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General Commentary ARTICLE

Front. Cell. Infect. Microbiol., 24 October 2013 | https://doi.org/10.3389/fcimb.2013.00068

A central role for aspartate in Mycobacterium tuberculosis physiology and virulence

  • 1Centre National de la Recherche Scientifique, Institut de Pharmacologie et de Biologie Structurale, Toulouse, France
  • 2Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, Université Paul Sabatier, Toulouse, France

A commentary on
Mycobacterium tuberculosis nitrogen assimilation and host colonization require aspartate

by Gouzy, A., Larrouy-Maumus, G., Wu, T. D., Peixoto, A., Levillain, F., Lugo-Villarino, G., et al. (2013). Nat. Chem. Biol. doi: 10.1038/nchembio.1355

The tuberculosis (TB) bacillus, Mycobacterium tuberculosis, is a facultative intracellular pathogen that multiplies inside macrophages, in which it resides within a specialized compartment, the phagosome, where nutrient sources are likely limited. A number of studies provided compelling evidence that M. tuberculosis has the ability to exploit host-derived carbon sources, such as triglycerides, cholesterol and glucose, and to proliferate inside host cells (Mckinney et al., 2000; Pandey and Sassetti, 2008; Daniel et al., 2011; Marrero et al., 2013). In addition to carbon, nitrogen is an essential constituent of all living organisms. As compared to carbon requirements, little is known about the nature of nitrogen-containing molecules utilized by the TB bacillus during infection. We recently discovered that nitrogen incorporation from exogenous aspartate is required for host colonization by M. tuberculosis (Gouzy et al., 2013). This study highlights, for the first time, the potential of amino acids, and aspartate in particular, as a major nitrogen reservoir supporting M. tuberculosis virulence in vivo. It also opens a series of questions to be addressed in the future.

Chief among these questions is the origin and molecular mechanism(s) of transport allowing mycobacterial access to aspartate inside host cells. TB lesions are enriched in aspartate, and this likely relies on global metabolic changes in immune cells during granuloma formation (Somashekar et al., 2011). How nutrients, such as aspartate, access the M. tuberculosis phagosome during infection is an intriguing issue. Using mass spectrometry imaging, we further showed that aspartate can access the mycobacterial phagosome, at least in vitro (Gouzy et al., 2013). In mammalian cells, aspartate uptake relies on transporters of the solute carriers (SLC) superfamily, and in particular on those of the high-affinity glutamate and neutral amino acid (SLC1) and of the cationic amino acid (SLC7) transporter families. Among these transporters, SLC1A2 was reported to mediate aspartate transport inside macrophages (Rimaniol et al., 2001; Ye et al., 2010). Interestingly, we reported the expression of the slc1a2 gene is increased in human macrophages upon M. tuberculosis infection (Tailleux et al., 2008). Increased expression of this transporter could impact considerably the ability of M. tuberculosis to multiply inside macrophages, as it is the case for the neutral amino acid transporter SLC1A5 in the context of Francisella tularensis and Legionella pneumophila infection (Wieland et al., 2005; Barel et al., 2012). These studies suggested the transport of yet to be identified amino acid(s) is important to sustain intracellular multiplication of these two bacterial species. Whether SLC1A2 and/or other members of the SLC superfamily allow M. tuberculosis to access aspartate and multiply inside host cells remains to be evaluated.

Another issue raised by our study is: how is nitrogen transferred from aspartate to other nitrogen-containing molecules in M. tuberculosis? Once acquired by the bacillus through its unique aspartate importer AnsP1, we showed this amino acid species mostly serves as a nitrogen provider and barely enters carbon metabolism through the Krebs cycle (Gouzy et al., 2013). Nitrogen assimilation from aspartate must rely on transamination steps allowing the transfer of aspartate-derived nitrogen to glutamate, which in turn, together with glutamine, provides nitrogen to most of biosynthesis pathways. In M. tuberculosis, two aspartate transaminases, called AspB and AspC, are predicted to mediate nitrogen transfer from aspartate to glutamate (Cole et al., 1998). Interestingly, the aspC gene is thought to be essential in M. tuberculosis (Sassetti et al., 2003), which may indicate that AspC is involved mostly in aspartate biosynthesis, rather than in aspartate catabolism and glutamate synthesis. Genetic inactivation of aspC may thus result in aspartate auxotrophy. The aspB gene, on the opposite, is not essential in vitro (Sassetti et al., 2003). Interestingly, aspB is expressed at higher level in bacteria withstanding conditions mimicking the intracellular environment (Fisher et al., 2002). AspB may thus be involved in aspartate-derived nitrogen assimilation during infection, and may be required for M. tuberculosis virulence. Noteworthy, AspB and AspC possess two homologue proteins, called Rv0858c and Rv1178, that are predicted to act as aspartate aminotransferases, and that might provide alternative pathways for aspartate-derived nitrogen assimilation in the TB bacillus (Cole et al., 1998).

