Cholesterol Catabolism by Mycobacterium tuberculosis Requires Transcriptional and Metabolic Adaptations

Abstract

SUMMARY To understand the adaptation of Mycobacterium tuberculosis to the intracellular environment, we used comprehensive metabolite profiling to identify the biochemical pathways utilized during growth on cholesterol, a critical carbon source during chronic infection. Metabolic alterations observed during cholesterol catabolism centered on propionyl-CoA and pyruvate pools. Consequently, growth on this substrate required the transcriptional induction of the propionyl-CoA assimilating methylcitrate cycle (MCC) enzymes, via the Rv1129c regulatory protein. We show that both Rv1129c and the MCC enzymes are required for intracellular growth in macrophages, and the growth defect of MCC mutants is largely attributable to the degradation of host-derived cholesterol. Together, these observations define a coordinated transcriptional and metabolic adaptation that is required for scavenging carbon during intracellular growth.

Mycobacterial Gene cuvA Is Required for Optimal Nutrient Utilization and Virulence

“…This inference is consistent with a recent study using genetic approaches and metabolic profiling to map the pathway of cholesterol catabolism in M. tuberculosis, in which the Rv1422 protein was not identified as being required for any step in cholesterol degradation (17). These data, together with rescue of the growth defect by addition of glucose, also suggest that the growth defect is not the result of accumulation of a toxic metabolite from catabolism of the several carbon sources on which the ⌬cuvA strain grew less well than the wild type.…”

“…Second, the M. smegmatis ⌬cuvA cholesterol growth defect was fully reversed by the addition of glucose, whereas the growth defects of the icl1 icl2 and prpC prpD M. tuberculosis strains were not reversed by glycerol. In addition, the growth-inhibitory effect of propionate on the M. smegmatis cuvA mutant was not reversed by addition of vitamin B 12 , in contrast to the case with the prpC prpD and icl1 icl2 mutants, in which B 12 addition allows cells to grow on propionate via utilization of the B 12 -dependent methylmalonyl-coenzyme A (CoA) pathway (17,45). Thus, cuvA does not appear to be required for the glyoxylate or methylcitrate pathways.…”

“…Though there is some similarity in the phenotypes of the ⌬cuvA strains and the strains disrupted in glyoxylate or methylcitrate pathways, there are important differences that suggest that CuvA does not participate directly in these pathways. First, the M. tuberculosis strains with defects in the glyoxylate or methylcitrate pathways show minimal growth on cholesterol or propionate in liquid MM (17,45,46). In contrast, the M. tuberculosis ⌬cuvA strain did not have a significant defect in cholesterol-containing liquid medium, and the M. smegmatis ⌬cuvA strain grew more slowly but did achieve sustained growth in MM plus cholesterol and MM plus propionate.…”

“…Each compound was added to a final concentration of 0.01%, the concentration of cholesterol that has been used in mycobacterial growth media because of its low aqueous solubility (2,17). As shown in Fig.…”

“…We identified growth defects on multiple carbon sources for deletion mutants of M. tuberculosis Rv1422 and the Mycobacterium smegmatis homologue MSMEG_3080 compared to the wild-type strains. Because of the importance of cholesterol as a carbon source for M. tuberculosis during infection (2,16,17), we used cholesterol as a representative carbon source for additional investigation of the function of Rv1422. When the M. smegmatis and M. tuberculosis mutant strains were grown on cholesterol, both had morphological abnormalities, including bulging and shortened cell length, indicating a role for Rv1422 in cell wall synthesis.…”