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Restoration of IFNcR Subunit Assembly, IFNc Signaling and Parasite Clearance in Leishmania donovani Infected Macrophages: Role of Membrane Cholesterol

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Title Restoration of IFNcR Subunit Assembly, IFNc Signaling
and Parasite Clearance in Leishmania donovani Infected
Macrophages: Role of Membrane Cholesterol
 
Creator Sen, Subha
Roy, Koushik
Mukherjee, Sandip
Mukhopadhyay, Rupkatha
Roy, Syamal
 
Subject Infectious Diseases and Immunology
 
Description Despite the presence of significant levels of systemic Interferon gamma (IFNc), the host protective cytokine, Kala-azar
patients display high parasite load with downregulated IFNc signaling in Leishmania donovani (LD) infected macrophages
(LD-MØs); the cause of such aberrant phenomenon is unknown. Here we reveal for the first time the mechanistic basis of
impaired IFNc signaling in parasitized murine macrophages. Our study clearly shows that in LD-MØs IFNc receptor (IFNcR)
expression and their ligand-affinity remained unaltered. The intracellular parasites did not pose any generalized defect in
LD-MØs as IL-10 mediated signal transducer and activator of transcription 3 (STAT3) phosphorylation remained unaltered
with respect to normal. Previously, we showed that LD-MØs are more fluid than normal MØs due to quenching of
membrane cholesterol. The decreased rigidity in LD-MØs was not due to parasite derived lipophosphoglycan (LPG) because
purified LPG failed to alter fluidity in normal MØs. IFNcR subunit 1 (IFNcR1) and subunit 2 (IFNcR2) colocalize in raft upon
IFNc stimulation of normal MØs, but this was absent in LD-MØs. Oddly enough, such association of IFNcR1 and IFNcR2
could be restored upon liposomal delivery of cholesterol as evident from the fluorescence resonance energy transfer (FRET)
experiment and co-immunoprecipitation studies. Furthermore, liposomal cholesterol treatment together with IFNc allowed
reassociation of signaling assembly (phospho-JAK1, JAK2 and STAT1) in LD-MØs, appropriate signaling, and subsequent
parasite killing. This effect was cholesterol specific because cholesterol analogue 4-cholestene-3-one failed to restore the
response. The presence of cholesterol binding motifs [(L/V)-X1–5-Y-X1–5-(R/K)] in the transmembrane domain of IFNcR1 was
also noted. The interaction of peptides representing this motif of IFNcR1 was studied with cholesterol-liposome and
analogue-liposome with difference of two orders of magnitude in respective affinity (KD: 4.2761029 M versus
2.6961027 M). These observations reinforce the importance of cholesterol in the regulation of function of IFNcR1 proteins.
This study clearly demonstrates that during its intracellular life-cycle LD perturbs IFNcR1 and IFNcR2 assembly and
subsequent ligand driven signaling by quenching MØ membrane cholesterol.
 
Date 2011
 
Type Article
PeerReviewed
 
Format application/pdf
 
Identifier http://www.eprints.iicb.res.in/1339/1/PLOS_PATHOGENS___7_(_9)Article_Number_e1002229;2011[42].pdf
Sen, Subha and Roy, Koushik and Mukherjee, Sandip and Mukhopadhyay, Rupkatha and Roy, Syamal (2011) Restoration of IFNcR Subunit Assembly, IFNc Signaling and Parasite Clearance in Leishmania donovani Infected Macrophages: Role of Membrane Cholesterol. PLoS Pathogens, 7 (9).
 
Relation http://dx.doi.org/10.1371/journal.ppat.1002229
http://www.eprints.iicb.res.in/1339/