Identification and purification of a receptor on macrophages for the dengue virus-induced suppressor cytokine.
Dengue type 2 virus (DV)-induced suppressor cytokine (SF) binds to macrophages to transmit the suppressor signal to recruit the second subpopulation of suppressor T cells. The present study was undertaken to identify and purify the receptor for SF (SF-R) on macrophages. The binding of 125I-SF to macrophages was saturable and reversible. Scatchard analysis showed the presence of both high (54,000/cell) and low (1.78 x 10(6)/cell) affinity receptor sites. The binding of 125I-SF to macrophages was inhibited by pretreatment of macrophages with anti-SF antiserum but not by a heterologous antiserum. Normal mouse peritoneal macrophage membrane was solubilized with Triton-X-100 and the components separated by low pressure liquid chromatography (LPLC) to purify SF-R. The presence of SF binding moiety (SF-R) was screened at each step of purification. The purified SF-R resolved into two bands of 45-50 kD mol. wt on SDS-PAGE. 125I-SF+SF-R complex run on SDS-PAGE showed a single band at about 55-60 kD mol. wt by autoradiography. Anti-SF-R antiserum reacted with SF-R in a Western blot test; the reaction was abolished by pretreatment of the blots with proteinase K, but not by pretreatment with periodic acid. SF-R was composed of two polypeptide chains (alpha and beta) which were obtained in pure form by high performance liquid chromatography (HPLC) of dithiothreitol- and iodoacetamide-treated SF-R. Only the beta chain bound SF.
Mukherjee
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R
Postgraduate Department of Microbiology, K. G. Medical College, Lucknow, India.
Chaturvedi
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P
Chaturvedi
U C
UC
D000818:Animals; D016207:Cytokines / Q000378:metabolism*; D003716:Dengue Virus / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D007106:Immune Sera / Q000276:immunology; D015151:Immunoblotting; D008264:Macrophages / Q000737:chemistry*; D051379:Mice; D011956:Receptors, Cell Surface / Q000302:isolation & purification*; D013491:Suppressor Factors, Immunologic / Q000378:metabolism*
D016207:Cytokines; D007106:Immune Sera; D011956:Receptors, Cell Surface; D013491:Suppressor Factors, Immunologic