Mycobacterium tuberculosis expresses a novel pH-dependent divalent cation transporter belonging to the Nramp family

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• Mammalian natural resistance-associated macrophage protein (Nramp) homologues influence susceptibility to intracellular pathogens.
• Mycobacteria, including Mycobacterium tuberculosis and Mycobacterium bovis (bacillus Calmette-Guérin [BCG]), encode an Nramp homologue called Mramp.
• Eukaryotic Nramp homologues are known to transport divalent cations like Fe(2+), Mn(2+), Zn(2+), and Cu(2+).

Purpose of the Study:

• To characterize the function and regulation of the Mycobacterium tuberculosis Nramp homologue (Mramp).
• To investigate the role of Mramp in metal ion transport and its potential interaction with host Nramp1.
• To explore the implications of Mramp for the intracellular survival of mycobacteria.

Main Methods:

• Xenopus laevis oocyte expression system to assay Mramp-mediated metal ion uptake.
• Semiquantitative and quantitative reverse transcription PCR to measure Mramp and y39 mRNA levels in Mycobacterium tuberculosis under varying metal ion concentrations.
• Culturing of Mycobacterium bovis BCG axenically and within THP-1 cells to detect Mramp mRNA expression.

Main Results:

• RNA encoding Mramp induced significant increases in Zn(2+) and Fe(2+) uptake in Xenopus oocytes, dependent on acidic pH.
• Mramp-mediated transport was inhibited by Mn(2+) and Cu(2+), indicating interaction with multiple divalent cations.
• Mramp mRNA levels in M. tuberculosis increased with low concentrations of Fe(2+) and Cu(2+), in parallel with the metal-transporting P-type ATPase y39.
• Higher Cu(2+) concentrations (5-70 microM) resulted in a fourfold decrease in Mramp/y39 mRNA ratios.
• Mramp mRNA was detected in both axenically cultured M. bovis BCG and BCG within THP-1 cells.

Conclusions:

• Mramp represents a novel prokaryotic class of metal ion transporter.
• Mramp's expression is regulated by environmental metal ion concentrations, particularly iron and copper.
• Competition between Mramp and host Nramp1 for divalent cations within phagosomes could be a critical factor for mycobacterial intracellular survival.