Enhanced immunogenicity of Mycobacterium bovis BCG through CRISPRi mediated depletion of AftC

Bala T.S.A. Madduri; Lauren Allen; Stephen C. Taylor; Gurdyal S. Besra; Luke J. Alderwick

doi:10.1016/j.tcsw.2022.100088

Enhanced immunogenicity of Mycobacterium bovis BCG through CRISPRi mediated depletion of AftC

Bala T.S.A. Madduri, Lauren Allen, Stephen C. Taylor, Gurdyal S. Besra^*, Luke J. Alderwick

^*Corresponding author for this work

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Abstract

Mycobacterium tuberculosis causes the disease tuberculosis and affects a third of the world's population. The recent COVID-19 pandemic exacerbated the situation with a projected 27% increase in tuberculosis related deaths. M. tuberculosis has an elaborate cell wall consisting of peptidoglycan, arabinogalactan and mycolic acids which shield the bacilli from the toxic bactericidal milieu within phagocytes. Amongst, the numerous glycosyltransferase enzymes involved in mycobacterial cell wall biosynthesis, arabinofuranosyltransferase C (aftC) is responsible for the branching of the arabinan domain in both arabinogalactan and lipoarabinomannan. Using Clustered Regularly Interspaced Short Palindromic Repeats interference (CRISPRi) we have generated aftC knockdowns in Mycobacterium bovis BCG and demonstrated the generation of a truncated, immunogenic lipoarabinomannan within its cell envelope. The aftC depleted BCG mutants were unable to form characteristic mycobacterial pellicular biofilms and elicit a potent immunostimulatory phenotype compared to wild type M. bovis BCG in a THP1 cell line. This study paves the way to further explore novel BCG mutants as promising vaccine boosters in preventing pulmonary tuberculosis.

Original language	English
Article number	100088
Journal	Cell Surface
Volume	8
Early online date	11 Nov 2022
DOIs	https://doi.org/10.1016/j.tcsw.2022.100088
Publication status	Published - Dec 2022

Bibliographical note

Funding Information: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:
Gurdyal Singh Besra reports financial support was provided by Medical Research Council. Bala T. S. A. Madduri reports financial support was provided by Darwin Trust of Edinburgh. Gurdyal Singh Besra reports a relationship with Microbiology Society that includes: travel reimbursement.
GSB also acknowledges support in the form of a Personal Research Chair from Mr James Bardrick, a Royal Society Wolfson Research Merit Award, The Medical Research Council (MR/S000542/1, MR/R001154/1 and MR/P015859/1), BM was awarded a Postgraduate Research Scholarship by the University of Birmingham and Darwin Trust of Edinburgh. We thank Dr CM Santosh Kumar who initially helped with the generation of mutants

Open Access: This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/

Publisher Copyright: © 2022 The Author(s). Published by Elsevier B.V.

Citation: Bala T.S.A. Madduri, Lauren Allen, Stephen C. Taylor, Gurdyal S. Besra, Luke J. Alderwick, Enhanced immunogenicity of Mycobacterium bovis BCG through CRISPRi mediated depletion of AftC, The Cell Surface, Volume 8, 2022, 100088,
ISSN 2468-2330, https://doi.org/10.1016/j.tcsw.2022.100088.

DOI: https://doi.org/10.1016/j.tcsw.2022.100088.

Keywords

Arabinofuranosytransferase C
CRISPR interference
Lipoarabinomannan
Mycobacterium bovis BCG
Transcriptional repression

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1016/j.tcsw.2022.100088Licence: CC BY

Madduri2022EnhancedImmunogenicityOfMycobacteriumBovisBCGThroughCRISPRiMediatedDepletionOfAftCCellSurfFinal published version, 4.72 MBLicence: CC BY

Cite this

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title = "Enhanced immunogenicity of Mycobacterium bovis BCG through CRISPRi mediated depletion of AftC",

abstract = "Mycobacterium tuberculosis causes the disease tuberculosis and affects a third of the world's population. The recent COVID-19 pandemic exacerbated the situation with a projected 27% increase in tuberculosis related deaths. M. tuberculosis has an elaborate cell wall consisting of peptidoglycan, arabinogalactan and mycolic acids which shield the bacilli from the toxic bactericidal milieu within phagocytes. Amongst, the numerous glycosyltransferase enzymes involved in mycobacterial cell wall biosynthesis, arabinofuranosyltransferase C (aftC) is responsible for the branching of the arabinan domain in both arabinogalactan and lipoarabinomannan. Using Clustered Regularly Interspaced Short Palindromic Repeats interference (CRISPRi) we have generated aftC knockdowns in Mycobacterium bovis BCG and demonstrated the generation of a truncated, immunogenic lipoarabinomannan within its cell envelope. The aftC depleted BCG mutants were unable to form characteristic mycobacterial pellicular biofilms and elicit a potent immunostimulatory phenotype compared to wild type M. bovis BCG in a THP1 cell line. This study paves the way to further explore novel BCG mutants as promising vaccine boosters in preventing pulmonary tuberculosis.",