Finally, our study suggests aspartate is an important metabolite required for mycobacterial virulence, and this raises several questions regarding the therapeutic potential of inhibitors of enzymes using aspartate as a substrate. In addition to serve as a nitrogen provider, aspartate possesses non-redundant and essential functions in bacterial metabolism. As a consequence, several enzymes involved in aspartate metabolism, namely the aspartate decarboxylase PanD, the aspartokinase ASK, and the aspartate-β-semialdehyde dehydrogenase ASD, are considered promising targets for novel antituberculous compounds (Shafiani et al., 2005; Gopalan et al., 2006; Chaitanya et al., 2010; Sharma et al., 2012a,b). PanD is the sole enzyme in M. tuberculosis mediating the decarboxylation of aspartate, which allows the formation of vitamin B5, a precursor of the coenzyme A (CoA) co-factor. CoA is an acyl-carrier co-enzyme involved in fatty acid metabolism and is essential for the viability of several microorganisms, including M. tuberculosis (Ambady et al., 2012). Consequently, deletion of the panD gene was shown to result in a severe attenuation of M. tuberculosis virulence in the mouse model (Sambandamurthy et al., 2002). Another essential pathway relying on aspartate metabolism is the synthesis of amino acids belonging to the aspartate amino acid family, namely methionine, threonine and isoleucine, and involving the ASK and ASD enzymes, that play a unique and essential part in aspartate incorporation into these biosynthesis pathways. Furthermore, in addition to their role in the biosynthesis of these amino acids, both enzymes are involved in the synthesis of diaminopimelic acid, an intermediate of the lysine biosynthesis pathway, which is an essential constituent of the bacterial cell wall constituent peptidoglycan. As a consequence, ASK and ASD are attractive drug targets for new antituberculous molecules since i/ they possess no homologue in humans, and ii/ their inhibition in M. tuberculosis may cause both amino acid auxotrophy and cell wall destruction (Shafiani et al., 2005; Chaitanya et al., 2010).

In conclusion, we believe aspartate acquisition and assimilation pathways should be further considered as promising targets for novel anti-TB therapies.

Acknowledgments

The authors received no specific funding for this work. The laboratory of Olivier Neyrolles is supported by the Centre National de la Recherche Scientifique (CNRS), the Fondation pour la Recherche Médicale (FRM), the Agence Nationale de la Recherche, the European Union, and the Fondation Mérieux. Alexandre Gouzy holds a fellowship from the FRM. The funders had no role in the decision to publish this article or in its preparation.

References

Ambady, A., Awasthy, D., Yadav, R., Basuthkar, S., Seshadri, K., and Sharma, U. (2012). Evaluation of CoA biosynthesis proteins of Mycobacterium tuberculosis as potential drug targets. Tuberculosis (Edinb). 92, 521–528. doi: 10.1016/j.tube.2012.08.001

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Barel, M., Meibom, K., Dubail, I., Botella, J., and Charbit, A. (2012). Francisella tularensis regulates the expression of the amino acid transporter SLC1A5 in infected THP-1 human monocytes. Cell Microbiol. 14, 1769–1783. doi: 10.1111/j.1462-5822.2012.01837.x

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Chaitanya, M., Babajan, B., Anuradha, C. M., Naveen, M., Rajasekhar, C., Madhusudana, P., et al. (2010). Exploring the molecular basis for selective binding of Mycobacterium tuberculosis Asp kinase toward its natural substrates and feedback inhibitors: a docking and molecular dynamics study. J. Mol. Model. 16, 1357–1367. doi: 10.1007/s00894-010-0653-4

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Cole, S. T., Brosch, R., Parkhill, J., Garnier, T., Churcher, C., Harris, D., et al. (1998). Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature 393, 537–544. doi: 10.1038/31159

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Daniel, J., Maamar, H., Deb, C., Sirakova, T. D., and Kolattukudy, P. E. (2011). Mycobacterium tuberculosis uses host triacylglycerol to accumulate lipid droplets and acquires a dormancy-like phenotype in lipid-loaded macrophages. PLoS Pathog. 7:e1002093. doi: 10.1371/journal.ppat.1002093

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Fisher, M. A., Plikaytis, B. B., and Shinnick, T. M. (2002). Microarray analysis of the Mycobacterium tuberculosis transcriptional response to the acidic conditions found in phagosomes. J. Bacteriol. 184, 4025–4032. doi: 10.1128/JB.184.14.4025-4032.2002

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Gopalan, G., Chopra, S., Ranganathan, A., and Swaminathan, K. (2006). Crystal structure of uncleaved L-aspartate-alpha-decarboxylase from Mycobacterium tuberculosis. Proteins 65, 796–802. doi: 10.1002/prot.21126

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Gouzy, A., Larrouy-Maumus, G., Wu, T. D., Peixoto, A., Levillain, F., Lugo-Villarino, G., et al. (2013). Mycobacterium tuberculosis nitrogen assimilation and host colonization require aspartate. Nat. Chem. Biol. doi: 10.1038/nchembio.1355. [Epub ahead of print].

