A putative ABC transporter, hatABCDE, is among molecular determinants of pyomelanin production in Pseudomonas...

Area of Science:

• Microbiology
• Biochemistry
• Genetics

Background:

• Pyomelanin overproduction is common in Pseudomonas aeruginosa from cystic fibrosis and UTIs, suggesting a role in microbial persistence.
• The precise mechanisms governing pyomelanin production, particularly homogentisic acid (HGA) precursor secretion, remain largely unelucidated.

Purpose of the Study:

• To identify novel genetic factors involved in pyomelanin biosynthesis and homogentisic acid (HGA) production in Pseudomonas aeruginosa.
• To characterize the function of a newly identified ABC transporter system in the secretion of HGA.

Main Methods:

• Transposon mutagenesis was employed to identify genes contributing to melanogenesis in a clinical P. aeruginosa isolate.
• Quantitative analysis of HGA production and intracellular accumulation was performed using mutant strains.
• Quantitative real-time PCR was used to assess gene expression levels.

Main Results:

• Beyond known enzymes, 26 genes regulating, metabolizing, transporting, or hypothetically involved in HGA production were identified.
• A novel ABC transporter, HatABCDE, was found to be essential for extracellular HGA secretion; mutants showed reduced HGA production and intracellular accumulation.
• Intracellular HGA accumulation triggered the upregulation of the hatABCDE transport genes.

Conclusions:

• The hatABCDE operon encodes a critical ABC transporter responsible for secreting homogentisic acid (HGA), the pyomelanin precursor, in Pseudomonas aeruginosa.
• Dysfunction of this transporter leads to intracellular HGA buildup and impacts pyomelanin production.
• This discovery provides new insights into the regulation and transport mechanisms of pyomelanin biosynthesis in P. aeruginosa.