Selective enrichment for temperature-sensitive secretion mutants of mammalian cells using plant lectin, concanavalin A.
The use of conditional mutants as a genetic approach to study protein secretion in mammalian cells requires the isolation of a large number of mutants. Because a procedure for the direct selection of mutants with secretion defects is not available, their isolation depends upon the selective enrichment of mutant phenotypes in a cell population. We have devised an enrichment strategy in which rat hepatoma cells unable to replace surface membrane receptors of a plant lectin, concanavalin A, are resistant to the cytotoxic effects of this lectin when administered at a nonpermissive temperature. This treatment yields a population highly enriched in cells that demonstrate temperature-sensitive secretion. Therefore, this selection strategy has important application in isolating temperature-sensitive mutants for use in the study of the mammalian cell secretion pathway.
Henry
K W
KW
Department of Biology, University of Nevada, Reno 89557.
Brown
J L
JL
McCracken
A A
AA
D000818:Animals; D003208:Concanavalin A; D005821:Genetic Techniques; D008114:Liver Neoplasms, Experimental / Q000378:metabolism; D016296:Mutagenesis; D010641:Phenotype; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D013696:Temperature; D014407:Tumor Cells, Cultured
D011506:Proteins; D003208:Concanavalin A