The use of transposon TnphoA to detect genes for cell envelope proteins subject to a common regulatory stimulus

Gutierrez, Claude; Barondess, James J.; Manoil, Colin; Beckwith, Jon

doi:10.1016/0022-2836(87)90650-4

Cited by 208 publications

(151 citation statements)

References 33 publications

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“…Isocitrate dehydrogenase (EC 1.1.1.41), a cytoplasmic enzyme in M. xanthus (25), was assayed based on a procedure by Watson and Dworkin (25), using 4 (4)(5)(6)(7)16). In these systems, 1-2% of the TnphoA insertion mutants exhibited PhoA enzyme activity (7,16). Since all of the PhoA substrates we tested were inhibitory to the growth of M. xanthus, we devised an in vitro scheme to identify mutants that produced enzymatically active PhoA fusion proteins.…”

Section: Methodsmentioning

confidence: 99%

Transposon tagging of genes for cell-cell interactions in Myxococcus xanthus.

Kalos¹,

Zissler²

1990

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

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The prokaryote Myxococcus xanthus is a model for cell interactions important in multicellular behavior. We used the transposon TnphoA to specifically identify genes for cell-surface factors involved in cell interactions. From a library of 10,700 insertions of TnphoA, we isolated 36 that produced alkaline phosphatase activity. Three TnphoA insertions tagged cell motility genes, called cgi, which control the adventurous movement of cells. The products of the tagged cgl genes could function in trans upon other cells and were localized primarily in the cell envelope and extracellular space, consistent with TnphoA tagging genes for extracellular factors controlling motility.Multicellular organisms are dependent on many molecules to communicate messages between cells. These molecules could include developmental factors guiding cell migration, growth factors stimulating cell proliferation and maturation,

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Section: Methodsmentioning

confidence: 99%

Transposon tagging of genes for cell-cell interactions in Myxococcus xanthus.

Kalos¹,

Zissler²

1990

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

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“…100% survival is defined as the absorbance of each strain after 180 min of growth in LB medium without CuCl 2 . The LD 50 is defined as the concentration of CuCl 2 that killed 50% of the cells after 15 h of growth.…”

Section: Novel Phenotypes Of E Coli Tat Mutants 47546mentioning

confidence: 99%

Novel Phenotypes of Escherichia coli tat Mutants Revealed by Global Gene Expression and Phenotypic Analysis

Ize

Porcelli

Lucchini

et al. 2004

Journal of Biological Chemistry

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The Tat protein export system serves to export folded proteins harboring an N-terminal twin arginine signal peptide across the cytoplasmic membrane. In this study, we have used gene expression profiling of Escherichia coli supported by phenotypic analysis to investigate how cells respond to a defect in the Tat pathway. Previous work has demonstrated that strains mutated in genes encoding essential Tat pathway components are defective in the integrity of their cell envelope because of the mislocalization of two amidases involved in cell wall metabolism (Ize, B., Stanley, N. R., Buchanan, G., and Palmer, T. (2003) Mol. Microbiol. 48, 1183-1193). To distinguish between genes that are differentially expressed specifically because of the cell envelope defect and those that result from other effects of the tatC deletion, we also analyzed two different transposon mutants of the ⌬tatC strain that have their outer membrane integrity restored. Approximately 50% of the genes that were differentially expressed in the tatC mutant are linked to the envelope defect, with the products of many of these genes involved in self-defense or protection mechanisms, including the production of exopolysaccharide. Among the changes that were not explicitly linked to envelope integrity, we characterized a role for the Tat system in iron acquisition and copper homeostasis. Finally, we have demonstrated that overproduction of the Tat substrate SufI saturates the Tat translocon and produces effects on global gene expression that are similar to those resulting from the ⌬tatC mutation.

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“…EXPERIMENTAL 2.1. Bacierial strain, plasmids, bacteriophage and medium E. co/iCC118 [5], plasmids pUC18-1.6cenA [6] and pB-1 [5], and bacteriophage hTnphoA-1 [7] were described previously. Cultures were grown in LB medium [8].…”

Section: Introduction Endoglucanasementioning

confidence: 99%

Fusion to an endoglucanase allows alkaline phosphatase to bind to cellulose

et al. 1989

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Endoglucanase CenA of Cellulomonas fimi comprises an N‐terminal cellulose‐binding domain and a C‐terminal catalytic domain joined together by a sequence of 23 proline and threonine residues (the Pro‐Thr box). The domains function independently when separated by proteolysis. TnphoA has been used to generate cenA′‐′phoA fusions. CenA′‐′PhoA fusion polypeptides which contain the entire cellulose‐binding domain of CenA bind to cellulose, allowing their purification from periplasmic extracts in a single, facile step. This result has implications for purification or immobilisation of chimeric proteins on a cheap cellulose matrix.

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The use of transposon TnphoA to detect genes for cell envelope proteins subject to a common regulatory stimulus

Cited by 208 publications

References 33 publications

Transposon tagging of genes for cell-cell interactions in Myxococcus xanthus.

Transposon tagging of genes for cell-cell interactions in Myxococcus xanthus.

Novel Phenotypes of Escherichia coli tat Mutants Revealed by Global Gene Expression and Phenotypic Analysis

Fusion to an endoglucanase allows alkaline phosphatase to bind to cellulose

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