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Plasma Membrane Profiling Reveals Upregulation of ABCA1 by Infected Macrophages Leading to Restriction of Mycobacterial Growth.

Long J, Basu Roy R, Zhang YJ, Antrobus R, Du Y, Smith DL, Weekes MP, Javid B - Front Microbiol (2016)

Bottom Line: The plasma membrane represents a critical interface between the internal and extracellular environments, and harbors multiple proteins key receptors and transporters that play important roles in restriction of intracellular infection.We quantified 559 PM proteins in BCG-infected THP-1 cells.One significantly upregulated cell-surface protein was the cholesterol transporter ABCA1.

View Article: PubMed Central - PubMed

Affiliation: Collaboration Innovation Centre for the Diagnosis and Treatment of Infectious Diseases, School of Medicine, Tsinghua University Beijing, China.

ABSTRACT
The plasma membrane represents a critical interface between the internal and extracellular environments, and harbors multiple proteins key receptors and transporters that play important roles in restriction of intracellular infection. We applied plasma membrane profiling, a technique that combines quantitative mass spectrometry with selective cell surface aminooxy-biotinylation, to Bacille Calmette-Guérin (BCG)-infected THP-1 macrophages. We quantified 559 PM proteins in BCG-infected THP-1 cells. One significantly upregulated cell-surface protein was the cholesterol transporter ABCA1. We showed that ABCA1 was upregulated on the macrophage cell-surface following infection with pathogenic mycobacteria and knockdown of ABCA1 resulted in increased mycobacterial survival within macrophages, suggesting that it may be a novel mycobacterial host-restriction factor.

No MeSH data available.


Related in: MedlinePlus

ABCA1 is upregulated in response to mycobacterial infection and is a mycobacterial restriction factor. Flow cytometry histograms (left) and graph (right) showing cell-surface ABCA1 staining of THP-1 macrophages infected by (A) fluorescently labeled live or heat-killed BCG (MOI: 5:1), (B)M. tuberculosis-H37Rv-GFP (MOI: 10:1), (C)M. abscessus-GFP (MOI: 5:1). (D) Relative cell-surface expression of ABCA1 on THP-1 macrophages following lentiviral-mediated knock-down as measured by flow cytometry. (E) Relative survival of M. abscessus following infection of THP-1 macrophages after 44 h following mock or ABCA1 knockdown. (F) Relative survival of M. tuberculosis-H37Rv following infection of THP-1 macrophages after 24 h following mock or ABCA1 knockdown. ∗∗p < 0.01 by Student’s t-test.
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Figure 4: ABCA1 is upregulated in response to mycobacterial infection and is a mycobacterial restriction factor. Flow cytometry histograms (left) and graph (right) showing cell-surface ABCA1 staining of THP-1 macrophages infected by (A) fluorescently labeled live or heat-killed BCG (MOI: 5:1), (B)M. tuberculosis-H37Rv-GFP (MOI: 10:1), (C)M. abscessus-GFP (MOI: 5:1). (D) Relative cell-surface expression of ABCA1 on THP-1 macrophages following lentiviral-mediated knock-down as measured by flow cytometry. (E) Relative survival of M. abscessus following infection of THP-1 macrophages after 44 h following mock or ABCA1 knockdown. (F) Relative survival of M. tuberculosis-H37Rv following infection of THP-1 macrophages after 24 h following mock or ABCA1 knockdown. ∗∗p < 0.01 by Student’s t-test.

Mentions: The cholesterol transporter ABCA1 pumps cholesterol from inside the cell to the outside (Jin et al., 2015) and may therefore alter the cellular content of cholesterol available to mycobacteria. We wished to investigate the functional role for upregulation of ABCA1 on mycobacterial infection. We noted a robust upregulation of ABCA1 following stimulation of activated THP-1 cells by both live and heat-killed BCG (Figure 4A). This was suggestive of a host response to mycobacterial infection. To verify that ABCA1 upregulation was not a generalized response to particle phagocytosis we fed activated THP-1 macrophages with fluorescently labeled beads. Phagocytosis of the beads was associated with a small down-regulation of surface ABCA1 expression (Supplementary Figure S1), suggesting that ABCA1 upregulation was a specific response to stimulation by mycobacterial components.


