A 78-kDa glucose-regulated protein is involved in the decrease of interleukin-6 secretion by lead treatment from astrocytes.
Interleukin (IL)-6 is a cytokine produced mainly by microglia and astrocytes and plays a pleiotropic role in the central nervous system. In this study, we cloned rat IL-6 cDNA into an enhanced green fluorescent protein (EGFP) or a red fluorescent protein (DsRed2) vector and rat 78-kDa glucose-regulated protein (GRP78) cDNA into an EGFP vector to construct IL-6-EGFP, IL-6-DsRed2, and GRP78-EGFP chimeras for the investigation of the mechanism of IL-6 secretion from astrocytes. The data showed that constructed IL-6-EGFP and IL-6-DsRed2 chimeras retained the secretory property, and the secretion of IL-6-EGFP from astrocytes could be attenuated by GRP78 depletion with double-stranded RNA interference. Coexpression of IL-6-DsRed2 and dysfunctional GRP78-EGFP abolished IL-6-DsRed2 secretion, and two chimeric proteins colocalized inside living astrocytes. Coimmunoprecipitation analysis indicated that IL-6 and GRP78 resided in the same complex. The data further revealed that IL-6-EGFP secretion from astrocytes was blocked by the heavy metal lead (Pb) in a concentration-dependent manner. Analysis of the Pb interaction with protein on a Pb-affinity column demonstrated that Pb bound to GRP78 but failed to bind to IL-6. Therefore, these data suggest that IL-6-EGFP or IL-6-DsRed2 chimeras can be used as imaging probes to study IL-6 secretion from living cells, that GRP78 is involved in IL-6 secretion from astrocytes, and that Pb can block IL-6 secretion from astrocytes via targeting GRP78.
Qian
Yongchang
Y
Dept. of Veterinary Integrative Biosciences, Texas A&M Univ., College Station, TX 77843-4458, USA. yqian@cvm.tamu.edu
Zheng
Ying
Y
Weber
Deanna
D
Tiffany-Castiglioni
Evelyn
E
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D002478:Cells, Cultured; D049452:Green Fluorescent Proteins / Q000235:genetics; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism*; D007854:Lead / Q000633:toxicity*; D008164:Luminescent Proteins / Q000235:genetics; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D009498:Neurotoxins / Q000633:toxicity; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D000091342:Endoplasmic Reticulum Chaperone BiP
D049452:Green Fluorescent Proteins; D006360:Heat-Shock Proteins; D015850:Interleukin-6; D007854:Lead; D008164:Luminescent Proteins; D018832:Molecular Chaperones; D009498:Neurotoxins; D011993:Recombinant Fusion Proteins; C051684:GRP78 protein, rat; D000091342:Endoplasmic Reticulum Chaperone BiP; C413663:fluorescent protein 583