A genome‐wide strategy for the identification of essential genes in Staphylococcus aureus

Summary To address the need for new approaches to antibiotic drug development, we have identified a large number of essential genes for the bacterial pathogen, Staphylococcus aureus, using a rapid shotgun antisense RNA method. Staphylococcus aureus chromosomal DNA fragments were cloned into a xylose...

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Bibliographic Details
Published in:Molecular microbiology 2002-03, Vol.43 (6), p.1387-1400
Main Authors: Forsyth, R. Allyn, Haselbeck, Robert J., Ohlsen, Kari L., Yamamoto, Robert T., Xu, Howard, Trawick, John D., Wall, Daniel, Wang, Liangsu, Brown‐Driver, Vickie, Froelich, Jamie M., C., Kedar G., King, Paula, McCarthy, Melissa, Malone, Cheryl, Misiner, Brian, Robbins, David, Tan, Zehui, Zhu, Zhan‐yang, Carr, Grant, Mosca, Deborah A., Zamudio, Carlos, Foulkes, J. Gordon, Zyskind, Judith W.
Format: Article
Language:English
Subjects:
Anti-Bacterial Agents - pharmacology
Bacterial Proteins - genetics
Bacterial Proteins - metabolism
Cloning, Molecular
Computational Biology - methods
Gene Targeting
Genes, Essential
Genome, Bacterial
Mycoplasma - genetics
Mycoplasma - metabolism
Plasmids
RNA, Antisense
RNA, Messenger - genetics
Staphylococcus aureus - drug effects
Staphylococcus aureus - genetics
Staphylococcus aureus - growth & development
Transformation, Bacterial
Xylose - pharmacology
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Summary:Summary To address the need for new approaches to antibiotic drug development, we have identified a large number of essential genes for the bacterial pathogen, Staphylococcus aureus, using a rapid shotgun antisense RNA method. Staphylococcus aureus chromosomal DNA fragments were cloned into a xylose‐inducible expression plasmid and transformed into S. aureus. Homology comparisons between 658 S. aureus genes identified in this particular antisense screen and the Mycoplasma genitalium genome, which contains 517 genes in total, yielded 168 conserved genes, many of which appear to be essential in M. genitalium and other bacteria. Examples are presented in which expression of an antisense RNA specifically reduces its cognate mRNA. A cell‐based, drug‐screening assay is also described, wherein expression of an antisense RNA confers specific sensitivity to compounds targeting that gene product. This approach enables facile assay development for high throughput screening for any essential gene, independent of its biochemical function, thereby greatly facilitating the search for new antibiotics.
ISSN:0950-382X
1365-2958
DOI:10.1046/j.1365-2958.2002.02832.x