A genetic selection for supercoiling mutants of Escherichia coli reveals proteins implicated in chromosome structure.

Abstract:

:Chromosomes are divided into topologically independent regions, called domains, by the action of uncharacterized barriers. With the goal of identifying domain barrier components, we designed a genetic selection for mutants with reduced negative supercoiling of the Escherichia coli chromosome. We employed a strain that contained two chromosomally located reporter genes under the control of a supercoiling-sensitive promoter and used transposon mutagenesis to generate a wide range of mutants. We subjected the selected mutants to a series of secondary screens and identified five proteins as modulators of chromosomal supercoiling in vivo. Three of these proteins: H-NS, Fis and DksA, have clear ties to chromosome biology. The other two proteins, phosphoglucomutase (Pgm) and transketolase (TktA), are enzymes involved in carbohydrate metabolism and have not previously been shown to affect DNA. Deletion of any of the identified genes specifically affected chromosome topology, without affecting plasmid supercoiling. We suggest that at least H-NS, Fis and perhaps TktA assist directly in the supercoiling of domains by forming topological barriers on the E. coli chromosome.

journal_name

Mol Microbiol

journal_title

Molecular microbiology

authors

Hardy CD,Cozzarelli NR

doi

10.1111/j.1365-2958.2005.04799.x

subject

Has Abstract

pub_date

2005-09-01 00:00:00

pages

1636-52

issue

6

eissn

0950-382X

issn

1365-2958

pii

MMI4799

journal_volume

57

pub_type

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