Cosmid based mutagenesis causes genetic instability in Streptomyces coelicolor, as shown by targeting of the lipoprotein signal peptidase gene

John T. Munnoch; David A. Widdick; Govind Chandra; Iain C. Sutcliffe; Tracy Palmer; Matthew I. Hutchings

doi:10.1038/srep29495

Cosmid based mutagenesis causes genetic instability in Streptomyces coelicolor, as shown by targeting of the lipoprotein signal peptidase gene

Scientific Reports ◽

10.1038/srep29495 ◽

2016 ◽

Vol 6 (1) ◽

Cited By ~ 3

Author(s):

John T. Munnoch ◽

David A. Widdick ◽

Govind Chandra ◽

Iain C. Sutcliffe ◽

Tracy Palmer ◽

...

Keyword(s):

Genetic Instability ◽

Streptomyces Coelicolor ◽

Signal Peptidase

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Cosmid based mutagenesis causes genetic instability in Streptomyces coelicolor, as shown by targeting of the lipoprotein signal peptidase gene

10.1101/049320 ◽

2016 ◽

Author(s):

John T Munnoch ◽

David A. Widdick ◽

Govind Chandra ◽

Iain C. Sutcliffe ◽

Tracy Palmer ◽

...

Keyword(s):

Cell Division ◽

Gene Duplication ◽

Small Rna ◽

Genetic Instability ◽

Genetic Manipulation ◽

Streptomyces Coelicolor ◽

Extracellular Proteins ◽

Signal Peptidase ◽

Wild Type ◽

Developmental Defects

AbstractBacterial lipoproteins are a class of extracellular proteins tethered to cell membranes by covalently attached lipids. Deleting the lipoprotein signal peptidase (lsp) gene in Streptomyces coelicolor results in growth and developmental defects that cannot be restored by reintroducing the lsp. We report resequencing of the genomes of the wild-type M145 and the cis-complemented Δlsp mutant (BJT1004), mapping and identifying secondary mutations, including an insertion into a novel putative small RNA, scr6809. Disruption of scr6809 led to a range of developmental phenotypes. However, these secondary mutations do not increase the efficiency of disrupting lsp suggesting they are not lsp specific suppressors. Instead we suggest that these were induced by introducing the cosmid St4A10Δlsp as part of the Redirect mutagenesis protocol, which transiently duplicates a number of important cell division genes. Disruption of lsp using no gene duplication resulted in the previously observed phenotype. We conclude that lsp is not essential in S. coelicolor but loss of lsp does lead to developmental defects due to the loss of lipoproteins from the cell. Significantly, our results indicate the use of cosmid libraries for the genetic manipulation of bacteria can lead to unexpected phenotypes not necessarily linked to the gene or pathway of interest.

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Introduction of the Foreign Transposon Tn4560 in Streptomyces coelicolor Leads to Genetic Instability near the Native Insertion Sequence IS1649

Journal of Bacteriology ◽

10.1128/jb.00983-07 ◽

2007 ◽

Vol 189 (24) ◽

pp. 9108-9116 ◽

Cited By ~ 11

Author(s):

Elizabeth M. Widenbrant ◽

Camilla M. Kao

Keyword(s):

Genetic Instability ◽

Type Strain ◽

Insertion Sequence ◽

Structural Changes ◽

Wild Type Strain ◽

Streptomyces Coelicolor ◽

Wild Type ◽

Insertion Elements ◽

Foreign Dna

ABSTRACT We report an altered pattern of genetic instability for Streptomyces coelicolor when the bacterium harbored a foreign transposon, Tn4560. Deletions, amplifications, and circularizations of the linear 8.7-Mb chromosome occurred more frequently at sites adjacent to native insertion elements, notably IS1649. In contrast, deletions, amplifications, and circularizations of a wild-type strain happened at heterogeneous sites within the chromosome. In 50 strains examined, structural changes removed or duplicated hundreds of contiguous S. coelicolor genes, altering up to 33% of the chromosome. S. coelicolor shows a bias toward one type of genetic instability during this particular assault from the environment, the invasion of foreign DNA.

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Streptomyces coelicolor Undergoes Spontaneous Chromosomal End Replacement

Journal of Bacteriology ◽

10.1128/jb.01049-07 ◽

2007 ◽

Vol 189 (24) ◽

pp. 9117-9121 ◽

Cited By ~ 5

Author(s):

Elizabeth M. Widenbrant ◽

Hsiu-Hui Tsai ◽

Carton W. Chen ◽

Camilla M. Kao

Keyword(s):

Genetic Instability ◽

Type Strain ◽

Wild Type Strain ◽

Streptomyces Coelicolor ◽

The Other ◽

Wild Type ◽

Genetic Changes

ABSTRACT We report a previously unobserved form of genetic instability for Streptomyces coelicolor, the replacement of one chromosome end by the other end. These genetic changes occurred spontaneously in both a wild-type strain and strains harboring a foreign transposon. Deleted and duplicated DNA comprises up to 33% of the genome.

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