**2. ER stress response: The cytoprotective unfolded protein response**

The ER is a highly dynamic organelle, which mediates several cellular functions, such as the folding and post-translational modification of secretory proteins and protein quality control in addition to maintaining Ca2+ homeostasis (Schröder 2008). The loading of unfolded pro‐ tein in the lumen of ER for maturation is tightly controlled and dependent on the cellular requirements. Under stress conditions, the folding capacity of the ER can be overloaded causing the accumulation of unfolded proteins and disruption of cellular homeostasis (Xu, Bailly-Maitre & Reed 2005). To cope with this stress condition, eukaryotic cells evolved a so‐ phisticated signaling mechanism referred to as unfolded protein response (UPR; Malhotra and Kaufman., 2007). In mammalian cells, the UPR is transduced through three distinct ERtransmembrane sensors: PERK (protein kinase RNA-like ER kinase), Ire1 (inositol-requiring enzyme-1) and the basic leucine zipper transcription factor ATF6 (activating transcription factor-6; Ron and Walter, 2007; Malhotra and Kaufman, 2007, Kapoor and Sanyal, 2009). The activation of the UPR allows the ER processing and folding capacities to be balanced with protein loading into the lumen of the organelle under conditions of ER stress (Malhotra and Kaufman, 2007). This balance is achieved by (i) shutting down protein synthesis via PERK activation, (ii) up-regulating the expression of ER-resident processing proteins, such as mo‐ lecular chaperones and foldases, via activation of Ire1 and ATF6, and (iii) inducing the ERassociated protein degradation (ERAD) machinery, through activation of Ire1, which mediates the targeting and subsequent degradation of unfolded proteins by the proteosome. However, if the ER stress is sustained, multiple apoptotic pathways can be activated in mammalian cells.

In plants, the UPR seems to operate as a bipartite module, as the ER stress signal is trans‐ duced through homologs of the Ire1 and ATF6 transducers, but a PERK-mediated branch of the UPR has not been shown (Urade 2009; Chen and Brandizzi., 2012). Two components of the Ire1-mediaded branch of the UPR is known. The first one is the Ire1 ortholog that is rep‐ resented by two copies in the Arabidopsis genome, Ire1a and Ire1b, and one copy, OsIre1, in the rice genome. Like the mammalian counterpart, plant Ire1 is associated with the ER mem‐ brane and exhibits ribonuclease activity and autophosphorylation activities, as shown for Ire1a, Ire1b and OsIre1(Koizumi et al., 2001; Okushima et al., 2002). The second component is ER membrane-associated transcription factor bZIP60. Upon ER stress, bZIP60 mRNA is spliced in an IRE1-mediated process to generate an alternatively spliced transcript that lacks the transmembrane domain-encoding sequences (Liu et al., 2007 e 2008 ; Deng et al., 2011; Nagashima et al., 2011). This splicing leads to the synthesis of a soluble and functional bZIP60 transfactor that can be translocated to the nucleus, where it activates ER stress indu‐ cible promoters, such as the BiP3 promoter. Likewise, OsbZIP74 or OsbZIP50 from rice, an ortholog of Arabidopsis AtbZIP60, is regulated through the IRE1-mediated splicing of its RNA to render the activation of ER stress-inducible promoters (Hayashi et al., 2011; Lu et al., 2011).

by the G protein in Arabidopsis, but it remains to be determined whether it operates in soy‐

The ER is a highly dynamic organelle, which mediates several cellular functions, such as the folding and post-translational modification of secretory proteins and protein quality control in addition to maintaining Ca2+ homeostasis (Schröder 2008). The loading of unfolded pro‐ tein in the lumen of ER for maturation is tightly controlled and dependent on the cellular requirements. Under stress conditions, the folding capacity of the ER can be overloaded causing the accumulation of unfolded proteins and disruption of cellular homeostasis (Xu, Bailly-Maitre & Reed 2005). To cope with this stress condition, eukaryotic cells evolved a so‐ phisticated signaling mechanism referred to as unfolded protein response (UPR; Malhotra and Kaufman., 2007). In mammalian cells, the UPR is transduced through three distinct ERtransmembrane sensors: PERK (protein kinase RNA-like ER kinase), Ire1 (inositol-requiring enzyme-1) and the basic leucine zipper transcription factor ATF6 (activating transcription factor-6; Ron and Walter, 2007; Malhotra and Kaufman, 2007, Kapoor and Sanyal, 2009). The activation of the UPR allows the ER processing and folding capacities to be balanced with protein loading into the lumen of the organelle under conditions of ER stress (Malhotra and Kaufman, 2007). This balance is achieved by (i) shutting down protein synthesis via PERK activation, (ii) up-regulating the expression of ER-resident processing proteins, such as mo‐ lecular chaperones and foldases, via activation of Ire1 and ATF6, and (iii) inducing the ERassociated protein degradation (ERAD) machinery, through activation of Ire1, which mediates the targeting and subsequent degradation of unfolded proteins by the proteosome. However, if the ER stress is sustained, multiple apoptotic pathways can be activated in

