scholarly journals Co-expressed subunits of dual genetic origin define a conserved supercomplex mediating essential protein import into chloroplasts

2020 ◽  
Author(s):  
Silvia Ramundo ◽  
Yukari Asakura ◽  
Patrice A. Salomé ◽  
Daniela Strenkert ◽  
Morgane Boone ◽  
...  
Keyword(s):  
Ribosome Biogenesis ◽  
Protein Import ◽  
Molecular Composition ◽  
Chloroplast Envelope ◽  
Genetic Origin ◽  
Unfolded Protein ◽  
Photosynthetic Eukaryotes ◽  
Genes Encoding ◽  
Protein Response ◽  
Envelope Membranes

AbstractIn photosynthetic eukaryotes, thousands of proteins are translated in the cytosol and imported into the chloroplast through the concerted action of two translocons — termed TOC and TIC — located in the outer and inner membranes of the chloroplast envelope, respectively. The degree to which the molecular composition of the TOC and TIC complexes is conserved over phylogenetic distances has remained controversial. Here, we combine transcriptomic, biochemical, and genetic tools in the green alga Chlamydomonas (Chlamydomonas reinhardtii) to demonstrate that, despite a lack of evident sequence conservation for some of its components, the algal TIC complex mirrors the molecular composition of a TIC complex from Arabidopsis thaliana. The Chlamydomonas TIC complex contains three nuclear-encoded subunits, Tic20, Tic56 and Tic100, and one chloroplast-encoded subunit, Tic214, and interacts with the TOC complex, as well as with several uncharacterized proteins to form a stable supercomplex (TicToc), indicating that protein import across both envelope membranes is mechanistically coupled. Expression of the nuclear and chloroplast genes encoding both known and the here newly identified TicToc components is highly coordinated, suggesting that a mechanism for regulating its biogenesis across compartmental boundaries must exist. Conditional repression of Tic214, the only chloroplast-encoded subunit in the TicToc complex, impairs the import of chloroplast proteins with essential roles in chloroplast ribosome biogenesis and protein folding and induces a pleiotropic stress response, including several proteins involved in the chloroplast unfolded protein response. These findings underscore the functional importance of the TicToc supercomplex in maintaining chloroplast proteostasis.

scholarly journals Coexpressed subunits of dual genetic origin define a conserved supercomplex mediating essential protein import into chloroplasts

2020 ◽  
Vol 117 (51) ◽  
pp. 32739-32749
Author(s):  
Silvia Ramundo ◽  
Yukari Asakura ◽  
Patrice A. Salomé ◽  
Daniela Strenkert ◽  
Morgane Boone ◽  
...  
Keyword(s):  
Ribosome Biogenesis ◽  
Protein Import ◽  
Molecular Composition ◽  
Chloroplast Envelope ◽  
Genetic Origin ◽  
Unfolded Protein ◽  
Photosynthetic Eukaryotes ◽  
Genes Encoding ◽  
Protein Response ◽  
Envelope Membranes

In photosynthetic eukaryotes, thousands of proteins are translated in the cytosol and imported into the chloroplast through the concerted action of two translocons—termed TOC and TIC—located in the outer and inner membranes of the chloroplast envelope, respectively. The degree to which the molecular composition of the TOC and TIC complexes is conserved over phylogenetic distances has remained controversial. Here, we combine transcriptomic, biochemical, and genetic tools in the green alga Chlamydomonas (Chlamydomonas reinhardtii) to demonstrate that, despite a lack of evident sequence conservation for some of its components, the algal TIC complex mirrors the molecular composition of a TIC complex fromArabidopsis thaliana.The Chlamydomonas TIC complex contains three nuclear-encoded subunits, Tic20, Tic56, and Tic100, and one chloroplast-encoded subunit, Tic214, and interacts with the TOC complex, as well as with several uncharacterized proteins to form a stable supercomplex (TIC-TOC), indicating that protein import across both envelope membranes is mechanistically coupled. Expression of the nuclear and chloroplast genes encoding both known and uncharacterized TIC-TOC components is highly coordinated, suggesting that a mechanism for regulating its biogenesis across compartmental boundaries must exist. Conditional repression of Tic214, the only chloroplast-encoded subunit in the TIC-TOC complex, impairs the import of chloroplast proteins with essential roles in chloroplast ribosome biogenesis and protein folding and induces a pleiotropic stress response, including several proteins involved in the chloroplast unfolded protein response. These findings underscore the functional importance of the TIC-TOC supercomplex in maintaining chloroplast proteostasis.

