Arabidopsis IRE1 catalyses unconventional splicing of bZIP60 mRNA to produce the active transcription factor

IRE1 plays an essential role in the endoplasmic reticulum (ER) stress response in yeast and mammals. We found that a double mutant of Arabidopsis IRE1A and IRE1B (ire1a/ire1b) is more sensitive to the ER stress inducer tunicamycin than the wild-type. Transcriptome analysis revealed that genes whose induction was reduced in ire1a/ire1b largely overlapped those in the bzip60 mutant. We observed that the active form of bZIP60 protein detected in the wild-type was missing in ire1a/ire1b. We further demonstrated that bZIP60 mRNA is spliced by ER stress, removing 23 ribonucleotides and therefore causing a frameshift that replaces the C-terminal region of bZIP60 including the transmembrane domain (TMD) with a shorter region without a TMD. This splicing was detected in ire1a and ire1b single mutants, but not in the ire1a/ire1b double mutant. We conclude that IRE1A and IRE1B catalyse unconventional splicing of bZIP60 mRNA to produce the active transcription factor

Alteration of the bZIP60/IRE1 Pathway Affects Plant Response to ER Stress in Arabidopsis thaliana

The Unfolded Protein Response (UPR) is elicited under cellular and environmental stress conditions that disrupt protein folding in the endoplasmic reticulum (ER). Through the transcriptional induction of genes encoding ER resident chaperones and proteins involved in folding, the pathway contributes to alleviating ER stress by increasing the folding capacity in the ER. Similarly to other eukaryotic systems, one arm of the UPR in Arabidopsis is set off by a non-conventional splicing event mediated by ribonuclease kinase IRE1b. The enzyme specifically targets mature bZIP60 RNA for cleavage, which results in a novel splice variant encoding a nuclear localized transcription factor. Although it is clear that this molecular switch widely affects the transcriptome, its exact role in overall plant response to stress has not been established and mutant approaches have not provided much insight. In this study, we took a transgenic approach to manipulate the pathway in positive and negative fashions. Our data show that the ER-resident chaperone BiP accumulates differentially depending on the level of activation of the pathway. In addition, phenotypes of the transgenic lines suggest that BiP accumulation is positively correlated with plant tolerance to chronic ER stress

IRE1/bZIP60-Mediated Unfolded Protein Response Plays Distinct Roles in Plant Immunity and Abiotic Stress Responses

Endoplasmic reticulum (ER)-mediated protein secretion and quality control have been shown to play an important role in immune responses in both animals and plants. In mammals, the ER membrane-located IRE1 kinase/endoribonuclease, a key regulator of unfolded protein response (UPR), is required for plasma cell development to accommodate massive secretion of immunoglobulins. Plant cells can secrete the so-called pathogenesis-related (PR) proteins with antimicrobial activities upon pathogen challenge. However, whether IRE1 plays any role in plant immunity is not known. Arabidopsis thaliana has two copies of IRE1, IRE1a and IRE1b. Here, we show that both IRE1a and IRE1b are transcriptionally induced during chemically-induced ER stress, bacterial pathogen infection and treatment with the immune signal salicylic acid (SA). However, we found that IRE1a plays a predominant role in the secretion of PR proteins upon SA treatment. Consequently, the ire1a mutant plants show enhanced susceptibility to a bacterial pathogen and are deficient in establishing systemic acquired resistance (SAR), whereas ire1b is unaffected in these responses. We further demonstrate that the immune deficiency in ire1a is due to a defect in SA- and pathogen-triggered, IRE1-mediated cytoplasmic splicing of the bZIP60 mRNA, which encodes a transcription factor involved in the expression of UPR-responsive genes. Consistently, IRE1a is preferentially required for bZIP60 splicing upon pathogen infection, while IRE1b plays a major role in bZIP60 processing upon Tunicamycin (Tm)-induced stress. We also show that SA-dependent induction of UPR-responsive genes is altered in the bzip60 mutant resulting in a moderate susceptibility to a bacterial pathogen. These results indicate that the IRE1/bZIP60 branch of UPR is a part of the plant response to pathogens for which the two Arabidopsis IRE1 isoforms play only partially overlapping roles and that IRE1 has both bZIP60-dependent and bZIP60-independent functions in plant immunity

An Arabidopsis transcription factor, AtbZIP60, regulates the endoplasmic reticulum stress response in a manner unique to plants

Analysis of transcripts of 75 genes encoding putative basic leucine zipper (bZIP) transcription factors in the Arabidopsis genome identified AtbZIP60, which was induced by tunicamycin. AtbZIP60 encodes a predicted protein of 295 aa with a putative transmembrane domain near its C terminus after a bZIP domain. A truncated form of AtbZIP60 without a transmembrane domain (AtbZIP60ΔC) fused with GFP localized to the nucleus, suggesting translocation of native protein to the nucleus by release from the membrane. AtbZIP60 was also induced by DTT and azetidine-2-carboxylate, which induce the endoplasmic reticulum (ER) stress response (also called the unfolded protein response). Expression of AtbZIP60ΔC clearly activated any of three BiP and two calnexin promoters in a dual luciferase assay using protoplasts of cultured cells. The induction was considered to be through cis-elements plant-specific unfolded protein response element and ER stress-response element. Interestingly, AtbZIP60ΔC also appeared to induce the expression of AtbZIP60 through an ER stress-response element-like sequence in the promoter of AtbZIP60. These characteristics of AtbZIP60 imply a signal transduction pathway of the ER stress response unique to plants