Heat induces the splicing by IRE1 of a mRNA encoding a transcription factor involved in the unfolded protein response in Arabidopsis

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189

The unfolded protein response (UPR) endows plants with the capacity to perceive, respond, and protect themselves from adverse environmental conditions. The UPR signaling pathway in Arabidopsis has two "arms," one arm involving the bifunctional protein kinase (PK)/ribonuclease, IRE1, a RNA splicing enzyme, and another involving membrane-associated transcription factors, such as basic leucine zipper transcription factor 28 (bZIP28). Because of functional redundancies, single gene mutations in the plant UPR signaling pathway generally have not resulted in prominent phenotypes. In this study we generated multiple mutations in the UPR signaling pathway, such as an ire1a ire1b double mutant, which showed defects in stress tolerance and vegetative growth and development. Complementation of ire1a ire1b with constructs containing site-specific mutations in the PK or RNase domains of IRE1b demonstrated that a functional RNase domain is required for endoplasmic reticulum stress tolerance, and that both the PK and RNase domains are required for normal vegetative growth under unstressed conditions. Root growth under stress conditions was dependent on the splicing target of IRE1b, bZIP60 mRNA, and on regulated IRE1-dependent decay of target genes. However, root and shoot growth in the absence of stress was independent of bZIP60. Blocking both arms of the UPR signaling pathway in a triple ire1a ire1b bzip28 mutant was lethal, impacting pollen viability under unstressed conditions. Complementation with IRE1b constructs showed that both the PK and RNase domains are required for normal gametophyte development, but bZIP60 is not. Hence, the UPR plays a critical role in stress tolerance, and in normal vegetative growth and reproductive development in plants.T he plant unfolded protein response (UPR) is important in protecting plants from environmental stress. Adverse environmental conditions can interfere with sensitive biosynthetic processes in plants such as protein folding. The UPR has been subject of many recent reviews (for example, see ref. 1) and results from the accumulation of misfolded proteins in the endoplasmic reticulum (ER). The UPR signaling pathway in plants consists of two "arms," one involving membrane associated transcription factors such as basic leucine zipper transcription factor 17 and 28 (bZIP17 and bZIP28) and another arm involving Inositol requiring enzyme1 (IRE1) (2). IRE1 is a dual PK/ribonuclease that is conserved from yeast to man. In response to stress, IRE1 is activated and splices a specific target mRNA in the cytoplasm that encodes a stress-response transcription factor.IRE1 is a single-pass transmembrane protein in the ER membrane with its N terminus facing the ER lumen, serving as a sensor domain. The C terminus of the protein faces the cytosol and contains the PK and ribonuclease domains. Upon activation, IRE1 dimerizes or oligomerizes and undergoes transphosphorylation and activation of its ribonuclease domain (3, 4). Splicing involves cleavage in each of two loops of the substrate RNA, removal...

Section: Discussionsupporting

confidence: 82%

Section: Significancementioning

confidence: 99%

See 1 more Smart Citation

Protein kinase and ribonuclease domains of IRE1 confer stress tolerance, vegetative growth, and reproductive development in Arabidopsis

Deng

Srivastava

Howell

2013

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118

189

“…Activation mechanism of XBP1 is similar to that of Hac1 in yeast in which IRE1-regulated unconventional splicing results in an ORF shift and elimination of the transmembrane domain, enabling the nuclear localization of the newly encoded XBP1 (29,30). Using the model plant Arabidopsis, two ER membrane-associated transcription factors, bZIP28 and bZIP60, were identified as the plant counterparts of mamalian ATF6 and XBP1, which are activated through the regulated intramembrane proteolysis and unconventional splicing in response to ER stress, respectively (31)(32)(33). In general, when the membrane-associated transcription factors are activated, they recruit general transcription factors (GTFs) and RNA polymerase II (RNAPII) machinery (34) to the promoters of ER stress-responsive genes during transcription activation.…”

Section: Significancementioning

confidence: 99%

Transcription factor interaction with COMPASS-like complex regulates histone H3K4 trimethylation for specific gene expression in plants