Mechanism of protein import across the chloroplast envelope

2000 ◽  
Vol 28 (4) ◽  
pp. 485-491 ◽  
Author(s):  
K. Chen ◽  
X. Chen ◽  
D. J. Schnell
Keyword(s):  
Protein Import ◽  
Essential Elements ◽  
Chloroplast Envelope ◽  
Envelope Membrane ◽  
Outer Envelope ◽  
Double Membrane ◽  
Protein Subunits ◽  
Targeting Signals ◽  
Outer Envelope Membrane ◽  
Envelope Membranes

The development and maintenance of chloroplasts relies on the contribution of protein subunits from both plastid and nuclear genomes. Most chloroplast proteins are encoded by nuclear genes and are post-translationally imported into the organelle across the double membrane of the chloroplast envelope. Protein import into the chloroplast consists of two essential elements: the specific recognition of the targeting signals (transit sequences) of cytoplasmic preproteins by receptors at the outer envelope membrane and the subsequent translocation of preproteins simultaneously across the double membrane of the envelope. These processes are mediated via the co-ordinate action of protein translocon complexes in the outer (Toe apparatus) and inner (Tic apparatus) envelope membranes.

scholarly journals Transcriptional Levels of Autophagy and UPR Markers in the Brain and Liver of Pregnant Rats

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Saeed Alizadeh ◽  
Ghasem Ghasempour ◽  
Elnaz Golestaneh ◽  
Yasaman Safian Isfahani ◽  
Arya Emami ◽  
...  
Keyword(s):  
Er Stress ◽  
Control Group ◽  
Mrna Levels ◽  
Pregnant Rats ◽  
Unfolded Protein ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Autophagy Pathway ◽  
Protein Response ◽  
The Brain

Background: Pregnancy is associated with oxidative stress that results in endoplasmic reticulum (ER) stress and unfolded protein response (UPR). Prolonged-unalleviated ER stress causes the activation of the autophagy pathway via UPR. Expression of genes encoding glucose-regulated protein 78 (GRP78) and BECLIN1 are induced in UPR and autophagy. Objectives: We studied the mRNA expression of the aforementioned genes in the liver and brain of Nulligravida versus saline and ethanol-treated pregnant rats. Methods: Control pregnant rats were orally treated with normal saline, and test animals received ethanol 250 mg/kg or resveratrol 120 mg/kg from day 1 to day 21 of gestation. Nulligravida rats treated by saline comprised the non-pregnant control group. On day 21, mRNAs encoding GRP78 and BECLIN1 were extracted from the liver and brain tissues and assessed using real-time PCR. Results: Our results showed that the level of transcripts encoding GRP78 and BECLIN1 was higher in the liver of pregnant rats compared to Nulligravida ones. Further, ethanol decreased the mRNA levels of GRP78 and BECLIN1 in the liver of pregnant rats, an effect that was reversed by resveratrol. Levels of GRP78 transcripts were decreased, and those of BECLIN1 remained unchanged in the brain of ethanol exposed pregnant rats. Conclusions: Levels of mRNAs for GRP78 and BECLIN1 are up-regulated during pregnancy. These levels are reduced in the liver of ethanol-treated rats, and resveratrol compensates these effects.

scholarly journals ADD66, a Gene Involved in the Endoplasmic Reticulum-associated Degradation of α-1-Antitrypsin-Z in Yeast, Facilitates Proteasome Activity and Assembly

2007 ◽  
Vol 18 (10) ◽  
pp. 3776-3787 ◽  
Author(s):  
Craig M. Scott ◽  
Kristina B. Kruse ◽  
Béla Z. Schmidt ◽  
David H. Perlmutter ◽  
Ardythe A. McCracken ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Expression System ◽  
20S Proteasome ◽  
Yeast Expression System ◽  
Unfolded Protein ◽  
Endoplasmic Reticulum Associated Degradation ◽  
Antitrypsin Deficiency ◽  
Genes Encoding ◽  
The Unfolded Protein Response ◽  
Protein Response

Antitrypsin deficiency is a primary cause of juvenile liver disease, and it arises from expression of the “Z” variant of the α-1 protease inhibitor (A1Pi). Whereas A1Pi is secreted from the liver, A1PiZ is retrotranslocated from the endoplasmic reticulum (ER) and degraded by the proteasome, an event that may offset liver damage. To better define the mechanism of A1PiZ degradation, a yeast expression system was developed previously, and a gene, ADD66, was identified that facilitates A1PiZ turnover. We report here that ADD66 encodes an ∼30-kDa soluble, cytosolic protein and that the chymotrypsin-like activity of the proteasome is reduced in add66Δ mutants. This reduction in activity may arise from the accumulation of 20S proteasome assembly intermediates or from qualitative differences in assembled proteasomes. Add66p also seems to be a proteasome substrate. Consistent with its role in ER-associated degradation (ERAD), synthetic interactions are observed between the genes encoding Add66p and Ire1p, a transducer of the unfolded protein response, and yeast deleted for both ADD66 and/or IRE1 accumulate polyubiquitinated proteins. These data identify Add66p as a proteasome assembly chaperone (PAC), and they provide the first link between PAC activity and ERAD.

