Isolation and characterization of peanut agglutinin-resistant embryonal carcinoma cell-surface variants

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Abstract
The isolation and characterization of variant embryonal carcinoma (EC) cells possessing altered cell‐surface structures is described. The lectin peanut agglutinin (PNA), which binds to EC cells but not their differentiated derivatives, was used to select the variants. Clones resistant to the cytotoxic effect of PNA were isolated at a frequency of 4 × 10–5 following mutagenesis. The resistant phenotype was stable in the absence of selection in all eight clones tested. The increased frequency of resistant clones following mutagenesis and the stability of the phenotype suggests a mutational origin. Somatic cell hybrids constructed between wild‐type cells and two different PNA‐resistant cell lines were sensitive to PNA; this suggests that the resistant phenotype is recessive. Binding assays demonstrated that resistant cells exhibited a twofold to fourfold reduction in the total amount of PNA bound. Together with the recessive behavior of the phenotype, this suggests that resistant cells are deficient for PNA receptors. The PNA‐resistant cells also showed reduced binding of monoclonal antibody against stage‐specific embryonic antigen 1 (SSEA–1) in indirect cytotoxicity tests. All eight PNA‐resistant lines isolated were tumorigenic in syngeneic mice and gave rise to well‐differentiated teratocarcinomas. The PNA‐resistant cells behaved like their wild‐type parents in a cell recognition assay; when incubated in suspension with endodermal cells, they sorted out to form simple embryoid bodies (a core of EC cells surrounded by an endodermal rind). Thus, EC cells can form tumors, differentiate, and recognize differentiated cells in a sorting assay despite a reduction in expression of the embryo‐specific cell surface structures (s) that bind PNA and anti‐SSEA‐1 antibody.