TY - JOUR
T1 - Cytosolic and nuclear binding proteins for 2,3,7,8-tetrachlorodibenzo-p-dioxin in the rat thymus
AU - Lund, Johan
AU - Hath Kurl, Rabinder
AU - Poellinger, Lorenz
AU - Gustafsson, Jan Åke
PY - 1982/5/5
Y1 - 1982/5/5
N2 - The existence of a high-affinity, low-capacity 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-binding species was demonstrated in cytosol from rat thymus. It was sensitive to heat and to pronase, trypsin or chymotrypsin but not to DNAase or RNAase, indicating that it was a protein. An excess of unlabelled 2,3,7,8-tetrachlorodibenzofuran or β-naphthoflavone displaced [3H]TCDD from the binder whereas phenobarbital, pregnenolone-16-α-carbonitrile or dexamethasone did not compete. Using a dextra-coated charcoal assay, the apparent dissociation constant (Kd) of the [3H]TCDD-binder complex was determined to 0.36 nM and the apparent maximum amount of binding sites (Bmax) to 68 fmol/mg of cytosolic protein. When analyzed by sucrose density-gradient centrifugation at high ionic strength, the [3H]TCDD-binder complex sedimented at 4-5 S; at low ionic strength the complex sedimented more rapidly, probably due to aggregation. All these data support the interpretation that the demonstrated cytosolic TCDD-binder represents the receptor protein for TCDD, as previously described for rat and mouse liver. Following intravenous administration of [3H]TCDD, a low-capacity [3H]TCDD-macromolecule complex was extractable from thymic cell neuclei; this complex behaved identically to the cytosolic [3H]TCDD-receptor complex when exposed to heat or to hydrolytic enzymes and was therefore alos identified as a protein. The nuclear [3H]TCDD-protein complex sedimented at 4-5 S at high ionic strength. Furthermore, a maximum uptake of [3H]TCDD in thymic nuclei was observed simultaneously with a decline in cytosolic radioactivity (at 3 h post-injection). These findings suggest that the nuclear [3H]TCDD-protein complex represented [3H]TCDD-receptor complex translocated from the cytoplasm. In conclusion, the rat thymus contains a cytosolic TCDD receptor at a concentration similar to that of the rat hepatic receptor. However, in vivo experiments showed that the nuclear uptake of [3H]TCDD (expressed as dpm/mg GNA) in the thymus was only about 6% of that in liver. Further studies are needed for an understanding of the mechanism behind this discrepancy.
AB - The existence of a high-affinity, low-capacity 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-binding species was demonstrated in cytosol from rat thymus. It was sensitive to heat and to pronase, trypsin or chymotrypsin but not to DNAase or RNAase, indicating that it was a protein. An excess of unlabelled 2,3,7,8-tetrachlorodibenzofuran or β-naphthoflavone displaced [3H]TCDD from the binder whereas phenobarbital, pregnenolone-16-α-carbonitrile or dexamethasone did not compete. Using a dextra-coated charcoal assay, the apparent dissociation constant (Kd) of the [3H]TCDD-binder complex was determined to 0.36 nM and the apparent maximum amount of binding sites (Bmax) to 68 fmol/mg of cytosolic protein. When analyzed by sucrose density-gradient centrifugation at high ionic strength, the [3H]TCDD-binder complex sedimented at 4-5 S; at low ionic strength the complex sedimented more rapidly, probably due to aggregation. All these data support the interpretation that the demonstrated cytosolic TCDD-binder represents the receptor protein for TCDD, as previously described for rat and mouse liver. Following intravenous administration of [3H]TCDD, a low-capacity [3H]TCDD-macromolecule complex was extractable from thymic cell neuclei; this complex behaved identically to the cytosolic [3H]TCDD-receptor complex when exposed to heat or to hydrolytic enzymes and was therefore alos identified as a protein. The nuclear [3H]TCDD-protein complex sedimented at 4-5 S at high ionic strength. Furthermore, a maximum uptake of [3H]TCDD in thymic nuclei was observed simultaneously with a decline in cytosolic radioactivity (at 3 h post-injection). These findings suggest that the nuclear [3H]TCDD-protein complex represented [3H]TCDD-receptor complex translocated from the cytoplasm. In conclusion, the rat thymus contains a cytosolic TCDD receptor at a concentration similar to that of the rat hepatic receptor. However, in vivo experiments showed that the nuclear uptake of [3H]TCDD (expressed as dpm/mg GNA) in the thymus was only about 6% of that in liver. Further studies are needed for an understanding of the mechanism behind this discrepancy.
KW - (Rat)
KW - Dextran-coated charcoal
KW - Tetrachlorodibenzodioxin receptor
KW - Thymic atrophy
KW - Toxin
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UR - http://www.scopus.com/inward/citedby.url?scp=0020318406&partnerID=8YFLogxK
U2 - 10.1016/0304-4165(82)90197-0
DO - 10.1016/0304-4165(82)90197-0
M3 - Article
C2 - 6284245
AN - SCOPUS:0020318406
VL - 716
SP - 16
EP - 23
JO - Biochimica et Biophysica Acta - General Subjects
JF - Biochimica et Biophysica Acta - General Subjects
SN - 0304-4165
IS - 1
ER -