title: Efficient, long-term transgene expression in Xenopus laevis dermal melanophores.
abstract: Xenopus laevis dermal melanophores provide an excellent model system for the investigation of complex cellular processes. Specifically, the expression of exogenous genes in Xenopus melanophores is the basis of recombinant bioassays for the study of receptor-ligand interactions. However, due to their slow rate of cell division and to the relatively low efficiency of current transfection protocols, long-term expression of exogenous genes and the generation of stable melanophore cell lines remains problematic. In this report we demonstrate the efficient, long-term expression of two exogenous proteins, the enhanced green fluorescent protein (EGFP) and the human CD4 (hCD4) cell surface receptor, following stable introduction into Xenopus melanophores via an HIV-1 based vector. Transduction of melanophores with the EGFP expression vector resulted in up to 80% EGFP+ cells. After 1 year in continuous culture in the absence of antibiotic selection, more than 60% of the cells remained EGFP+. Furthermore, we demonstrate the expression of hCD4 melanophores for over 9 months in continuous culture in the absence of antibiotic selection. Our results indicate that lentivirus vectors provide an efficient means of introducing genetic information into Xenopus melanophores, resulting in sustained levels of gene expression. The significance of this gene transfer system for the study of cellular signal transduction pathways is discussed.
authors.affiliation: Department of Internal Medicine, University of Texas Southwestern Medical Center at Dallas, 75390-9113, USA.
authors.forename: J
authors.initials: J
authors.lastname: Gatlin
authors.forename: D J
authors.initials: DJ
authors.lastname: Unett
authors.forename: M R
authors.initials: MR
authors.lastname: Lerner
authors.forename: J V
authors.initials: JV
authors.lastname: Garcia
mesh_terms: D000818:Animals; D015704:CD4 Antigens / Q000235:genetics; D002478:Cells, Cultured; D020405:Dermis / Q000166:cytology*; D005434:Flow Cytometry; D015870:Gene Expression / Q000502:physiology; D005822:Genetic Vectors; D015497:HIV-1 / Q000235:genetics; D016086:Lentivirus / Q000235:genetics; D008547:Melanophores / Q000502:physiology*; D008565:Membrane Proteins / Q000235:genetics; D010957:Plasmids; D014161:Transduction, Genetic / Q000379:methods*; D019076:Transgenes / Q000235:genetics*; D014982:Xenopus laevis
chemical_list: D015704:CD4 Antigens; D008565:Membrane Proteins