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Leishmania donovani infection enhances lateral mobility of macrophage membrane protein which is reversed by liposomal cholesterol.

Ghosh M, Roy K, Das Mukherjee D, Chakrabarti G, Roy Choudhury K, Roy S - PLoS Negl Trop Dis (2014)

Bottom Line: The protozoan parasite Leishmania donovani (LD) reduces cellular cholesterol of the host possibly for its own benefit.To our knowledge this is the first direct demonstration that LD parasites during their intracellular life cycle increases lateral mobility of membrane proteins and decreases F-actin level in infected macrophages.Such defects may contribute to ineffective intracellular signaling and other cellular functions.

View Article: PubMed Central - PubMed

Affiliation: Infectious Diseases and Immunology Division, CSIR-Indian Institute of Chemical Biology, Kolkata, India.

ABSTRACT

Background: The protozoan parasite Leishmania donovani (LD) reduces cellular cholesterol of the host possibly for its own benefit. Cholesterol is mostly present in the specialized compartment of the plasma membrane. The relation between mobility of membrane proteins and cholesterol depletion from membrane continues to be an important issue. The notion that leishmania infection alters the mobility of membrane proteins stems from our previous study where we showed that the distance between subunits of IFNγ receptor (R1 and R2) on the cell surface of LD infected cell is increased, but is restored to normal by liposomal cholesterol treatment.

Methodology/principal findings: We determined the lateral mobility of a membrane protein in normal, LD infected and liposome treated LD infected cells using GFP-tagged PLCδ1 as a probe. The mobility of PLCδ1 was computationally analyzed from the time lapse experiment using boundary distance plot and radial profile movement. Our results showed that the lateral mobility of the membrane protein, which is increased in infection, is restored to normal upon liposomal cholesterol treatment. The results of FRAP experiment lent further credence to the above notion. The membrane proteins are intimately linked with cellular actin and alteration of cellular actin may influence lateral mobility. We found that F-actin is decreased in infection but is restored to normal upon liposomal cholesterol treatment as evident from phalloidin staining and also from biochemical analysis by immunoblotting.

Conclusions/significances: To our knowledge this is the first direct demonstration that LD parasites during their intracellular life cycle increases lateral mobility of membrane proteins and decreases F-actin level in infected macrophages. Such defects may contribute to ineffective intracellular signaling and other cellular functions.

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Related in: MedlinePlus

Radial distance map of a cell.A) Blue line shows cell boundary (radial distance = 1). Red line show contours of equal radial distance within the cell (radial distance <1). Green lines show contours of equal radial distance outside the cell (radial distance >1). B) Plot of the distribution of PLCδ1-GFP expression as a function of distance from center of the cell. The red line shows the average activity as a function of distance, obtained via a smoothing line. The x-axis (radial distance) is truncated at 1.5 to avoid contamination of PLCδ1-GFP expression from other cells.
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pntd-0003367-g002: Radial distance map of a cell.A) Blue line shows cell boundary (radial distance = 1). Red line show contours of equal radial distance within the cell (radial distance <1). Green lines show contours of equal radial distance outside the cell (radial distance >1). B) Plot of the distribution of PLCδ1-GFP expression as a function of distance from center of the cell. The red line shows the average activity as a function of distance, obtained via a smoothing line. The x-axis (radial distance) is truncated at 1.5 to avoid contamination of PLCδ1-GFP expression from other cells.

Mentions: The expression of PLCδ1 in N-MΦ, I-MΦ and I-MΦ-CL was quantified by computational method. The boundary distance co-ordinates were computed for each cell using the Euclidean distance transform after presmoothing and oversampling of cellular boundaries to normalize with respect to variations in cell shape and size. The general shape profile based on GFP expression in N-MΦ, I-MΦ and I-MΦ-CL were similar (Fig. 2A). An adaptive piecewise linear model is used to compare expression gradients in intra, peri and extra cellular zones.We have considered the center of the cell as ‘0’ and the cell surface as ‘1’ on the x-axis. The distribution of PLCδ1 are relatively flat in the interior of the cell (between 0 and 1 on the axis), followed by a sharp rise at the plasma membrane (at 1 on the x-axis) and finally a sharp drop off (points >1 on the x-axis) outside the cell (Fig. 2B). The average expression curves show the same basic spatial distribution in N-MΦ, I-MΦ and I-MΦ-CL: low near the center of the cell, peak near the boundary and then a drop to ∼‘0’ some distance outside the boundary (Fig. 3).


