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Screen for localized proteins in Caulobacter crescentus.

Russell JH, Keiler KC - PLoS ONE (2008)

Bottom Line: Precise localization of individual proteins is required for processes such as motility, chemotaxis, cell-cycle progression, and cell division in bacteria, but the number of proteins that are localized in bacterial species is not known.The localized hypothetical proteins included one protein that was localized in a helix-like structure, and two proteins for which the localization changed as a function of the cell cycle, suggesting that complex three-dimensional patterns and cell cycle-dependent localization are likely to be common in bacteria.Other mutants produced localized fusion proteins even though the transposon has inserted near the 5' end of a gene, demonstrating that short peptides can contain sufficient information to localize bacterial proteins.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, United States of America.

ABSTRACT
Precise localization of individual proteins is required for processes such as motility, chemotaxis, cell-cycle progression, and cell division in bacteria, but the number of proteins that are localized in bacterial species is not known. A screen based on transposon mutagenesis and fluorescence activated cell sorting was devised to identify large numbers of localized proteins, and employed in Caulobacter crescentus. From a sample of the clones isolated in the screen, eleven proteins with no previously characterized localization in C. crescentus were identified, including six hypothetical proteins. The localized hypothetical proteins included one protein that was localized in a helix-like structure, and two proteins for which the localization changed as a function of the cell cycle, suggesting that complex three-dimensional patterns and cell cycle-dependent localization are likely to be common in bacteria. Other mutants produced localized fusion proteins even though the transposon has inserted near the 5' end of a gene, demonstrating that short peptides can contain sufficient information to localize bacterial proteins. The screen described here could be used in most bacterial species.

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Related in: MedlinePlus

Localization of CC0572 does not depend on the MreB helix.C. crescentus producing CC0572(1–314)-GFP or GFP-MreB were incubated without A22 (top) or with A22 (bottom) for 30 min and imaged by epifluorescence (left panels) and DIC microscopy (right panels). Under these conditions, the GFP-MreB helix was completely disrupted, but the CC0572(1–314)-GFP helix was still present. Scale bars indicating 1 µm are shown.
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pone-0001756-g007: Localization of CC0572 does not depend on the MreB helix.C. crescentus producing CC0572(1–314)-GFP or GFP-MreB were incubated without A22 (top) or with A22 (bottom) for 30 min and imaged by epifluorescence (left panels) and DIC microscopy (right panels). Under these conditions, the GFP-MreB helix was completely disrupted, but the CC0572(1–314)-GFP helix was still present. Scale bars indicating 1 µm are shown.

Mentions: Cell-cycle regulated helical localization patterns have been reported for the actin-like protein MreB in C. crescentus [28]. To determine if CC0572 required MreB for localization, the MreB structure was disrupted using the inhibitor A22 [32]. In cells treated with A22, fluorescence from a GFP-MreB fusion was rapidly delocalized (Fig. 7). The helical pattern produced by CC0572-GFP was unaffected under similar conditions (Fig. 7), indicating that localization is independent of the MreB structure.


Screen for localized proteins in Caulobacter crescentus.

Russell JH, Keiler KC - PLoS ONE (2008)

Localization of CC0572 does not depend on the MreB helix.C. crescentus producing CC0572(1–314)-GFP or GFP-MreB were incubated without A22 (top) or with A22 (bottom) for 30 min and imaged by epifluorescence (left panels) and DIC microscopy (right panels). Under these conditions, the GFP-MreB helix was completely disrupted, but the CC0572(1–314)-GFP helix was still present. Scale bars indicating 1 µm are shown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2262157&req=5

pone-0001756-g007: Localization of CC0572 does not depend on the MreB helix.C. crescentus producing CC0572(1–314)-GFP or GFP-MreB were incubated without A22 (top) or with A22 (bottom) for 30 min and imaged by epifluorescence (left panels) and DIC microscopy (right panels). Under these conditions, the GFP-MreB helix was completely disrupted, but the CC0572(1–314)-GFP helix was still present. Scale bars indicating 1 µm are shown.
Mentions: Cell-cycle regulated helical localization patterns have been reported for the actin-like protein MreB in C. crescentus [28]. To determine if CC0572 required MreB for localization, the MreB structure was disrupted using the inhibitor A22 [32]. In cells treated with A22, fluorescence from a GFP-MreB fusion was rapidly delocalized (Fig. 7). The helical pattern produced by CC0572-GFP was unaffected under similar conditions (Fig. 7), indicating that localization is independent of the MreB structure.

Bottom Line: Precise localization of individual proteins is required for processes such as motility, chemotaxis, cell-cycle progression, and cell division in bacteria, but the number of proteins that are localized in bacterial species is not known.The localized hypothetical proteins included one protein that was localized in a helix-like structure, and two proteins for which the localization changed as a function of the cell cycle, suggesting that complex three-dimensional patterns and cell cycle-dependent localization are likely to be common in bacteria.Other mutants produced localized fusion proteins even though the transposon has inserted near the 5' end of a gene, demonstrating that short peptides can contain sufficient information to localize bacterial proteins.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania, United States of America.

ABSTRACT
Precise localization of individual proteins is required for processes such as motility, chemotaxis, cell-cycle progression, and cell division in bacteria, but the number of proteins that are localized in bacterial species is not known. A screen based on transposon mutagenesis and fluorescence activated cell sorting was devised to identify large numbers of localized proteins, and employed in Caulobacter crescentus. From a sample of the clones isolated in the screen, eleven proteins with no previously characterized localization in C. crescentus were identified, including six hypothetical proteins. The localized hypothetical proteins included one protein that was localized in a helix-like structure, and two proteins for which the localization changed as a function of the cell cycle, suggesting that complex three-dimensional patterns and cell cycle-dependent localization are likely to be common in bacteria. Other mutants produced localized fusion proteins even though the transposon has inserted near the 5' end of a gene, demonstrating that short peptides can contain sufficient information to localize bacterial proteins. The screen described here could be used in most bacterial species.

Show MeSH
Related in: MedlinePlus