Studies on IL-1 receptors on D10S T-helper cells: demonstration of two molecularly and antigenically distinct IL-1 binding proteins.
Receptor binding studies were performed on the interleukin-1 (IL-1) sensitive T-helper cell line D10S, a stable line which proliferates to subfemtomolar concentrations of IL-1 in the absence of mitogens. IL-1 binds in a specific and saturable manner and Scatchard analysis at 4 degrees C reveals one class of binding affinity. On D10S cells, the Kd for IL-1 is 227 pM +/- 80, with 11,000 (range 3,300 to 23,800) sites per cell. EL4.6.1 cells, which are less sensitive to IL-1, bind with a single class of high affinity sites (55 pM; 4,000 sites). D10S cells incubated 18 h with IL-1 display reduced IL-1 receptor (IL-1R) numbers and affinities, consistent with reduced (75%, p less than 0.005) proliferation to subsequent IL-1; preincubation with IL-4 increases the number of IL-1R which is associated with increased (200%, p less than 0.001) proliferation to IL-1. The molecular mass of the major (80 kD) IL-1R binding [125I]IL-1 alpha on D10S cells was consistently observed at 73 kD as compared to the 80 kD molecule on the EL4 cells. On the other hand, crosslinking studies with [125I]IL-1 beta on D10S cells revealed a novel 46 kD band on gradient SDS-PAGE corresponding to a binding protein of 29 to 30 kD, which is antigenically distinct from the 80 kD IL-1R. Crosslinking of D10S or EL4 cells at 4 degrees C in the presence of phytohemagglutinin (PHA) and labeled IL-1 enhanced the appearance of the 30 kD IL-1 binding protein. The findings are consistent with a two-chain model for the IL-1R, although Scatchard analysis did not consistently indicate two classes of affinities. IL-1 binding to the 80 kD protein may form a heteroduplex with the 30 kD IL-1R which could account for the presence of the 120 to 130 kD IL-1 crosslinked proteins observed by several investigators.
Savage
N
N
Department of Medicine, Tufts University School of Medicine, Boston, MA 02111.
Puren
A J
AJ
Orencole
S F
SF
Ikejima
T
T
Clark
B D
BD
Dinarello
C A
CA
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002452:Cell Count; D002455:Cell Division; D002460:Cell Line; D007375:Interleukin-1 / Q000378:metabolism* / Q000494:pharmacology; D007376:Interleukin-2 / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology; D051379:Mice; D011971:Receptors, Immunologic / Q000378:metabolism*; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins / Q000494:pharmacology; D006377:T-Lymphocytes, Helper-Inducer / Q000378:metabolism*
D007375:Interleukin-1; D007376:Interleukin-2; D011971:Receptors, Immunologic; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins; D015847:Interleukin-4