The stroma of higher plant plastids contain ClpP and ClpC, functional homologs of Escherichia coli ClpP and ClpA: an archetypal two-component ATP-dependent protease.

Shanklin, John; DeWitt, Natalie; Flanagan, John M.

doi:10.1105/tpc.7.10.1713

Cited by 162 publications

(147 citation statements)

References 50 publications

(52 reference statements)

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“…Chloroplasts, consistent with their endosymbiotic origin, contain various proteases of bacterial ancestry Clarke et al, 2005). One of the first identified was the ATPdependent Clp protease localized primarily in the stroma (Shanklin et al, 1995). The model for the Ser-type Clp protease has long been the one in Escherichia coli, which consists of a proteolytic core flanked on one or both sides by a HSP100 molecular chaperone.…”

Section: Introductionmentioning

confidence: 99%

Structural and Functional Insights into the Chloroplast ATP-Dependent Clp Protease in Arabidopsis

et al. 2006

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In contrast with the model Escherichia coli Clp protease, the ATP-dependent Clp protease in higher plants has a remarkably diverse proteolytic core consisting of multiple ClpP and ClpR paralogs, presumably arranged within a dual heptameric ring structure. Using antisense lines for the nucleus-encoded ClpP subunit, ClpP6, we show that the Arabidopsis thaliana Clp protease is vital for chloroplast development and function. Repression of ClpP6 produced a proportional decrease in the Clp proteolytic core, causing a chlorotic phenotype in young leaves that lessened upon maturity. Structural analysis of the proteolytic core revealed two distinct subcomplexes that likely correspond to single heptameric rings, one containing the ClpP1 and ClpR1-4 proteins, the other containing ClpP3-6. Proteomic analysis revealed several stromal proteins more abundant in clpP6 antisense lines, suggesting that some are substrates for the Clp protease. A proteolytic assay developed for intact chloroplasts identified potential substrates for the stromal Clp protease in higher plants, most of which were more abundant in young Arabidopsis leaves, consistent with the severity of the chlorotic phenotype observed in the clpP6 antisense lines. The identified substrates all function in more general housekeeping roles such as plastid protein synthesis, folding, and quality control, rather than in metabolic activities such as photosynthesis.

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Section: Introductionmentioning

confidence: 99%

Structural and Functional Insights into the Chloroplast ATP-Dependent Clp Protease in Arabidopsis

et al. 2006

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“…Double knockout of both genes causes lethality, indicating that Hsp93 is essential (Kovacheva et al, 2007). In addition, Hsp93 has been proposed to act as a regulatory subunit of the ClpP protease in chloroplasts (Shanklin et al, 1995;Desimone et al, 1997;Sokolenko et al, 1998;Halperin et al, 2001). Interestingly, although Hsp93 was not detected in the ClpP core complex isolated from chloroplasts, the land plant chloroplast ClpP complex contains two additional peripheral subunits, ClpT1 and ClpT2 (previously named ClpS1 and ClpS2), that have high sequence similarity to the N-terminal portion of Hsp93 (Peltier et al, 2004).…”

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confidence: 99%

The Amino-Terminal Domain of Chloroplast Hsp93 Is Important for Its Membrane Association and Functions in Vivo