keywords = "Arabinofuranosytransferase C, CRISPR interference, Lipoarabinomannan, Mycobacterium bovis BCG, Transcriptional repression",

author = "Madduri, {Bala T.S.A.} and Lauren Allen and Taylor, {Stephen C.} and Besra, {Gurdyal S.} and Alderwick, {Luke J.}",

note = "Funding Information: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Gurdyal Singh Besra reports financial support was provided by Medical Research Council. Bala T. S. A. Madduri reports financial support was provided by Darwin Trust of Edinburgh. Gurdyal Singh Besra reports a relationship with Microbiology Society that includes: travel reimbursement. GSB also acknowledges support in the form of a Personal Research Chair from Mr James Bardrick, a Royal Society Wolfson Research Merit Award, The Medical Research Council (MR/S000542/1, MR/R001154/1 and MR/P015859/1), BM was awarded a Postgraduate Research Scholarship by the University of Birmingham and Darwin Trust of Edinburgh. We thank Dr CM Santosh Kumar who initially helped with the generation of mutants Open Access: This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/ Publisher Copyright: {\textcopyright} 2022 The Author(s). Published by Elsevier B.V. Citation: Bala T.S.A. Madduri, Lauren Allen, Stephen C. Taylor, Gurdyal S. Besra, Luke J. Alderwick, Enhanced immunogenicity of Mycobacterium bovis BCG through CRISPRi mediated depletion of AftC, The Cell Surface, Volume 8, 2022, 100088, ISSN 2468-2330, https://doi.org/10.1016/j.tcsw.2022.100088. DOI: https://doi.org/10.1016/j.tcsw.2022.100088.",

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AU - Allen, Lauren

AU - Taylor, Stephen C.

AU - Besra, Gurdyal S.

AU - Alderwick, Luke J.

N1 - Funding Information: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Gurdyal Singh Besra reports financial support was provided by Medical Research Council. Bala T. S. A. Madduri reports financial support was provided by Darwin Trust of Edinburgh. Gurdyal Singh Besra reports a relationship with Microbiology Society that includes: travel reimbursement. GSB also acknowledges support in the form of a Personal Research Chair from Mr James Bardrick, a Royal Society Wolfson Research Merit Award, The Medical Research Council (MR/S000542/1, MR/R001154/1 and MR/P015859/1), BM was awarded a Postgraduate Research Scholarship by the University of Birmingham and Darwin Trust of Edinburgh. We thank Dr CM Santosh Kumar who initially helped with the generation of mutants Open Access: This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/ Publisher Copyright: © 2022 The Author(s). Published by Elsevier B.V. Citation: Bala T.S.A. Madduri, Lauren Allen, Stephen C. Taylor, Gurdyal S. Besra, Luke J. Alderwick, Enhanced immunogenicity of Mycobacterium bovis BCG through CRISPRi mediated depletion of AftC, The Cell Surface, Volume 8, 2022, 100088, ISSN 2468-2330, https://doi.org/10.1016/j.tcsw.2022.100088. DOI: https://doi.org/10.1016/j.tcsw.2022.100088.

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AB - Mycobacterium tuberculosis causes the disease tuberculosis and affects a third of the world's population. The recent COVID-19 pandemic exacerbated the situation with a projected 27% increase in tuberculosis related deaths. M. tuberculosis has an elaborate cell wall consisting of peptidoglycan, arabinogalactan and mycolic acids which shield the bacilli from the toxic bactericidal milieu within phagocytes. Amongst, the numerous glycosyltransferase enzymes involved in mycobacterial cell wall biosynthesis, arabinofuranosyltransferase C (aftC) is responsible for the branching of the arabinan domain in both arabinogalactan and lipoarabinomannan. Using Clustered Regularly Interspaced Short Palindromic Repeats interference (CRISPRi) we have generated aftC knockdowns in Mycobacterium bovis BCG and demonstrated the generation of a truncated, immunogenic lipoarabinomannan within its cell envelope. The aftC depleted BCG mutants were unable to form characteristic mycobacterial pellicular biofilms and elicit a potent immunostimulatory phenotype compared to wild type M. bovis BCG in a THP1 cell line. This study paves the way to further explore novel BCG mutants as promising vaccine boosters in preventing pulmonary tuberculosis.

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KW - Lipoarabinomannan

KW - Mycobacterium bovis BCG

KW - Transcriptional repression

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Enhanced immunogenicity of Mycobacterium bovis BCG through CRISPRi mediated depletion of AftC

Abstract

Bibliographical note

Keywords

UN SDGs

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