Plasma Membrane Profiling Reveals Upregulation of ABCA1 by Infected Macrophages Leading to Restriction of Mycobacterial Growth.

Long J, Basu Roy R, Zhang YJ, Antrobus R, Du Y, Smith DL, Weekes MP, Javid B - Front Microbiol (2016)

ABCA1 is upregulated in response to mycobacterial infection and is a mycobacterial restriction factor. Flow cytometry histograms (left) and graph (right) showing cell-surface ABCA1 staining of THP-1 macrophages infected by (A) fluorescently labeled live or heat-killed BCG (MOI: 5:1), (B)M. tuberculosis-H37Rv-GFP (MOI: 10:1), (C)M. abscessus-GFP (MOI: 5:1). (D) Relative cell-surface expression of ABCA1 on THP-1 macrophages following lentiviral-mediated knock-down as measured by flow cytometry. (E) Relative survival of M. abscessus following infection of THP-1 macrophages after 44 h following mock or ABCA1 knockdown. (F) Relative survival of M. tuberculosis-H37Rv following infection of THP-1 macrophages after 24 h following mock or ABCA1 knockdown. ∗∗p < 0.01 by Student’s t-test.
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Figure 4: ABCA1 is upregulated in response to mycobacterial infection and is a mycobacterial restriction factor. Flow cytometry histograms (left) and graph (right) showing cell-surface ABCA1 staining of THP-1 macrophages infected by (A) fluorescently labeled live or heat-killed BCG (MOI: 5:1), (B)M. tuberculosis-H37Rv-GFP (MOI: 10:1), (C)M. abscessus-GFP (MOI: 5:1). (D) Relative cell-surface expression of ABCA1 on THP-1 macrophages following lentiviral-mediated knock-down as measured by flow cytometry. (E) Relative survival of M. abscessus following infection of THP-1 macrophages after 44 h following mock or ABCA1 knockdown. (F) Relative survival of M. tuberculosis-H37Rv following infection of THP-1 macrophages after 24 h following mock or ABCA1 knockdown. ∗∗p < 0.01 by Student’s t-test.
Mentions: The cholesterol transporter ABCA1 pumps cholesterol from inside the cell to the outside (Jin et al., 2015) and may therefore alter the cellular content of cholesterol available to mycobacteria. We wished to investigate the functional role for upregulation of ABCA1 on mycobacterial infection. We noted a robust upregulation of ABCA1 following stimulation of activated THP-1 cells by both live and heat-killed BCG (Figure 4A). This was suggestive of a host response to mycobacterial infection. To verify that ABCA1 upregulation was not a generalized response to particle phagocytosis we fed activated THP-1 macrophages with fluorescently labeled beads. Phagocytosis of the beads was associated with a small down-regulation of surface ABCA1 expression (Supplementary Figure S1), suggesting that ABCA1 upregulation was a specific response to stimulation by mycobacterial components.

Bottom Line: The plasma membrane represents a critical interface between the internal and extracellular environments, and harbors multiple proteins key receptors and transporters that play important roles in restriction of intracellular infection.We quantified 559 PM proteins in BCG-infected THP-1 cells.One significantly upregulated cell-surface protein was the cholesterol transporter ABCA1.

View Article: PubMed Central - PubMed

Affiliation: Collaboration Innovation Centre for the Diagnosis and Treatment of Infectious Diseases, School of Medicine, Tsinghua University Beijing, China.

ABSTRACT
The plasma membrane represents a critical interface between the internal and extracellular environments, and harbors multiple proteins key receptors and transporters that play important roles in restriction of intracellular infection. We applied plasma membrane profiling, a technique that combines quantitative mass spectrometry with selective cell surface aminooxy-biotinylation, to Bacille Calmette-Guérin (BCG)-infected THP-1 macrophages. We quantified 559 PM proteins in BCG-infected THP-1 cells. One significantly upregulated cell-surface protein was the cholesterol transporter ABCA1. We showed that ABCA1 was upregulated on the macrophage cell-surface following infection with pathogenic mycobacteria and knockdown of ABCA1 resulted in increased mycobacterial survival within macrophages, suggesting that it may be a novel mycobacterial host-restriction factor.

No MeSH data available.


Related in: MedlinePlus