Further <i>in vivo</i> studies on the role of the molecular chaperone, Hsp93, in plastid protein import

S, Kovacheva; Bédard, Jocelyn; Wardle, Anthony; Patel, Ramesh N.; Jarvis, Paul

doi:10.1111/j.1365-313x.2007.03060.x

Cited by 117 publications

(125 citation statements)

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“…The +/tic110-1, hsp93-V-1, tic40-4, ppi1-1 and hsp93-V-1 hsp93-III-1 (hsp93-V/III) mutants employed to derive the calibration relationships have all been described previously (Jarvis et al 1998;Kovacheva et al 2005;Kovacheva et al 2007). Similarly, the pph-1 and pao1 mutants have been presented in earlier reports (Pruzinska et al 2005;Schelbert et al 2009).…”

Section: Plant Materials and Growth Conditionsmentioning

confidence: 96%

Use of a SPAD-502 meter to measure leaf chlorophyll concentration in Arabidopsis thaliana

2010

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The SPAD-502 meter is a hand-held device that is widely used for the rapid, accurate and non-destructive measurement of leaf chlorophyll concentrations. It has been employed extensively in both research and agricultural applications, with a range of different plant species. However, its utility has not been fully exploited in relation to the most intensively studied model organism for plant science research, Arabidopsis thaliana. Measurements with the SPAD-502 meter produce relative SPAD meter values that are proportional to the amount of chlorophyll present in the leaf. In order to convert these values into absolute units of chlorophyll concentration, calibration curves must be derived and utilized. Here, we present calibration equations for Arabidopsis that can be used to convert SPAD values into total chlorophyll per unit leaf area (nmol/cm 2 ; R 2 = 0.9960) or per unit fresh weight of leaf tissue (nmol/mg; R 2 = 0.9809). These relationships were derived using a series of Arabidopsis chloroplast biogenesis mutants that exhibit chlorophyll deficiencies of varying severity, and were verified by the subsequent analysis of senescent or light-stressed leaves. Our results revealed that the converted SPAD values differ from photometric measurements of solventextracted chlorophyll by just ~6% on average.

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Section: Plant Materials and Growth Conditionsmentioning

confidence: 96%

Use of a SPAD-502 meter to measure leaf chlorophyll concentration in Arabidopsis thaliana

2010

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“…Hsp93 is sufficient to support export of Tic110 or that other chaperones can assist the function of Hsp93 in this conservative sorting pathway. On the other hand, an increased expression rate of Tic110 in these mutants might also cause a seemingly unchanged amount of protein, but the expression level is not documented [16].…”

Section: Resultsmentioning

confidence: 97%

Requirements for a conservative protein translocation pathway in chloroplasts

Vojta¹,

Soll²,

Bölter³

2007

FEBS Letters

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The chloroplast inner envelope translocon subunit Tic110 is imported via a soluble stromal translocation intermediate. In this study an in-organellar import system is established which allows for an accumulation of this intermediate in order to analyze its requirements for reexport. All results demonstrate that the re-export of Tic110 from the soluble intermediate stage into the inner envelope requires ATP hydrolysis, which cannot be replaced by other NTPs. Furthermore, the molecular chaperone Hsp93 seems prominently involved in the reexport pathway of Tic110, because other stromal intermediates like that of the oxygen evolving complex subunit OE33 (iOE33) en route to the thylakoid lumen interacts preferentially with Hsp70.

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“…3A, lanes 3 and 4). The level of Tic40 was analyzed as a loading control, since it has been shown that the protein levels of major Toc/Tic components are not affected in the hsp93V mutant (Constan et al, 2004;Kovacheva et al, 2007). The atHSP93V-DN transgene was also successfully expressed, and the atHsp93 protein level in the atHSP93V-DN transgenic plants, when adding together the amounts of atHsp93V-DN and atHsp93III, was comparable to the total amount of atHsp93 in the wild type (Fig.…”

Section: Athsp93v-dn Cannot Complement the Hsp93v Mutantmentioning

confidence: 99%

“…In chloroplasts, both stromal 93-kD heat shock protein (Hsp93/ClpC) and 70-kD heat shock protein (Hsp70) have been shown to be important for protein import, and both have been proposed to function as the motors that drive protein translocation across the envelope into the stroma Nielsen et al, 1997;Constan et al, 2004;Kovacheva et al, 2005Kovacheva et al, , 2007Shi and Theg, 2010;Su and Li, 2010). However, evidence directly demonstrating the motor function for either chaperone is still lacking.…”

mentioning

confidence: 99%

“…Knockout mutants of hsp93III are indistinguishable from the wild type, while hsp93V mutants are pale green and defective in protein import into chloroplasts (Constan et al, 2004;Kovacheva et al, 2005). 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).…”

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confidence: 99%

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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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Further in vivo studies on the role of the molecular chaperone, Hsp93, in plastid protein import

Cited by 117 publications

References 69 publications

Use of a SPAD-502 meter to measure leaf chlorophyll concentration in Arabidopsis thaliana

Use of a SPAD-502 meter to measure leaf chlorophyll concentration in Arabidopsis thaliana

Requirements for a conservative protein translocation pathway in chloroplasts

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

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