Transcriptomic and phenotypic analysis of paralogous spx gene function in Bacillus anthracis Sterne

Skye Barendt, Hyunwoo Lee, Cierra Birch, Michiko Nakano, Marcus Jones, Peter Zuber

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

Spx of Bacillus subtilis is a redox-sensitive protein, which, under disulfide stress, interacts with RNA polymerase to activate genes required for maintaining thiol homeostasis. Spx orthologs are highly conserved among low %GC Gram-positive bacteria, and often exist in multiple paralogous forms. In this study, we used B. anthracis Sterne, which harbors two paralogous spx genes, spxA1 and spxA2, to examine the phenotypes of spx null mutations and to identify the genes regulated by each Spx paralog. Cells devoid of spxA1 were sensitive to diamide and hydrogen peroxide, while the spxA1 spoxA2 double mutant was hypersensitive to the thiol-specific oxidant, diamide. Bacillus anthracis Sterne strains expressing spxA1DD or spxA2DD alleles encoding protease-resistant products were used in microarray and quantitative real-time polymerase chain reaction (RT-qPCR) analyses in order to uncover genes under SpxA1, SpxA2, or SpxA1/SpxA2 control. Comparison of transcriptomes identified many genes that were upregulated when either SpxA1DD or SpxA2DD was produced, but several genes were uncovered whose transcript levels increased in only one of the two SpxADD-expression strains, suggesting that each Spx paralog governs a unique regulon. Among genes that were upregulated were those encoding orthologs of proteins that are specifically involved in maintaining intracellular thiol homeostasis or alleviating oxidative stress. Some of these genes have important roles in B. anthracis pathogenesis, and a large number of upregulated hypothetical genes have no homology outside of the B. cereus/thuringiensis group. Microarray and RT-qPCR analyses also unveiled a regulatory link that exists between the two spx paralogous genes. The data indicate that spxA1 and spxA2 are transcriptional regulators involved in relieving disulfide stress but also control a set of genes whose products function in other cellular processes.

Original languageEnglish (US)
Pages (from-to)695-714
Number of pages20
JournalMicrobiologyOpen
Volume2
Issue number4
DOIs
StatePublished - Aug 2013

Fingerprint

Bacillus anthracis
Genes
Sulfhydryl Compounds
Diamide
Disulfides
Homeostasis
Regulon
Gram-Positive Bacteria
DNA-Directed RNA Polymerases
Bacillus subtilis
Transcriptome
Oxidants
Hydrogen Peroxide
Oxidation-Reduction
Real-Time Polymerase Chain Reaction
Proteins
Oxidative Stress
Peptide Hydrolases
Alleles

Keywords

  • Bacillus anthracis
  • Oxidative stress
  • SpxA1
  • SpxA2
  • Transcriptomic

ASJC Scopus subject areas

  • Microbiology

Cite this

Transcriptomic and phenotypic analysis of paralogous spx gene function in Bacillus anthracis Sterne. / Barendt, Skye; Lee, Hyunwoo; Birch, Cierra; Nakano, Michiko; Jones, Marcus; Zuber, Peter.

In: MicrobiologyOpen, Vol. 2, No. 4, 08.2013, p. 695-714.

Research output: Contribution to journalArticle

Barendt, Skye ; Lee, Hyunwoo ; Birch, Cierra ; Nakano, Michiko ; Jones, Marcus ; Zuber, Peter. / Transcriptomic and phenotypic analysis of paralogous spx gene function in Bacillus anthracis Sterne. In: MicrobiologyOpen. 2013 ; Vol. 2, No. 4. pp. 695-714.
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