In plants, the UPR seems to operate as a bipartite module, as the ER stress signal is trans‐ duced through homologs of the Ire1 and ATF6 transducers, but a PERK-mediated branch of the UPR has not been shown (Urade 2009; Chen and Brandizzi., 2012). Two components of the Ire1-mediaded branch of the UPR is known. The first one is the Ire1 ortholog that is rep‐ resented by two copies in the Arabidopsis genome, Ire1a and Ire1b, and one copy, OsIre1, in the rice genome. Like the mammalian counterpart, plant Ire1 is associated with the ER mem‐ brane and exhibits ribonuclease activity and autophosphorylation activities, as shown for Ire1a, Ire1b and OsIre1(Koizumi et al., 2001; Okushima et al., 2002). The second component is ER membrane-associated transcription factor bZIP60. Upon ER stress, bZIP60 mRNA is spliced in an IRE1-mediated process to generate an alternatively spliced transcript that lacks the transmembrane domain-encoding sequences (Liu et al., 2007 e 2008 ; Deng et al., 2011; Nagashima et al., 2011). This splicing leads to the synthesis of a soluble and functional bZIP60 transfactor that can be translocated to the nucleus, where it activates ER stress indu‐ cible promoters, such as the BiP3 promoter. Likewise, OsbZIP74 or OsbZIP50 from rice, an ortholog of Arabidopsis AtbZIP60, is regulated through the IRE1-mediated splicing of its

**2. ER stress response: The cytoprotective unfolded protein response**

A Comprehensive Survey of International Soybean Research - Genetics, Physiology, Agronomy and Nitrogen

bean as well.

Relationships

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mammalian cells.

The second branch of UPR in plants mechanistically resembles the ATF6-mediated transduc‐ tion of the ER stress signal. Upon ER stress, the membrane-associated Arabidopsis ATF6 ho‐ mologs bZIP17 and bZIP28 are relocated to the Golgi, where their transcriptional domains are proteolytically released from the membrane by SP2 (Tajima et al., 2008: Che et al., 2010). The released bZIP domain of these transfactors is then translocated to the nucleus, where it acts in concert with the heterotrimeric NF-Y complex to activate UPR genes (Liu e Howel., 2010). The NF-Y complex is composed the transcriptional factors NF-YA4, NF-YB3 and NF-YC2.

Comprehensive genome-wide evaluations of ER stress-induced changes in gene expression have provided evidence that the UPR operates in a similar fashion in both soybean and Ara‐ bidopsis (Irsigler et al., 2007). Inducers of ER stress, such as tunicamycin and AZC, promote the up-regulation of a class of genes that functions in protein folding and ERAD. In the pro‐ tein folding category, the up-regulated genes include ER-resident molecular chaperones such as BiP, calreticulin, calnexin, and the folding catalyst protein disulfide isomerase (PDI). ERAD-associated genes that are up-regulated by ER stress in soybean include those encod‐ ing polyubiquitin, ubiquitin conjugating enzyme, the alpha subunit of the proteasome, CDC48 and Derlin. These genomic analyses suggested that soybean, like Arabidopsis, have evolved at least two different mechanisms that mediate UPR: (i) transcriptional induction of genes encoding chaperones and vesicle trafficking proteins and (ii) upregulation of the ERassociated protein degradation (ERAD) system for rapid disposal of unfolded proteins in the ER as a protective measure.

In addition to the cytoprotective bipartite response to ER stress in plants, two apparently distinct branches of the ER stress-induced pathways have been shown to transduce a cell death signal: (i) the ER membrane associated Gβ-Gγ heterodimer-mediated signaling events that trigger UPR-associated cell death in *Arabidopsis* (Wang et al., 2007) and (ii) the ER stressinduced NRP-mediated cell death response that has been uncovered in soybean (Reis and Fontes, 2012).