scholarly journals Distinct roles of activating transcription factor 6 (ATF6) and double-stranded RNA-activated protein kinase-like endoplasmic reticulum kinase (PERK) in transcription during the mammalian unfolded protein response

2002 ◽  
Vol 366 (2) ◽  
pp. 585-594 ◽  
Author(s):  
Tetsuya OKADA ◽  
Hiderou YOSHIDA ◽  
Rieko AKAZAWA ◽  
Manabu NEGISHI ◽  
Kazutoshi MORI
Keyword(s):  
Endoplasmic Reticulum ◽  
Transcription Factor ◽  
Protein Kinase ◽  
Er Stress ◽  
Double Stranded Rna ◽  
Unfolded Protein ◽  
Genes Encoding ◽  
Membrane Bound ◽  
Activating Transcription Factor ◽  
Protein Response

In response to accumulation of unfolded proteins in the endoplasmic reticulum (ER), a homoeostatic response, termed the unfolded protein response (UPR), is activated in all eukaryotic cells. The UPR involves only transcriptional regulation in yeast, and approx. 6% of all yeast genes, encoding not only proteins to augment the folding capacity in the ER, but also proteins working at various stages of secretion, are induced by ER stress [Travers, Patil, Wodicka, Lockhart, Weissman and Walter (2000) Cell (Cambridge, Mass.) 101, 249–258]. In the present study, we conducted microarray analysis of HeLa cells, although our analysis covered only a small fraction of the human genome. A great majority of human ER stress-inducible genes (approx. 1% of 1800 genes examined) were classified into two groups. One group consisted of genes encoding ER-resident molecular chaperones and folding enzymes, and these genes were directly regulated by the ER-membrane-bound transcription factor activating transcription factor (ATF) 6. The ER-membrane-bound protein kinase double-stranded RNA-activated protein kinase-like ER kinase (PERK)-mediated signalling pathway appeared to be responsible for induction of the remaining genes, which are not involved in secretion, but may be important after cellular recovery from ER stress. In higher eukaryotes, the PERK-mediated translational-attenuation system is known to operate in concert with the transcriptional-induction system. Thus we propose that mammalian cells have evolved a strategy to cope with ER stress different from that of yeast cells.

scholarly journals UPRmt scales mitochondrial network expansion with protein synthesis via mitochondrial import

2020 ◽  
Author(s):  
Tomer Shpilka ◽  
YunGuang Du ◽  
Qiyun Yang ◽  
Andrew Melber ◽  
Nandhitha U. Naresh ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Protein Synthesis ◽  
Protein Import ◽  
Dependent Manner ◽  
Mitochondrial Network ◽  
Mitochondrial Import ◽  
Network Expansion ◽  
Unfolded Protein ◽  
Protein Response ◽  
Mitochondrial Unfolded Protein Response

AbstractAs organisms develop, individual cells generate mitochondria to fulfill physiologic requirements. However, it remains unknown how mitochondrial network expansion is scaled to cell growth and impacted by environmental cues. The mitochondrial unfolded protein response (UPRmt) is a signaling pathway mediated by the transcription factor ATFS-1 which harbors a mitochondrial targeting sequence (MTS)1. Here, we demonstrate that ATFS-1 mediates an adaptable mitochondrial expansion program that is active throughout normal development. Developmental mitochondrial network expansion required the relatively inefficient MTS2 in ATFS-1, which allowed the transcription factor to be responsive to parameters that impact protein import capacity of the entire mitochondrial network. Increasing the strength of the ATFS-1 MTS impaired UPRmt activity throughout development due to increased accumulation within mitochondria. The insulin-like signaling-TORC13 and AMPK pathways affected UPRmt activation4,5 in a manner that correlated with protein synthesis. Manipulation to increase protein synthesis caused UPRmt activation. Alternatively, S6 kinase inhibition had the opposite effect due to increased mitochondrial accumulation of ATFS-1. However, ATFS-1 with a dysfunctional MTS6 constitutively increased UPRmt activity independent of TORC1 function. Lastly, expression of a single protein with a strong MTS, was sufficient to expand the muscle cell mitochondrial network in an ATFS-1-dependent manner. We propose that mitochondrial network expansion during development is an emergent property of the synthesis of highly expressed mitochondrial proteins that exclude ATFS-1 from mitochondrial import, causing UPRmt activation. Mitochondrial network expansion is attenuated once ATFS-1 can be imported.

scholarly journals 12h-clock control of central dogma information flow by XBP1s

2019 ◽  
Author(s):  
Yinghong Pan ◽  
Heather Ballance ◽  
Huan Meng ◽  
Naomi Gonzalez ◽  
Clifford C. Dacso ◽  
...  
Keyword(s):  
Information Flow ◽  
Ribosome Biogenesis ◽  
Marine Animals ◽  
Central Dogma ◽  
Unfolded Protein ◽  
Evolutionarily Conserved ◽  
A Cell ◽  
Protein Response