Leishmania donovani infection enhances lateral mobility of macrophage membrane protein which is reversed by liposomal cholesterol.

Ghosh M, Roy K, Das Mukherjee D, Chakrabarti G, Roy Choudhury K, Roy S - PLoS Negl Trop Dis (2014)

Radial distance map of a cell.A) Blue line shows cell boundary (radial distance = 1). Red line show contours of equal radial distance within the cell (radial distance <1). Green lines show contours of equal radial distance outside the cell (radial distance >1). B) Plot of the distribution of PLCδ1-GFP expression as a function of distance from center of the cell. The red line shows the average activity as a function of distance, obtained via a smoothing line. The x-axis (radial distance) is truncated at 1.5 to avoid contamination of PLCδ1-GFP expression from other cells.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256160&req=5

pntd-0003367-g002: Radial distance map of a cell.A) Blue line shows cell boundary (radial distance = 1). Red line show contours of equal radial distance within the cell (radial distance <1). Green lines show contours of equal radial distance outside the cell (radial distance >1). B) Plot of the distribution of PLCδ1-GFP expression as a function of distance from center of the cell. The red line shows the average activity as a function of distance, obtained via a smoothing line. The x-axis (radial distance) is truncated at 1.5 to avoid contamination of PLCδ1-GFP expression from other cells.
Mentions: The expression of PLCδ1 in N-MΦ, I-MΦ and I-MΦ-CL was quantified by computational method. The boundary distance co-ordinates were computed for each cell using the Euclidean distance transform after presmoothing and oversampling of cellular boundaries to normalize with respect to variations in cell shape and size. The general shape profile based on GFP expression in N-MΦ, I-MΦ and I-MΦ-CL were similar (Fig. 2A). An adaptive piecewise linear model is used to compare expression gradients in intra, peri and extra cellular zones.We have considered the center of the cell as ‘0’ and the cell surface as ‘1’ on the x-axis. The distribution of PLCδ1 are relatively flat in the interior of the cell (between 0 and 1 on the axis), followed by a sharp rise at the plasma membrane (at 1 on the x-axis) and finally a sharp drop off (points >1 on the x-axis) outside the cell (Fig. 2B). The average expression curves show the same basic spatial distribution in N-MΦ, I-MΦ and I-MΦ-CL: low near the center of the cell, peak near the boundary and then a drop to ∼‘0’ some distance outside the boundary (Fig. 3).

Bottom Line: The protozoan parasite Leishmania donovani (LD) reduces cellular cholesterol of the host possibly for its own benefit.To our knowledge this is the first direct demonstration that LD parasites during their intracellular life cycle increases lateral mobility of membrane proteins and decreases F-actin level in infected macrophages.Such defects may contribute to ineffective intracellular signaling and other cellular functions.

View Article: PubMed Central - PubMed

Affiliation: Infectious Diseases and Immunology Division, CSIR-Indian Institute of Chemical Biology, Kolkata, India.

ABSTRACT

Background: The protozoan parasite Leishmania donovani (LD) reduces cellular cholesterol of the host possibly for its own benefit. Cholesterol is mostly present in the specialized compartment of the plasma membrane. The relation between mobility of membrane proteins and cholesterol depletion from membrane continues to be an important issue. The notion that leishmania infection alters the mobility of membrane proteins stems from our previous study where we showed that the distance between subunits of IFNγ receptor (R1 and R2) on the cell surface of LD infected cell is increased, but is restored to normal by liposomal cholesterol treatment.

Methodology/principal findings: We determined the lateral mobility of a membrane protein in normal, LD infected and liposome treated LD infected cells using GFP-tagged PLCδ1 as a probe. The mobility of PLCδ1 was computationally analyzed from the time lapse experiment using boundary distance plot and radial profile movement. Our results showed that the lateral mobility of the membrane protein, which is increased in infection, is restored to normal upon liposomal cholesterol treatment. The results of FRAP experiment lent further credence to the above notion. The membrane proteins are intimately linked with cellular actin and alteration of cellular actin may influence lateral mobility. We found that F-actin is decreased in infection but is restored to normal upon liposomal cholesterol treatment as evident from phalloidin staining and also from biochemical analysis by immunoblotting.

Conclusions/significances: To our knowledge this is the first direct demonstration that LD parasites during their intracellular life cycle increases lateral mobility of membrane proteins and decreases F-actin level in infected macrophages. Such defects may contribute to ineffective intracellular signaling and other cellular functions.

Show MeSH
Related in: MedlinePlus