cDNA clones of the neural cell adhesion molecule (N-CAM) lacking a membrane-spanning region consistent with evidence for membrane attachment via a phosphatidylinositol intermediate.

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Abstract
In embryonic chicken brains, the neural cell adhesion molecule N-CAM is expressed mainly as two polypeptides, the large intracellular-domain polypeptide (Id), (Mr=160,000) and the small intracellular-domain polypeptide (sd) (Mr=130,000) chains, that differ in their cytoplasmic domains and that arise by alternative splicing of RNA transcribed from a single gene. There is evidence for a minor N-CAM polypeptide of Mr=120,000 that is similar to the Id and sd chains for most of its amino-terminal sequence, but which lacks a cytoplasmic domain. We report here the isolation and characterization of a cDNA clone, .lambda.N151, that appears to encode this third N-CAM polypeptide, which we designate the ssd (small surface-domain) polypeptide chain. The cDNA insert of .lambda.N151 consists of 2437 base pairs (bp). DNA hybridization and sequencing indicate that the first 1721 bp are nearly identical to the corresponding sequence of clone .lambda.N208, which encodes the Id chain. Following in the same reading frame, .lambda.N151 encodes 25 amino acids not present in .lambda.N208. The rest of .lambda.N151 consists of a 637-bp noncoding region containing an AATACA polyadenylylation sequence and a 55-bp poly(A) tract. Messenger RNAs complementary to .lambda.N151 appear later in development than those complementary to the Id and sd chains, and their appearance is correlated with the appearance of the ssd polypeptide. Although the polypeptide encoded by .lambda.N151 lacks a membrane region that would define a cytoplasmic domain, it does contain at its carboxyl end a relatively hydrophobic stretch of amino acids similar to those seen in precursors of membrane proteins that are attached to membranes via the lipid phosphatidylinositol. We show here that the ssd chain of chicken N-CAM can be released from brain vesicles by treatment with phospholipase C, suggesting that it too may have a phosphatidylinositol anchor. These results define two additional modes by which N-CAM expression can be modulated: by RNA splicing at a new site and by differential membrane attachment of the resulting polypeptide through a lipid intermediate.