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Marrero, J., Trujillo, C., Rhee, K. Y., and Ehrt, S. (2013). Glucose phosphorylation is required for Mycobacterium tuberculosis persistence in mice. PLoS Pathog. 9:e1003116. doi: 10.1371/journal.ppat.1003116

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Mckinney, J. D., Honer Zu Bentrup, K., Munoz-Elias, E. J., Miczak, A., Chen, B., Chan, W. T., et al. (2000). Persistence of Mycobacterium tuberculosis in macrophages and mice requires the glyoxylate shunt enzyme isocitrate lyase. Nature 406, 735–738. doi: 10.1038/35021074

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Pandey, A. K., and Sassetti, C. M. (2008). Mycobacterial persistence requires the utilization of host cholesterol. Proc. Natl. Acad. Sci. U.S.A. 105, 4376–4380. doi: 10.1073/pnas.0711159105

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Rimaniol, A. C., Mialocq, P., Clayette, P., Dormont, D., and Gras, G. (2001). Role of glutamate transporters in the regulation of glutathione levels in human macrophages. Am. J. Physiol. Cell Physiol. 281, C1964–C1970.

Pubmed Abstract | Pubmed Full Text

Sambandamurthy, V. K., Wang, X., Chen, B., Russell, R. G., Derrick, S., Collins, F. M., et al. (2002). A pantothenate auxotroph of Mycobacterium tuberculosis is highly attenuated and protects mice against tuberculosis. Nat. Med. 8, 1171–1174. doi: 10.1038/nm765

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Sassetti, C. M., Boyd, D. H., and Rubin, E. J. (2003). Genes required for mycobacterial growth defined by high density mutagenesis. Mol. Microbiol. 48, 77–84. doi: 10.1046/j.1365-2958.2003.03425.x

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Shafiani, S., Sharma, P., Vohra, R. M., and Tewari, R. (2005). Cloning and characterization of aspartate-beta-semialdehyde dehydrogenase from Mycobacterium tuberculosis H37 Rv. J. Appl. Microbiol. 98, 832–838. doi: 10.1111/j.1365-2672.2004.02505.x

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Sharma, R., Florea, M., Nau, W. M., and Swaminathan, K. (2012a). Validation of drug-like inhibitors against Mycobacterium tuberculosis L-aspartate alpha-decarboxylase using nuclear magnetic resonance (1H NMR). PLoS ONE 7:e45947. doi: 10.1371/journal.pone.0045947

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Sharma, R., Kothapalli, R., Van Dongen, A. M., and Swaminathan, K. (2012b). Chemoinformatic identification of novel inhibitors against Mycobacterium tuberculosis L-aspartate alpha-decarboxylase. PLoS ONE 7:e33521. doi: 10.1371/journal.pone.0033521

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Somashekar, B. S., Amin, A. G., Rithner, C. D., Troudt, J., Basaraba, R., Izzo, A., et al. (2011). Metabolic profiling of lung granuloma in Mycobacterium tuberculosis infected guinea pigs: ex vivo 1H magic angle spinning NMR studies. J. Proteome Res. 10, 4186–4195. doi: 10.1021/pr2003352

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Tailleux, L., Waddell, S. J., Pelizzola, M., Mortellaro, A., Withers, M., Tanne, A., et al. (2008). Probing host pathogen cross-talk by transcriptional profiling of both Mycobacterium tuberculosis and infected human dendritic cells and macrophages. PLoS ONE 3:e1403. doi: 10.1371/journal.pone.0001403

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Wieland, H., Ullrich, S., Lang, F., and Neumeister, B. (2005). Intracellular multiplication of Legionella pneumophila depends on host cell amino acid transporter SLC1A5. Mol. Microbiol. 55, 1528–1537. doi: 10.1111/j.1365-2958.2005.04490.x

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Ye, R., Rhoderick, J. F., Thompson, C. M., and Bridges, R. J. (2010). Functional expression, purification and high sequence coverage mass spectrometric characterization of human excitatory amino acid transporter EAAT2. Protein Expr. Purif. 74, 49–59. doi: 10.1016/j.pep.2010.04.006

Pubmed Abstract | Pubmed Full Text | CrossRef Full Text

Keywords: Mycobacterium tuberculosis, macrophage, phagosome, aspartate, nitrogen

Citation: Gouzy A, Poquet Y and Neyrolles O (2013) A central role for aspartate in Mycobacterium tuberculosis physiology and virulence. Front. Cell. Infect. Microbiol. 3:68. doi: 10.3389/fcimb.2013.00068

Received: 30 September 2013; Accepted: 09 October 2013;
Published online: 24 October 2013.

Edited by:

Monique Barel, Institut National de la Santé et de la Recherche Médicale, France

Reviewed by:

Greg Cook, University of Otago, New Zealand

Copyright © 2013 Gouzy, Poquet and Neyrolles. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

*Correspondence: olivier.neyrolles@ipbs.fr