Recovery of Soluble Sheep Erythrocyte Receptor from the T Lymphocyte Surface by Proteolytic Cleavage

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Abstract
Proteolytic digestion of the human T lymphoblastoid cell line (Molt-4) and of peripheral blood lymphocytes by trypsin, chymotrypsin, and pronase results in a progressive, time- and dose-dependent diminution of T lymphocyte — sheep red blood cell (SRBC) rosette formation, whereas thrombin, plasmin, collagenase, DNAse, and phospholipase have no effect. Complete abrogation of SRBC binding is achieved when lymphocytes (1 × 108/ml) are incubated with either trypsin or chymotrypsin at 10 μg/ml for 30 min, and τ;50% abrogation is observed between 3 to 10 min. Preincubation of SRBC with the 10 min and 20 min lymphocyte digest supernatants inhibited their subsequent binding by normal T lymphocytes by as much as 64%. Thirty-minute digests were less inhibitory. Equivalent digests from several human B lumphoblastoid cell lines and from a non-rosetting clone of Molt-4 cells were not inhibitory. Polyacrylamide gel electrophoresis followed by elution of serial gel slices revealed four distinct inhibitory bands (I–IV) in the 20-min digest supernatant whereas only bands I–III and band IV were present in the 10-min and 30-min digest supernatants, respectively, suggesting progressive proteolysis of a distinct receptor. These experiments indicate that the binding of SRBC by human T lymphocytes represents a receptor-ligand interaction rather than a nonspecific electrical charge phenomenon and that the receptor is a discrete molecular species which can be isolated from the surface of T but not B lymphocytes by limited enzymatic proteolysis.