Transfer of functional adenovirus E1A transcription activator proteins into mammalian cells by protoplast fusion.
Human adenovirus 2/5 E1A proteins were used to evaluate protoplast fusion as a method of transferring functional proteins into mammalian cells. Both the E1A 13 and 12 S mRNA products expressed in Escherichia coli are shown to activate in trans adenovirus gene expression following transfer into monkey kidney cells by protoplast fusion. Approximately 20% of the recipient mammalian cells exhibited positive nuclear E1A-specific immunofluorescence following fusion with protoplasts containing E1A protein. E. coli-expressed E1A protein was modified post-translationally in Vero cells following protoplast fusion, as evidenced by its shift in sodium dodecyl sulfate-polyacrylamide gel electrophoresis mobility. These results establish protoplast fusion as a simple rapid method for examining the functional activity, intracellular distribution, and post-translational modification of E. coli-expressed proteins in intact mammalian cells.
D017456:Adenovirus Early Proteins; D000260:Adenoviruses, Human / Q000235:genetics* / Q000254:growth & development; D000818:Animals; D002472:Cell Transformation, Viral; D004926:Escherichia coli / Q000235:genetics; D005796:Genes; D005814:Genes, Viral; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D010957:Plasmids; D011523:Protoplasts / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic; D014709:Vero Cells; D014775:Virus Activation
D017456:Adenovirus Early Proteins; D009856:Oncogene Proteins, Viral; D012333:RNA, Messenger