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Identification of gene products involved in the oxidative stress response of Moraxella catarrhalis
Todd C Hoopman1, Wei Liu, Stephanie N Joslin
1Department of Internal Medicine, Pulmonary and Critical Care Medicine, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
Abstract:
Moraxella catarrhalis is subjected to oxidative stress from both internal and environmental sources. A previous study (C. D. Pericone, K. Overweg, P. W. Hermans, and J. N. Weiser, Infect. Immun. 68:3990-3997, 2000) indicated that a wild-type strain of M. catarrhalis was very resistant to killing by exogenous hydrogen peroxide (H₂O₂). The gene encoding OxyR, a LysR family transcriptional regulator, was identified and inactivated in M. catarrhalis strain O35E, resulting in an increase in sensitivity to killing by H₂O₂ in disk diffusion assays and a concomitant aerobic serial dilution effect. Genes encoding a predicted catalase (KatA) and an alkyl hydroperoxidase (AhpCF) showed dose-dependent upregulation in wild-type cells exposed to H₂O₂. DNA microarray and real-time reverse transcription-PCR (RT-PCR) analyses identified M. catarrhalis genes whose expression was affected by oxidative stress in an OxyR-dependent manner. Testing of M. catarrhalis O35E katA and ahpC mutants for their abilities to scavenge exogenous H₂O₂ showed that the KatA catalase was responsible for most of this activity in the wild-type parent strain. The introduction of the same mutations into M. catarrhalis strain ETSU-4 showed that the growth of a ETSU-4 katA mutant was markedly inhibited by the addition of 50 mM H₂O₂ but that this mutant could still form a biofilm equivalent to that produced by its wild-type parent strain.
Insights
Moraxella catarrhalis utilizes OxyR to regulate genes like KatA, crucial for resisting hydrogen peroxide (H₂O₂) oxidative stress. KatA is the primary enzyme responsible for detoxifying H₂O₂ in this bacterium.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Oxidative Stress Response
Background:
- Moraxella catarrhalis faces significant oxidative stress from internal and external sources.
- Previous research indicated wild-type M. catarrhalis exhibits high resistance to exogenous hydrogen peroxide (H₂O₂).
Purpose of the Study:
- To investigate the role of the OxyR regulator in M. catarrhalis's response to oxidative stress.
- To identify specific genes involved in H₂O₂ detoxification and their regulation by OxyR.
Main Methods:
- Gene inactivation of OxyR in M. catarrhalis strain O35E.
- Disk diffusion assays and aerobic serial dilution to assess H₂O₂ sensitivity.
- DNA microarray and real-time reverse transcription-PCR (RT-PCR) to analyze gene expression.
- Construction and testing of katA and ahpC mutants.
Main Results:
- OxyR inactivation increased M. catarrhalis sensitivity to H₂O₂.
- KatA (catalase) and AhpCF (alkyl hydroperoxidase) expression were upregulated by H₂O₂ in an OxyR-dependent manner.
- KatA was identified as the primary enzyme responsible for scavenging exogenous H₂O₂ in wild-type M. catarrhalis.
- A katA mutant in strain ETSU-4 showed significant growth inhibition with H₂O₂, but biofilm formation remained unaffected.
Conclusions:
- OxyR is a key regulator of oxidative stress response in M. catarrhalis, controlling the expression of detoxification enzymes.
- KatA plays a critical role in protecting M. catarrhalis from H₂O₂-induced damage.
- While KatA is vital for growth under oxidative stress, its absence does not impair biofilm formation in M. catarrhalis.
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