The molecular chaperone Hsp90 delivers precursor proteins to the chloroplast import receptor Toc64

Qbadou, Soumya; Becker, Thomas; Mirus, Oliver; Tews, Ivo; Soll, Jürgen; Schleiff, Enrico

doi:10.1038/sj.emboj.7601091

Cited by 154 publications

(212 citation statements)

References 29 publications

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“…Cytosolic Hsp90 has been shown to participate in the posttranslational targeting of preproteins to the TOC-TIC and translocase of the outer membrane-translocase of the inner membrane (TOM-TIM) import machinery of chloroplasts and mitochondria, respectively (30,40). It was recently reported that cytosolic Hsp90 participates in translocation of antigen across the endosomal membrane into the cytosol in mammalian cells and also in dislocation of the cholera toxin A1 subunit from the ER into the cytosol (41,42).…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, previous studies have implicated cytosolic Hsp90 in the targeting of preproteins to the TOC translocon (30), and the use of urea-denatured recombinant pre-SSU-3xFLAG eliminated the possibility that the effect on import was due to inhibition of cytosolic Hsp90 present in the reticulocyte lysate of in vitro translated preproteins (31). Three general stages of import have previously been defined (2,3,(21)(22)(23)(24)(25): energy-independent binding to the TOC receptors, early import intermediates spanning both TOC and TIC translocons formed in the presence of 0.1 mM ATP/GTP, and late import intermediates captured in the presence of high concentrations of ATP (>1 mM) that are fully translocated across the TOC translocon and remain engaged with the TIC complex.…”

Section: Dsp (mentioning

confidence: 99%

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An essential role for chloroplast heat shock protein 90 (Hsp90C) in protein import into chloroplasts

Inoue

Schnell

2013

Proc. Natl. Acad. Sci. U.S.A.

108

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Chloroplast heat shock protein 90 (Hsp90C) represents a highly conserved subfamily of the Hsp90 family of molecular chaperones whose function has not been defined. We identified Hsp90C as a component that interacts with import intermediates of nuclear-encoded preproteins during posttranslational import into isolated chloroplasts. Hsp90C was specifically coprecipitated with a complex of protein import components, including Tic110, Tic40, Toc75, Tic22, and the stromal chaperones, Hsp93 and Hsp70. Radicicol, an inhibitor of Hsp90 ATPase activity, reversibly inhibited the import of a variety of preproteins during translocation across the inner envelope membrane, indicating that Hsp90C functions in membrane translocation into the organelle. Hsp90C is encoded by a single gene in Arabidopsis thaliana, and insertion mutations in the Hsp90C gene are embryo lethal, indicating an essential function for the chaperone in plant viability. On the basis of these results, we propose that Hsp90C functions within a chaperone complex in the chloroplast stroma to facilitate membrane translocation during protein import into the organelle.

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Section: Discussionmentioning

confidence: 99%

Section: Dsp (mentioning

confidence: 99%

An essential role for chloroplast heat shock protein 90 (Hsp90C) in protein import into chloroplasts

Inoue

Schnell

2013

Proc. Natl. Acad. Sci. U.S.A.

108

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“…Although we do not know whether these differences are significant, it is perhaps germane that the substrate for the Table 2 experiments was produced in a wheat germ translation medium, which has been reported to contain additional factors such as 14-3-3 proteins and molecular chaperones (42,47) that can stimulate protein import. Although additional experiments are required to explore this point further, our data are consistent with the view that these factors might serve in some way to increase the efficiency of energy utilization during the import process.…”

Section: Discussionmentioning

confidence: 99%

“…This system yields relatively high amounts of soluble precursor, obviating the need for urea denaturation required of our bacterially expressed precursors before the import experiments. Additionally, precursors synthesized in this manner are added to the import reactions along with chaperones and other elements of the wheat germ translation mix that have been reported to be active in chloroplast import experiments (42). The radiolabeled precursor used in these experiments was GFP preceded by a stromal targeting signal comprising the prSSU transit peptide followed by 22 amino acids of the mature protein (tp22-GFP, Materials and Methods).…”

Section: Effect Of Inhibitors On Intrinsic Background Atpase Activitymentioning

confidence: 99%

Energetic cost of protein import across the envelope membranes of chloroplasts

Shi

Theg

2012

Proc. Natl. Acad. Sci. U.S.A.

