The Journal of Neuroscience, August 1, 2000, 20(15):5696-5702
Clinical Mutations in the L1 Neural Cell Adhesion Molecule Affect
Cell-Surface Expression
Hugh D.
Moulding,
Robert L.
Martuza, and
Samuel D.
Rabkin
Interdisciplinary Program for Neuroscience and Department of
Neurosurgery, Georgetown University Medical Center, Washington, DC
20007
Mutations in the L1 neural cell adhesion molecule, a transmembrane
glycoprotein, cause a spectrum of congenital neurological syndromes,
ranging from hydrocephalus to mental retardation. Many of these
mutations are single amino acid changes that are distributed throughout
the various domains of the protein. Defective herpes simplex virus
vectors were used to express L1 protein with the clinical missense
mutations R184Q and D598N in the Ig2 and Ig6 extracellular domains,
respectively, and S1194L in the cytoplasmic domain. All three mutant
proteins were expressed at similar levels in infected cells. Neurite
outgrowth of cerebellar granule cells was stimulated on astrocytes
expressing wild-type or S1194L L1, whereas those expressing R184Q and
D598N L1 failed to increase neurite length. Live cell immunofluorescent
staining of L1 demonstrated that most defective vector-infected cells
did not express R184Q or D598N L1 on their cell surface. This greatly
diminished cell-surface expression occurred in astrocytes, neurons, and
non-neural cells. In contrast to wild-type or S1194L L1, the R184Q and
D598N L1 proteins had altered apparent molecular weights and remained
completely endoglycosidase H (endoH)-sensitive, suggesting incomplete
post-translational processing. We propose that some missense mutations
in human L1 impede correct protein trafficking, with functional
consequences independent of protein activity. This provides a rationale
for how expressed, full-length proteins with single amino acid changes could cause clinical phenotypes similar in severity to knock-out mutants.
Key words:
gene transfer; herpes simplex virus; protein trafficking; membrane glycoproteins; mental retardation; neurite outgrowth
Copyright © 2000 Society for Neuroscience 0270-6474/00/20155696-07$05.00/0
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