The insulin-like growth factor II/mannose-6-phosphate receptor is present in fetal and maternal sheep serum.
A large mol wt binding protein for insulin-like growth factor II (IGF-II) has been described in fetal sheep serum. We now provide evidence to demonstrate that this binding protein is the IGF-II/mannose-6-phosphate (Man-6-P) receptor. Serum and plasma were gel filtered on Sephadex G-200, and the column fractions were assayed for binding of radiolabeled IGF-II. There was significant binding of [125I]IGF-II to the void volume fractions in addition to binding to the 150K and 40K carrier proteins. Binding to the void volume fractions was increased in fetal serum as well as maternal serum and dramatically decreased in the nonpregnant adult. Competitive binding studies with [125I]IGF-II and the void volume pools from fetal and maternal sheep serum demonstrated that IGF-I competed less potently than IGF-II, and insulin did not compete. There was no specific binding of [125I]IGF-I to the void volume pools of either fetal or maternal samples. Chemical cross-linking of [125I]IGF-II to aliquots of the void volume pools from fetal and maternal sheep serum samples and analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the presence of dithiothreitol demonstrated a specific band at about 240K. Western blotting using a specific antiserum (no. 3637) against rat IGF-II/Man-6-P receptor was performed on aliquots of the Sephadex G-200 void volume pools of fetal, maternal, uterine vein, and adult sheep serum; a band of approximately 210K (without dithiothreitol) was seen. The IGF-II/Man-6-P receptor band was more intense in fetal serum than in either maternal or adult nonpregnant sheep serum. There was also increased binding of [125I]IGF-II in the 40K region of the Sephadex G-200 column fractions in the maternal serum compared to that in serum from nonpregnant adult ewes. When fetal, maternal, and adult nonpregnant sheep serum Sephadex G-200 pools were gel filtered on Sephadex G-50 in 1 mol/liter acetic acid to separate bound from free IGF, and IGF-II was measured by RRA, approximately 50% of the circulating IGF-II was associated with this IGF-II/Man-6-P receptor in fetal serum compared to 7% in maternal serum and 3% in adult nonpregnant serum. These data demonstrate that the IGF-II/Man-6-P receptor circulates in fetal sheep serum as well as maternal serum and appears to be a significant carrier for IGF-II in fetal sheep.
Gelato
M C
MC
Division of Endocrinology, Health Sciences Center, State University of New York, Stony Brook 11794.
Rutherford
C
C
Stark
R I
RI
Daniel
S S
SS
D000818:Animals; D002850:Chromatography, Gel; D005260:Female; D005312:Fetal Blood / Q000378:metabolism*; D006600:Hexosephosphates / Q000097:blood*; D015151:Immunoblotting; D007335:Insulin-Like Growth Factor II / Q000097:blood*; D008360:Mannosephosphates / Q000097:blood*; D008970:Molecular Weight; D011247:Pregnancy; D011270:Pregnancy, Animal / Q000097:blood*; D017527:Receptor, IGF Type 2; D011956:Receptors, Cell Surface / Q000097:blood* / Q000302:isolation & purification / Q000378:metabolism*; D017451:Receptors, Somatomedin; D012756:Sheep; D013002:Somatomedins / Q000097:blood*
D006600:Hexosephosphates; D008360:Mannosephosphates; D017527:Receptor, IGF Type 2; D011956:Receptors, Cell Surface; D017451:Receptors, Somatomedin; D013002:Somatomedins; D007335:Insulin-Like Growth Factor II