Song

Sun

et al. 2015

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104

Accumulation of unfolded or misfolded proteins causes endoplasmic reticulum (ER) stress, which activates a set of ER membraneassociated transcription factors for protein homeostasis regulation. Previous genome-wide chromatin immunoprecipitation analysis shows a strong correlation between histone H3K4 trimethylation (H3K4me3) and active gene expression. However, how the histone modification complex is specifically and timely recruited to the active promoters remains unknown. Using ER stress responsive gene expression as a model system, we demonstrate that sequencespecific transcription factors interact with COMPASS-like components and affect H3K4me3 formation at specific target sites in Arabidopsis. Gene profiling analysis reveals that membrane-associated basic leucine zipper (bZIP) transcription factors bZIP28 and bZIP60 regulate most of the ER stress responsive genes. Loss-offunctions of bZIP28 and bZIP60 impair the occupancy of H3K4me3 on promoter regions of ER stress responsive genes. Further, in vitro pull-down assays and in vivo bimolecular fluorescence complementation (BiFC) experiments show that bZIP28 and bZIP60 interact with Ash2 and WDR5a, both of which are core COMPASS-like components. Knockdown expression of either Ash2 or WDR5a decreased the expression of several ER stress responsive genes. The COMPASSlike complex is known to interact with histone methyltransferase to facilitate preinitiation complex (PIC) assembly and generate H3K4me3 during transcription elongation. Thus, our data shows that the ER stress stimulus causes the formation of PIC and deposition of H3K4me3 mark at specific promoters through the interaction between transcription factor and COMPASS-like components.COMPASS-like | ER stress | H3K4 trimethylation | transcription factor | unfolded protein response

“…In addition to Tm and DTT, heat is known to induce the cytoplasmic splicing of bZIP60 (9). Therefore, the profiles of BiP3, bZIP60s, PR-4, PRX34, and AT1G78850 after Tm, DTT, and heat treatment were monitored by qPCR in WT and ire1a/b.…”

Section: Modulated Regulation Of Cytoplasmic Splicing and Ridd In Difmentioning

confidence: 99%

Defects in IRE1 enhance cell death and fail to degrade mRNAs encoding secretory pathway proteins in the Arabidopsis unfolded protein response

Mishiba

Nagashima

Suzuki

et al. 2013

169

167

The unfolded protein response (UPR) is a cellular response highly conserved in eukaryotes to obviate accumulation of misfolded proteins in the endoplasmic reticulum (ER). Inositol-requiring enzyme 1 (IRE1) catalyzes the cytoplasmic splicing of mRNA encoding bZIP transcription factors to activate the UPR signaling pathway. Arabidopsis IRE1 was recently shown to be involved in the cytoplasmic splicing of bZIP60 mRNA. In the present study, we demonstrated that an Arabidopsis mutant with defects in two IRE1 paralogs showed enhanced cell death upon ER stress compared with a mutant with defects in bZIP60 and wild type, suggesting an alternative function of IRE1 in the UPR. Analysis of our previous microarray data and subsequent quantitative PCR indicated degradation of mRNAs encoding secretory pathway proteins by tunicamycin, DTT, and heat in an IRE1-dependent manner. The degradation of mRNAs localized to the ER during the UPR was considered analogous to a molecular mechanism referred to as the regulated IRE1-dependent decay of mRNAs reported in metazoans. Another microarray analysis conducted in the condition repressing transcription with actinomycin D and a subsequent Gene Set Enrichment Analysis revealed the regulated IRE1-dependent decay of mRNAs-mediated degradation of a significant portion of mRNAs encoding the secretory pathway proteins. In the mutant with defects in IRE1, genes involved in the cytosolic protein response such as heat shock factor A2 were upregulated by tunicamycin, indicating the connection between the UPR and the cytosolic protein response.heat stress | protein folding | bioinformatics T he unfolded protein response (UPR) or the endoplasmic reticulum (ER) stress response is a cellular response that is highly conserved in eukaryotes to obviate accumulation of misfolded proteins and to alleviate protein overload in the ER (1-3). Inositol-requiring enzyme 1 (IRE1), which is the primary transducer of the UPR in various organisms, catalyzes the unconventional or cytoplasmic splicing of mRNAs encoding bZIP transcription factors to up-regulate the UPR-related genes, such as genes for the ER-resident molecular chaperones, through its ribonuclease domain. The cytoplasmic splicing by IRE1 activates the bZIP transcription factors HAC1, XBP1, and bZIP60 in yeast, animals, and plants, respectively, by producing active proteins. Although the fundamental mechanism of the cytoplasmic splicing by IRE1 appears to be highly conserved, the mechanism of transcription factor activation is diverse among organisms (4).In addition to the cytoplasmic splicing of mRNAs for transcription factors, other functions of metazoan IRE1 have been reported. One such function is the degradation of mRNAencoding proteins in the secretory pathway referred to as regulated IRE1-dependent decay (RIDD) (5-7). RIDD is considered to contribute to reducing the amount of proteins entering the ER in the UPR. The metazoan UPR has an alternative mechanism to reduce the amount of protein entering the ER, and this mechanism is regulated by PKR-li...