The mycobacterial cell envelope — a moving target

Dulberger, Charles L.; Rubín, Eric J.; Boutte, Cara C.

doi:10.1038/s41579-019-0273-7

Cited by 251 publications

(283 citation statements)

References 151 publications

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“…Loss of such cell-wall enzymes may result in collateral perturbation to the cell membrane proteome, causing LepA to become indispensable to the cell. LepA function in mycobacterial appears to be critical for maintenance of the mycomembrane, a very complex network of lipids and proteins [67].…”

Section: Discussionmentioning

confidence: 99%

A conserved translation factor is required for optimal synthesis of a membrane protein family in mycobacteria

Fishbein

Wolf

Dulberger

et al. 2019

Preprint

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14Ribosomes require the activity of associated GTPases to synthesize proteins. Despite strong 15 evolutionary conservation, the roles of many of these remain unknown. For example, LepA (also known 16 as elongation factor 4) is a ribosome-associated GTPase found in bacteria, mitochondria, and 17 chloroplasts, yet its physiological contribution to cell survival is not clear. Recently, we found that loss of 18 lepA in Mycobacterium smegmatis (Msm) altered tolerance to rifampin, a drug that targets a non-19 ribosomal cellular process. To uncover the determinants of LepA-mediated drug tolerance, we 20 characterized the whole-cell proteomes and transcriptomes of a lepA deletion mutant relative to a wild-21 type strain. We find that LepA is important for the steady-state abundance of an outer membrane porin, 22which is integral to nutrient uptake and drug susceptibility. Loss of LepA leads to a decreased amount 23 of porin in the membrane, resulting in the drug tolerance phenotype of the lepA mutant. LepA control 24 requires a sequence motif in the 5' region of the porin transcript. Thus, LepA controls the abundance of 25 specific proteins, likely through its activity during translation. 27 Importance 28Our understanding of how ribosomes properly synthesis an entire cellular proteome, in all its 29 complexity, is still evolving. Ribosomal GTPases are often highly conserved, but the roles of many are 30 not well understood. For example, elongation factor 4, or LepA, is a ribosome-associated GTPase 31 conserved across bacteria, mitochondria, and chloroplasts. Using whole-cell proteomics and RNA-32 sequencing of wild type and a lepA deletion mutant, we find that LepA improves translation of 33 mycobacterial porins in a message-specific manner. As porins play a key role in cell wall permeability, 34 loss of LepA produces a plethora of phenotypic changes. These findings underline the problem of 35 building proteins into a complex cell wall, such as that of mycobacteria, and point to a solution in the 36 use of GTPases such as LepA, that have evolved to aid in specific protein synthesis. 37 38 Keywords 39 LepA; ribosome; mycobacteria; porin; drug susceptibility 40 41 45 to adaptive, cellular responses. Indeed, the ribosome machinery and its associated factors/RNA 46 species adapt the cell during periods of environmental transition [2] [3, 4]. Recent advances in 47 techniques such as quantitative proteomics, ribosome profiling, and cryo-electron microscopy (cryo-48 EM) have revealed an intricate balance of mRNA sequences and associating proteins at the ribosome 49 that are necessary for the homeostatic cellular proteome [5-7]. Regardless of growth environment,50bacterial cells are likely composed of a heterogenous population of ribosomes [8, 9], with some inactive 51 and others actively translating parts of the proteome [10, 11]. Associating factors and RNA species tune 52 each ribosome, creating a cell with a spatially-and temporally-regulated proteome [8, 12]. 53The successful synthesis of a given protein at the ribosome is ...

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Section: Discussionmentioning

confidence: 99%

A conserved translation factor is required for optimal synthesis of a membrane protein family in mycobacteria

Fishbein

Wolf

Dulberger

et al. 2019

Preprint

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“…In dormant, non-replicating Mtb cells, cell wall synthesis is downregulated (Galagan et al 2013) and remodeled (Dulberger, Rubin, and Boutte 2019). This regulation of the wall protects Mtb from both the immune system and antibiotics during infection.…”

