1u are functionally interchangeable. Collectively, these data strongly suggest a novel, previously undescribed, role of XBP1u in the UPR, which is likely to be evolutionary conserved.
Keywords: UPR; IRE1a; XBP1s; XBP1u; Ustilago maydis; cell culture; clonogenic survival; transcriptional activity; luciferase assay; Cib1u; unfolded protein response; Cib1s; bZIP transcription factor; Ustilago maydis
