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Stable expression of acetylcholinesterase and associated collagenic subunits in transfected RBL cell lines: production of GPI-anchored dimers and collagen-tailed forms.
- Source :
-
European journal of cell biology [Eur J Cell Biol] 1995 Jul; Vol. 67 (3), pp. 254-60. - Publication Year :
- 1995
-
Abstract
- We obtained a stable expression of acetylcholinesterase (AChE, E.C. 3.1.1.7) in the rat basoleukemia cell line, RBL-2H3, which possesses a well developed secretory pathway, but expresses only very little endogenous AChE. Metabolic labeling showed that AChEH and AChET, differing by C-terminal peptides encoded by alternatively spliced exons, were synthesized at a similar rate. When transfected with AChEH, RBL cells efficiently produced GPI-anchored dimers, which were mostly exposed at the cell surface, as shown both by activity and immunofluorescence labeling. In contrast, when transfected with AChET, RBL cells produced about tenfold less activity, which was essentially retained in the cell, and the enzyme could not be detected at the cell surface by immunolabeling. The fate of the enzyme is therefore determined by its C-terminal alternative peptides. We were also able to coexpress the AChET subunit with the collagenic Q subunit. The cells produced small but significant amounts of collagen-tailed forms, essentially A4. The expression of these different catalytic and structural subunits in stably transfected RBL cells will be useful to explore the regulated posttranslational processes involved in protein maturation and transport.
- Subjects :
- Acetylcholinesterase genetics
Animals
Base Sequence
Cell Compartmentation
Cell Fractionation
Cell Membrane Permeability
Centrifugation, Density Gradient
Fluorescent Antibody Technique
Leukemia, Basophilic, Acute
Molecular Sequence Data
Precipitin Tests
Protein Conformation
Rats
Recombinant Proteins biosynthesis
Transfection
Tumor Cells, Cultured
Acetylcholinesterase biosynthesis
Collagen biosynthesis
Glycosylphosphatidylinositols biosynthesis
Subjects
Details
- Language :
- English
- ISSN :
- 0171-9335
- Volume :
- 67
- Issue :
- 3
- Database :
- MEDLINE
- Journal :
- European journal of cell biology
- Publication Type :
- Academic Journal
- Accession number :
- 7588881