Section: Discussionmentioning

confidence: 99%

“…PstP and Pkn A and B seem to have a mutual feedback regulatory loop (Iswahyudi et al 2019). Previous work has shown that dephosphorylation of PknA and PknB downregulates their activity (Sajid et al 2011; Boitel et al 2003) which would be expected to broadly downregulate cell growth (Betts et al 2002; Ortega et al 2014; Dulberger, Rubin, and Boutte 2019) The phosphorylation of PstP has been shown to stimulate its activity against small molecule substrates (Sajid et al 2011). However, dephosphorylation by PstP is known to both upregulate cell wall synthesis through KasA (Molle et al 2006) and to downregulate cell wall synthesis through CwlM (Boutte et al 2016) (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…These phospho-signaling events are likely involved in the transitions between growth and stasis during infection. However, peptidoglycan and lipid metabolism should be largely correlated (Dulberger, Rubin, and Boutte 2019), so PstP must be able to minutely toggle its substrate specificity between growth and stasis. For example, we expect that PstP should dephosphorylate KasA but not CwlM during growth, and should switch this specificity in stasis.…”

Section: Introductionmentioning

confidence: 99%

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Phosphorylation on PstP regulates cell wall metabolism and antibiotic tolerance in Mycobacterium smegmatis

Shamma

Papavinasasundaram

Bandekar

et al. 2019

Preprint

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13The mycobacterial cell wall is a dynamic structure that protects Mycobacterium 14 tuberculosis and its relatives from environmental stresses. Modulation of cell wall 15 metabolism under stress is thought to be responsible for decreased cell wall permeability 16 and increased tolerance to antibiotics. The signaling pathways that control cell wall 17 metabolism under stress, however, are poorly understood. Here, we examine the 18 signaling capacity of a cell wall master regulator, the Serine Threonine Phosphatase 19PstP, in the model organism Mycobacterium smegmatis. We studied how interference 20 with a regulatory phosphorylation site on PstP affects growth, cell wall metabolism and 21 antibiotic tolerance. We find that a phospho-mimetic mutation, pstP T171E, slows 22 growth, misregulates both mycolic acid and peptidoglycan metabolism in different 23 conditions, and interferes with antibiotic tolerance. These data suggest that 24 phosphorylation on PstP controls its substrate specificity and is important in the 25

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“…They can be covalently attached to the cell wall arabinogalactan or can be esterified to trehalose to produce trehalose monomycolate (TMM) and trehalose dimycolate (TDM) [25], or they can remain free. Given the importance of mycolic acids in the interaction of M. tuberculosis with the host [26], we subsequently assessed the mycolic acid content in both strains.…”

mentioning

confidence: 99%

A Phenotypic Characterization of Two Isolates of a Multidrug-Resistant Outbreak Strain of Mycobacterium tuberculosis with Opposite Epidemiological Fitness

Bei

Bigi

Lima

et al. 2020

BioMed Research International

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Tuberculosis (TB) is an infectious disease, caused by Mycobacterium tuberculosis, primarily affecting the lungs. The M. tuberculosis strain of the Haarlem family named M was responsible for a large multidrug-resistant TB (MDR-TB) outbreak in Buenos Aires. This outbreak started in the early 1990s and in the mid 2000s still accounted for 29% of all MDR-TB cases in Argentina. By contrast, a clonal variant of strain M, named 410, has caused a single tuberculosis case since the onset of the outbreak. The molecular bases of the high epidemiological fitness of the M strain remain unclear. To assess its unique molecular properties, herein, we performed a comparative protein and lipid analysis of a representative clone of the M strain (Mp) and the nonprosperous M variant 410. We also evaluated their growth in low pH. The variant 410 had higher levels of latency proteins under standard conditions and delayed growth at low pH, suggesting that it is more sensitive to stress stimuli than Mp. Moreover, Mp showed higher levels of mycolic acids covalently attached to the cell wall and lower accumulation of free mycolic acids in the outer layer than the 410 strain. The low expression of latency proteins together with the reduced content of surface mycolic acids may facilitate Mp to evade the host immune responses.

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The mycobacterial cell envelope — a moving target

Cited by 251 publications

References 151 publications

A conserved translation factor is required for optimal synthesis of a membrane protein family in mycobacteria

A conserved translation factor is required for optimal synthesis of a membrane protein family in mycobacteria

Phosphorylation on PstP regulates cell wall metabolism and antibiotic tolerance in Mycobacterium smegmatis

A Phenotypic Characterization of Two Isolates of a Multidrug-Resistant Outbreak Strain of Mycobacterium tuberculosis with Opposite Epidemiological Fitness

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