Student Theses and Dissertations

Date of Award

1991

Document Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

RU Laboratory

Edelman/Cunningham Laboratory

Abstract

The neuron-glia cell adhesion molecule, Ng-CAM, is a cell-surface glycoprotein expressed on neurons and Schwann cells in the chicken nervous system. It is expressed as a group of related components of -200 kD, 135 kD, and 80 kD, which are detected at particular sites during development. Ng-CAM mediates a variety of functions that are known to be important in specific cell-cell interactions and the histogenesis of the nervous system, including neuron-neuron adhesion, neuron-glia adhesion, neurite fasciculation, and neuronal migration on glial cells. This thesis describes the characterization of cDNA that encode the entire Ng-CAM protein and the determination of the origin of the polypeptide components. An analysis of the deduced amino acid sequence reveals the presence of immunoglobulin-like domains and fibronectin-type III repeats, and establishes the relationship of Ng-CAM to other neural CAMs containing similar structures. The thesis also describes the identification of a similar protein in the embryonic chicken nervous system, the Ng-CAM-related cell adhesion molecule (Nr-CAM). Its close structural and sequence similarities to Ng-CAM indicate its probable involvement in cell adhesion, and defines a closely related subfamily of neural CAMs. cDNA libraries prepared from embryonic nervous tissue were used to isolate clones encoding Ng-CAM. These clones encode a polypeptide that includes all three known components of Ng-CAM, and hybridize to a single mRNA species (6 kb) that is restricted to nervous tissue. Single bands were detected in genomic DNA blots with Ng-CAM cDNA probes, indicating that there is a single Ng-CAM gene in chicken. Together these data indicate that the polypeptide components of Ng-CAM all arise from a single translation product, and that the two smaller poljrpeptide components (135 kD and 80 kD) are products of posttranslational cleavage. The deduced amino acid sequence of Ng-CAM predicts the presence of six immunoglobulin-like domains, five fibronectin-type III repeats, a single transmembrane domain, and a short cytoplasmic region. Ng-CAM also contains a single RGD sequence in its third fibronectin-type III repeat, which in an equivalent position in several other proteins is known to mediate a cell-binding activity. Ng-CAM is similar to other neural CAMs containing immunoglobulin-like domains and is most similar to its previously presumed equivalent in mouse, Ll. However, overall the amino acid sequence of Ng-CAM is only 40% identical to Ll, and within the individual domains ranges from only 15% to 66% identity. Known equivalent CAMs (e.g., N-CAM) in the two species are much more closely related (80% identity), indicating that in contrast to previous assumptions Ng-CAM and Ll may be functionally different molecules. cDNA clones encoding another protein were isolated from embryonic chicken nervous system libraries using antibodies against Ng-CAM. The new protein shares at least one polypeptide epitope with Ng-CAM, and was designated the Ng-CAM-related cell adhesion molecule, or Nr-CAM. Like Ng-CAM, it is predicted to contain six immunoglobulin-like domains, five fibronectin-type III repeats, one transmembrane segment and a small cytoplasmic domain, each of which is most similar to corresponding segments in Ng-CAM and Ll. However, overall Nr-CAM shares only 40% identity to Ll, and is thus not the chicken equivalent of Ll. In addition, comparison of several similar Nr-CAM cDNA clones indicated that the fifth fibronectin-type III repeat (93 amino acids) and a segment between the second and third immunoglobulin-like domains (20 amino acids) may be differentially spliced in Nr-CAM mRNA; such alternative forms have not been seen for Ng-CAM and Ll. The combined results indicate that Ng-CAM, Nr-CAM and Ll define a structurally related subfamily of neural CAMs. Their similarities suggest that they may perform similar binding functions in the nervous system, possibly at different times and locations during neural development.

Comments

A thesis submitted to the faculty of the Rockefeller University in partial fulfillment of the requirements for the degree of Doctor of Philosophy

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