Mycolyltransferase-mediated Glycolipid Exchange in Mycobacteria

Abstract

Trehalose dimycolate (TDM), also known as cord factor, is a major surface glycolipid of the cell wall of mycobacteria. Because of its potent biological functions in models of infection, adjuvancy, and immunotherapy, it is important to determine how its biosynthesis is regulated. Here we show that glucose, a host-derived product that is not readily available in the environment, causes Mycobacterium avium to down-regulate TDM expression while up-regulating production of another major glycolipid with immunological roles in T cell activation, glucose monomycolate (GMM). In vitro, the mechanism of reciprocal regulation of TDM and GMM involves competitive substrate selection by antigen 85A. The switch from TDM to GMM biosynthesis occurs near the physiological concentration of glucose present in mammalian hosts. We further demonstrate that GMM is produced in vivo by mycobacteria growing in mouse lung. These results establish an enzymatic pathway for GMM production. More generally, these observations provide a specific enzymatic mechanism for dynamic alterations of cell wall glycolipid remodeling in response to the transition from noncellular to cellular growth environments, including factors that are monitored by the host immune system. Mycobacterium avium complex (MAC)2 includes a group of acid-fast bacteria that distribute widely in natural environments, including soil, water, aerosols, and dust (1). Although less virulent than Mycobacterium tuberculosis, these environmental mycobacteria occasionally infect humans, especially patients infected with human immunodeficiency virus type 1, where they represent a major cause of morbidity. The incidence of clinically overt MAC infection has increased significantly in recent years, and because of the multidrug resistance evolved by the microbes, MAC infection is difficult to clear with chemotherapeutic agents. Thus, M. tuberculosis and MAC are now the two major groups of mycobacteria species that require further efforts for prevention and treatment. Unlike M. tuberculosis, which transmits primarily from individuals with active disease, epidemiologic evidence suggests that such transmission pathways are unlikely for MAC. Rather, MAC infection appears to occur when susceptible individuals are exposed to environmental MAC. These observations predict that, upon infection, environmental MAC should undergo significant adaptive changes to allow its survival and replication within the host.Mycobacteria possess highly lipid-rich cell walls that are critical not simply for their acid-fast properties but also for their survival and replication. The cell wall contains mycolic acids, an ␣-alkyl-␤-hydroxy fatty acid with extremely long carbon chains (ϳC 80 ), which are densely aligned in covalent association with the 6-position of arabinose termini of the underlying arabinogalactan sugar layer or exist as free molecules complexed to sugars, either glucose or trehalose. Arabinogalactan-linked mycolates are proposed to extend outward and interact noncovalently with carbon chains of the so-c...

Exposure of Mycobacteria to Cell Wall-inhibitory Drugs Decreases Production of Arabinoglycerolipid Related to Mycolyl-arabinogalactan-peptidoglycan Metabolism

“…However, the glucose-containing medium allowed M. bovis BCG to produce another apolar glycolipid that migrates above TDM and that was subsequently identified by MS as a glucose monomycolate as recently reported in M. avium (Fig. 6B and data not shown) (35). Taken together, these results indicate that DMAG is present in various mycobacterial species and particularly in large quantities in slow growing species such as M. tuberculosis.…”

“…In their experiments, the investigators extracted DMAG using a biphasic solution of hexane/methanol/water (1:0.05:0.45, v/v/v), whereas our study included a less polar biphasic solution made of petroleum ether/methanol/water (1:0.9:0.1, v/v/v) (32)(33)(34). A second explanation may rely on differences in growth and culture conditions that are known to drastically influence the synthesis of bacterial surface glycoconjugates, including glycolipids (35) and polysaccharides (41). A relevant example is the synthesis of glucose monomycolate, which is up-regulated in M. avium by the presence of glucose in the growth environment (35).…”

“…A second explanation may rely on differences in growth and culture conditions that are known to drastically influence the synthesis of bacterial surface glycoconjugates, including glycolipids (35) and polysaccharides (41). A relevant example is the synthesis of glucose monomycolate, which is up-regulated in M. avium by the presence of glucose in the growth environment (35). This point is also emphasized in our study: the replacement of glycerol with glucose strongly reduced the production of DMAG and stimulated the production of glucose monomycolate.…”

Lipidomic analyses of Mycobacterium tuberculosis based on accurate mass measurements and the novel “Mtb LipidDB”

“…The abundance and biological importance of the Mtb lipids has resulted in extensive and elegant studies to elucidate their structures and functions ( 1-3 ). In many cases, the lipids of Mtb are unique to this pathogen or shared only with other members of this genus.Earlier studies demonstrate variability in lipid profi les among different strains of Mtb ( 7-11 ) and that minor variations in the structure of individual lipids can occur with changes in the growth environment (12)(13)(14)(15)(16)(17)(18)(19)(20). However, targeted and nontargeted assays that monitor changes in Mtb lipid profi les are generally performed by traditional TLC-based methods ( 21 ), and global lipidomics analyses in Mtb have been restricted due to limits in the technology to detect and rapidly identify a large number of lipids in a single experiment.…”

Trehalose Dimycolate Elicits Eosinophilic Skin Hypersensitivity in Mycobacteria-Infected Guinea Pigs

“…Mycobacterial mycolyltransferases are known to catalyze the final step of TDM synthesis from its precursor, TMM, by addition of a mycolyl acyl group. Upon exposure to host-derived glucose, however, competitive substrate selection of TMM and glucose by mycolyltransferases results in down-regulation of TDM accompanied with up-regulated biosynthesis of GMM (38). As shown in Fig.…”