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Chloroplasts are the organelles of green plants in which light energy is transduced into chemical energy, forming ATP and reduced carbon compounds upon which all life depends. The expenditure of this energy is one of the central issues of cellular metabolism. Chloroplasts contain ∼3,000 proteins, among which less than 100 are typically encoded in the plastid genome. The rest are encoded in the nuclear genome, synthesized in the cytosol, and posttranslationally imported into the organelle in an energy-dependent process. We report here a measurement of the amount of ATP hydrolyzed to import a protein across the chloroplast envelope membranesonly the second complete accounting of the cost in Gibbs free energy of protein transport to be undertaken. Using two different precursors prepared by three distinct techniques, we show that the import of a precursor protein into chloroplasts is accompanied by the hydrolysis of ∼650 ATP molecules. This translates to a ΔG protein transport of some 27,300 kJ/mol protein imported. We estimate that protein import across the plastid envelope membranes consumes ∼0.6% of the total light-saturated energy output of the organelle. T he majority of proteins in cells are synthesized in the cytoplasm on free or membrane-bound ribosomes. The fraction of proteins that are translocated across one or more cellular membranes has been estimated to be almost 50% in eukaryotic cells (1) and as high as 30% in bacteria (2, 3). Thus, protein movement out of the cytoplasm across membranes represents a considerable cellular activity involving numerous and varied protein transport systems. Protein translocation across membranes is also generally an energy-requiring process, making it a potentially costly activity for cells (4).The nature of the energy inputs to the different protein translocation systems has been extensively studied. We know, for example, that the posttranslational transport of proteins into the endoplasmic reticulum requires ATP hydrolysis alone (4-6), and that the transport of proteins into the thylakoid lumen or across the bacterial plasma membrane on the Tat pathway requires only the transmembrane protonmotive force with no contribution of ATP hydrolysis (7,8). A more complex energy input is required for many other protein transport systems. For instance, the import of proteins into the mitochondrial matrix requires both ATP hydrolysis and the Δψ across the inner mitochondrial membrane (9-11), and bacterial secretion and protein transport to the thylakoid lumen on their respective Sec pathways use ATP hydrolysis and the protonmotive force (3,7,12,13).In contrast to the abundance of work defining the nature of the energy driving protein transporters, few attempts have been made to quantitate the amount of energy required for these reactions. Without this information, it is not possible to assess the amount of metabolic energy that is spent on the cell's considerable protein trafficking activity. One such measurement of the energy required for protein transport is the so-called translo...

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“…In vitro cross-linking and co-immune precipitation experiments with antibodies against Toc64 revealed (Sohrt and Soll , 2000 ;Becker et al , 2004b ). It was later revealed that Toc64 only transiently associates with the TOC core complex (Schleiff et al , 2003b ) and functions in providing a docking site for Hsp90-affiliated preproteins via its TPR domain (Qbadou et al , 2006 ), indicating a function in post-translational protein translocation across the chloroplast outer envelopes. TPR domaincontaining proteins, which are found in all organisms in cellular compartments are known to contribute to posttranslational protein import by the interaction with cytosolic chaperones (Feldheim and Schekman , 1994 ;Young et al , 2003 ;Schlegel et al , 2007 ).…”

Section: The Molecular Framework Of the Toc Complexmentioning

confidence: 99%

The gateway to chloroplast: re-defining the function of chloroplast receptor proteins

Chang¹,

Soll²,

Bölter³

2012

Biological Chemistry

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: Chloroplast biogenesis often requires a tight orchestration between gene expression (both plastidial and nuclear) and translocation of ~ 3000 nuclear-encoded proteins into the organelle. Protein translocation is achieved via two multimeric import machineries at the outer (TOC) and inner (TIC) envelope of chloroplast, respectively. Three components constitute the core element of the TOC complex: a β -barrel protein translocation channel Toc75 and two receptor constituents, Toc159 and Toc34. A diverse set of distinct TOC complexes have recently been characterized and these diversified TOC complexes have evolved to coordinate the translocation of differentially expressed proteins. This review aims to describe the recent discoveries relating to the typical characteristics of these distinct TOC complexes, particularly the receptor constituents, which are the main contributors for TOC complex diversification.

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The molecular chaperone Hsp90 delivers precursor proteins to the chloroplast import receptor Toc64

Cited by 154 publications

References 29 publications

An essential role for chloroplast heat shock protein 90 (Hsp90C) in protein import into chloroplasts

An essential role for chloroplast heat shock protein 90 (Hsp90C) in protein import into chloroplasts

Energetic cost of protein import across the envelope membranes of chloroplasts

The gateway to chloroplast: re-defining the function of chloroplast receptor proteins

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