Chu

2012

Plant Physiology

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Chloroplast 93-kD heat shock protein (Hsp93/ClpC), an Hsp100 family member, is suggested to have various functions in chloroplasts, including serving as the regulatory chaperone for the ClpP protease in the stroma and acting as a motor component of the protein translocon at the envelope. Indeed, although Hsp93 is a soluble stromal protein, a portion of it is associated with the inner envelope membrane. The mechanism and functional significance of this Hsp93 membrane association have not been determined. Here, we mapped the region important for Hsp93 membrane association by creating various deletion constructs and found that only the construct with the amino-terminal domain deleted, Hsp93-DN, had reduced membrane association. When transformed into Arabidopsis (Arabidopsis thaliana), most atHsp93V-DN proteins did not associate with membranes and atHsp93V-DN failed to complement the pale-green and protein import-defective phenotypes of an hsp93V knockout mutant. The residual atHsp93V-DN at the membranes had further reduced association with the central protein translocon component Tic110. However, the degradation of chloroplast glutamine synthetase, a potential substrate for the ClpP protease, was not affected in the hsp93V mutant or in the atHSP93V-DN transgenic plants. Hsp93-DN also had the same ATPase activity as that of full-length Hsp93. These data suggest that the association of Hsp93 with the inner envelope membrane through its amino-terminal domain is important for the functions of Hsp93 in vivo.Chloroplasts are structurally complex organelles that perform diverse functions (Leister, 2003;Block et al., 2007). They are composed of three membranes, the outer and inner envelope membranes and the thylakoid membranes, and these membranes enclose three aqueous compartments, the intermembrane space, stroma, and thylakoid lumen. Although chloroplasts have their own genome, most chloroplast proteins are encoded by the nuclear genome and are translated in the cytosol as a precursor protein with an N-terminal extension called the transit peptide. Transit peptides direct the import of proteins into chloroplasts through the translocon complex located in the chloroplast envelope. Translocon components of the outer membrane are called Toc (for translocon at the outer envelope membrane of chloroplasts) proteins and those in the inner envelope membrane are called Tic (for translocon at the inner envelope membrane of chloroplasts) proteins. Three Toc components, Toc159, Toc34, and Toc75, form the Toc core complex. Toc159 and Toc34 function as receptors that recognize the precursors targeting to chloroplasts. Toc75 forms a protein-conducting channel across the outer membrane. Three proteins, Tic20, Tic21, and Tic110, have been suggested to function as the channel for precursor translocation across the inner membrane. Tic110 has also been shown to function as the stromal-side receptor for transit peptides and as a scaffold for assembling other stromal translocon components. Tic40 is a cochaperone that coordinates the actions of...

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“…We recently found that a ClpC1 subunit of the chloroplast Clp protease is involved in the regulation of CAO [25]. Since the ClpC1 subunit is a chaperone that recognizes the protease substrate sequences [31,32], it is tempting to speculate that the ClpC1 recognizes the A domain directly in a chlorophyll-b dependent manner. Nevertheless, the involvement of other types of proteases or other factors should not be excluded since mutation in the ClpC1 gene did not abolish the full regulation of the CAO protein levels [25].…”

Section: Discussionmentioning

confidence: 99%

Functional analysis of N-terminal domains of Arabidopsis chlorophyllide a oxygenase

Sakuraba

Yamasato

Tanaka

et al. 2007

Plant Physiology and Biochemistry

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Higher plants acclimate to various light environments by changing the antenna size of a light harvesting photosystem. The antenna size of a photosystem is partly determined by the amount of chlorophyll b in the light-harvesting complexes. Chlorophyllide a oxygenase (CAO) converts chlorophyll a to chlorophyll b in a two-step oxygenation reaction. In our previous study, we demonstrated that the cellular level of the CAO protein controls accumulation of chlorophyll b. We found that the amino acids sequences of CAO in higher plants consist of three domains (A, B, and C domains). The C domain exhibits a catalytic function, and we demonstrated that the combination of the A and B domains regulates the cellular level of CAO. However, the individual function of each of A and B domain has not been determined yet. Therefore, in the present study we constructed a series of deleted CAO sequences that were fused with green fluorescent protein and overexpressed in a chlorophyll b-less mutant of Arabidopsis thaliana, ch1-1, to further dissect functions of A and B domains. Subsequent comparative analyses of the transgenic plants overexpressing B-domain containing proteins and those lacking the B domain determined that there was no significant difference in CAO protein levels. These results indicate that the B domain is not involved in the regulation of the CAO protein levels. Taken together, we concluded that the A domain alone is involved in the regulatory mechanism of the CAO protein levels.

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The stroma of higher plant plastids contain ClpP and ClpC, functional homologs of Escherichia coli ClpP and ClpA: an archetypal two-component ATP-dependent protease.

Cited by 162 publications

References 50 publications

Structural and Functional Insights into the Chloroplast ATP-Dependent Clp Protease in Arabidopsis

Structural and Functional Insights into the Chloroplast ATP-Dependent Clp Protease in Arabidopsis

The Amino-Terminal Domain of Chloroplast Hsp93 Is Important for Its Membrane Association and Functions in Vivo

Functional analysis of N-terminal domains of Arabidopsis chlorophyllide a oxygenase

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