ABSTRACTOur group recently discovered a cell-autonomous mammalian 12h-clock regulating physiological unfolded protein response. Xbp1s ablation impairs 12h-transcript oscillations in vitro, and we now show liver-specific deletion of XBP1s globally impaired murine 12h-transcriptome, but not the circadian rhythms in vivo. XBP1s-dependent 12h-transcriptome is enriched for transcription, mRNA processing, ribosome biogenesis, translation, and protein ER-Golgi processing/sorting in a temporal order consistent with the progressive molecular processing sequence described by the central dogma information flow (CEDIF). The 12h-rhythms of CEDIF are cell-autonomous and evolutionarily conserved in circatidal marine animals. Mechanistically, we found the motif stringency of promoter XBP1s binding sites, but not necessarily XBP1s expression, dictates its ability to drive 12h-rhythms of transcription and further identified GABP as putative novel transcriptional regulator of 12h-clock. We hypothesize the 12h-rhythms of CEDIF allows rush hours’ gene expression and processing, with the particular genes processed at each rush hour regulated by circadian and/or tissue specific pathways.

scholarly journals Identification of intermediates in the pathway of protein import into chloroplasts and their localization to envelope contact sites.

1993 ◽  
Vol 120 (1) ◽  
pp. 103-115 ◽  
Author(s):  
D J Schnell ◽  
G Blobel
Keyword(s):  
Protein Import ◽  
Signal Sequence ◽  
Protein A ◽  
Early Time ◽  
Small Subunit ◽  
Chloroplast Envelope ◽  
Staphylococcal Protein A ◽  
Binding Domains ◽  
Igg Binding ◽  
Envelope Membranes

We have used a hybrid precursor protein to study the pathway of protein import into chloroplasts. This hybrid (pS/protA) consists of the precursor to the small subunit of Rubisco (pS) fused to the IgG binding domains of staphylococcal protein A. The pS/protA is efficiently imported into isolated chloroplasts and is processed to its mature form (S/protA). In addition to the mature stromal form, two intermediates in the pathway of pS/protA import were identified at early time points in the import reaction. The first intermediate represents unprocessed pS/protA bound to the outer surface of the chloroplast envelope and is analogous to a previously characterized form of pS that is specifically bound to the chloroplast surface and can be subsequently translocated in the stroma (Cline, K., M. Werner-Washburne, T. H. Lubben, and K. Keegstra. 1985. J. Biol. Chem. 260:3691-3696.) The second intermediate represents a partially translocated form of the precursor that remains associated with the envelope membrane. This form is processed to mature S/protA, but remains susceptible to exogenously added protease in intact chloroplasts. We conclude that the envelope associated S/protA is spanning both the outer and inner chloroplast membranes en route to the stroma. Biochemical and immunochemical localization of the two translocation intermediates indicates that both forms are exposed at the surface of the outer membrane at sites where the outer and inner membrane are closely apposed. These contact zones appear to be organized in a reticular network on the outer envelope. We propose a model for protein import into chloroplasts that has as its central features two distinct protein conducting channels in the outer and inner envelope membranes, each gated open by a distinct subdomain of the pS signal sequence.

IRE1 and efferent signaling from the endoplasmic reticulum

2000 ◽  
Vol 113 (21) ◽  
pp. 3697-3702 ◽  
Author(s):  
F. Urano ◽  
A. Bertolotti ◽  
D. Ron
Keyword(s):  
Gene Expression ◽  
Endoplasmic Reticulum ◽  
Transcription Factor ◽  
Er Stress ◽  
Signaling Pathway ◽  
Unfolded Protein ◽  
Genes Encoding ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Stress Signaling Pathway

Genetic analysis of the cellular adaptation to malfolded proteins in the endoplasmic reticulum (the unfolded protein response - UPR) has revealed a novel signaling pathway initiated by activation of IRE1, an ER-resident protein kinase and endonuclease. In yeast, Ire1p activates gene expression by promoting a non-conventional splicing event that converts the mRNA encoding the Hac1p transcription factor from an inefficiently translated inactive mRNA to an actively translated one. Hac1p binds to the promoters of genes encoding chaperones and other targets of the UPR and activates them. Recently, mammalian IRE1 homologues have been identified and their response to ER stress is regulated by binding to the ER chaperone BiP. The mechanisms by which mammalian IRE1 activates gene expression have not been completely characterized and mammalian HAC1 homologues have not been identified. Surprisingly, mammalian IRE1s are able to activate both JUN N-terminal kinases and an alternative ER-stress signaling pathway mediated by the transcription factor ATF6. This indicates that the mammalian UPR is more complex than that found in yeast.

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