B-279. Function and Expression of the hmu Heme Transport System in Corynebacterium Species

M. P. Schmitt, C. E. Allen, C. A. Kunkle;
FDA/CBER, Bethesda, MD.

The acquisition of iron is required by numerous bacterial pathogens to cause disease. Host compounds, such as heme and hemoglobin, are used as essential iron sources by many human pathogens including the virulent Corynebacterium species, C. diphtheriae and C. ulcerans. In this study, we examined the hmu heme transport region in C. diphtheriae, a genetic cluster that includes a previously described ABC-type heme transporter encoded by the hmuTUV genes and three additional genes, which we have designated htaA, htaB and htaC. The htaA gene is predicted to encode a cell surface or secreted protein that may function as a receptor for heme or hemoglobin. The htaA gene lies upstream from hmuTUV, and appears to be part of an iron and DtxR-regulated operon that includes hmuTUV. The htaC gene is divergently transcribed from htaA, while htaB is located downstream from hmuTUV and is expressed from a separate iron/DtxR-regulated promoter. A previously described allelic replacement method was used to construct several defined deletions within the hmu locus in C. diphtheriae and in C. ulcerans. The mutants were examined in a quantitative biological assay to determine their ability to use hemoglobin as a sole source of iron for growth. The findings from these studies demonstrate that the hmuTUV genes are involved in the use of heme as an iron source in both species; however, the hmuTUV deletion in C. ulcerans fully abolished heme utilization, while this mutation in C. diphtheriae caused only a partial defect. A non-polar deletion in htaA resulted in an approximately 50% reduction in heme-iron utilization in C. diphtheriae, but a similar deletion of htaA had no effect in C. ulcerans. A deletion of the entire hmu locus in C. diphtheriae resulted in a heme utilization phenotype similar to that of the hmuTUV deletion. In conclusion, these results suggests that factors other than those encoded by the hmu gene cluster are involved in heme transport in C. diphtheriae, and possibly in C. ulcerans, although the HmuTUV system appears to be essential for the use of heme-iron in C. ulcerans.