Limits...
Complete maturation of the plastid protein translocation channel requires a type I signal peptidase.

Inoue K, Baldwin AJ, Shipman RL, Matsui K, Theg SM, Ohme-Takagi M - J. Cell Biol. (2005)

Bottom Line: Next, we show that disruption of a gene encoding plastidic SPase I (Plsp1) resulted in the accumulation of immature forms of Toc75, severe reduction of plastid internal membrane development, and a seedling lethal phenotype.These phenotypes were rescued by the overexpression of Plsp1 complementary DNA.Plsp1 appeared to be targeted both to the envelope and to the thylakoidal membranes; thus, it may have multiple functions.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Sciences, College of Agricultural and Environmental Sciences, University of California, Davis, CA 95616, USA. kinoue@ucdavis.edu

ABSTRACT
The protein translocation channel at the plastid outer envelope membrane, Toc75, is essential for the viability of plants from the embryonic stage. It is encoded in the nucleus and is synthesized with a bipartite transit peptide that is cleaved during maturation. Despite its important function, the molecular mechanism and the biological significance of the full maturation of Toc75 remain unclear. In this study, we show that a type I signal peptidase (SPase I) is responsible for this process. First, we demonstrate that a bacterial SPase I converted Toc75 precursor to its mature form in vitro. Next, we show that disruption of a gene encoding plastidic SPase I (Plsp1) resulted in the accumulation of immature forms of Toc75, severe reduction of plastid internal membrane development, and a seedling lethal phenotype. These phenotypes were rescued by the overexpression of Plsp1 complementary DNA. Plsp1 appeared to be targeted both to the envelope and to the thylakoidal membranes; thus, it may have multiple functions.

Show MeSH

Related in: MedlinePlus

A bacterial SPase I can convert Toc75 precursor to its mature form. Radiolabeled precursor (pr75; lane 1; 10% input) was converted to the intermediate (i75) and mature (m75) forms by chloroplast protein import assay in vitro (lane 2). When pr75 or i75 was incubated with Lep, the production of m75 was detected, whereas m75 was not processed (lanes 3, 6, and 9, respectively). Heat-denatured Lep was used as a negative control (lanes 4, 7, and 10). The black line indicates grouping of images of different exposures.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2171254&req=5

fig1: A bacterial SPase I can convert Toc75 precursor to its mature form. Radiolabeled precursor (pr75; lane 1; 10% input) was converted to the intermediate (i75) and mature (m75) forms by chloroplast protein import assay in vitro (lane 2). When pr75 or i75 was incubated with Lep, the production of m75 was detected, whereas m75 was not processed (lanes 3, 6, and 9, respectively). Heat-denatured Lep was used as a negative control (lanes 4, 7, and 10). The black line indicates grouping of images of different exposures.

Mentions: As a first step to examine whether SPase I is responsible for the complete maturation of Toc75, we incubated the radiolabeled Toc75 precursor with an SPase I from Escherichia coli, leader peptidase (Lep). As shown in Fig. 1, the active but not heat-denatured Lep converted the Toc75 precursor to its mature form regardless of the presence of the first part of the transit peptide (lanes 3, 4, 6, and 7). This finding prompted us to seek SPases I in the model plant A. thaliana as candidates for the peptidase that is responsible for full maturation of Toc75. There are three SPases I predicted to be located in the plastid (Inoue et al., 2005). One of them, At2g30440, has already been identified as the thylakoidal processing peptidase (Chaal et al., 1998). The second protein, At1g06870, has not been identified in any proteome libraries, but its gene expression is evidenced by the presence of multiple cDNA clones in the database. Finally, the presence of the third protein, At3g24590, in chloroplasts had been confirmed by multidimensional chromatography (Kleffmann et al., 2004). We named this protein Plsp1 for plastidic type I signal peptidase 1 and decided to focus our research efforts on this protein.


Complete maturation of the plastid protein translocation channel requires a type I signal peptidase.

Inoue K, Baldwin AJ, Shipman RL, Matsui K, Theg SM, Ohme-Takagi M - J. Cell Biol. (2005)

A bacterial SPase I can convert Toc75 precursor to its mature form. Radiolabeled precursor (pr75; lane 1; 10% input) was converted to the intermediate (i75) and mature (m75) forms by chloroplast protein import assay in vitro (lane 2). When pr75 or i75 was incubated with Lep, the production of m75 was detected, whereas m75 was not processed (lanes 3, 6, and 9, respectively). Heat-denatured Lep was used as a negative control (lanes 4, 7, and 10). The black line indicates grouping of images of different exposures.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171254&req=5

fig1: A bacterial SPase I can convert Toc75 precursor to its mature form. Radiolabeled precursor (pr75; lane 1; 10% input) was converted to the intermediate (i75) and mature (m75) forms by chloroplast protein import assay in vitro (lane 2). When pr75 or i75 was incubated with Lep, the production of m75 was detected, whereas m75 was not processed (lanes 3, 6, and 9, respectively). Heat-denatured Lep was used as a negative control (lanes 4, 7, and 10). The black line indicates grouping of images of different exposures.
Mentions: As a first step to examine whether SPase I is responsible for the complete maturation of Toc75, we incubated the radiolabeled Toc75 precursor with an SPase I from Escherichia coli, leader peptidase (Lep). As shown in Fig. 1, the active but not heat-denatured Lep converted the Toc75 precursor to its mature form regardless of the presence of the first part of the transit peptide (lanes 3, 4, 6, and 7). This finding prompted us to seek SPases I in the model plant A. thaliana as candidates for the peptidase that is responsible for full maturation of Toc75. There are three SPases I predicted to be located in the plastid (Inoue et al., 2005). One of them, At2g30440, has already been identified as the thylakoidal processing peptidase (Chaal et al., 1998). The second protein, At1g06870, has not been identified in any proteome libraries, but its gene expression is evidenced by the presence of multiple cDNA clones in the database. Finally, the presence of the third protein, At3g24590, in chloroplasts had been confirmed by multidimensional chromatography (Kleffmann et al., 2004). We named this protein Plsp1 for plastidic type I signal peptidase 1 and decided to focus our research efforts on this protein.

Bottom Line: Next, we show that disruption of a gene encoding plastidic SPase I (Plsp1) resulted in the accumulation of immature forms of Toc75, severe reduction of plastid internal membrane development, and a seedling lethal phenotype.These phenotypes were rescued by the overexpression of Plsp1 complementary DNA.Plsp1 appeared to be targeted both to the envelope and to the thylakoidal membranes; thus, it may have multiple functions.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Sciences, College of Agricultural and Environmental Sciences, University of California, Davis, CA 95616, USA. kinoue@ucdavis.edu

ABSTRACT
The protein translocation channel at the plastid outer envelope membrane, Toc75, is essential for the viability of plants from the embryonic stage. It is encoded in the nucleus and is synthesized with a bipartite transit peptide that is cleaved during maturation. Despite its important function, the molecular mechanism and the biological significance of the full maturation of Toc75 remain unclear. In this study, we show that a type I signal peptidase (SPase I) is responsible for this process. First, we demonstrate that a bacterial SPase I converted Toc75 precursor to its mature form in vitro. Next, we show that disruption of a gene encoding plastidic SPase I (Plsp1) resulted in the accumulation of immature forms of Toc75, severe reduction of plastid internal membrane development, and a seedling lethal phenotype. These phenotypes were rescued by the overexpression of Plsp1 complementary DNA. Plsp1 appeared to be targeted both to the envelope and to the thylakoidal membranes; thus, it may have multiple functions.

Show MeSH
Related in: MedlinePlus