Reprogramming human fibroblasts using HIV-1 TAT recombinant proteins OCT4, SOX2, KLF4 and c-MYC.
It has been shown that human and murine fibroblasts can be reprogrammed by ectopic expression of transcription factors using viral vectors. For the purpose of human therapeutic applications, the integration of viral transgenes into the genome is unlikely to be accepted. We therefore produced recombinant transcription factor proteins in E. coli (OCT4, SOX2, c-MYC and KLF4) carrying the cell penetrating TAT domain from HIV1. The purified proteins were able to enter into mammalian cells when added to tissue culture medium but appeared not to translocate to the nucleus. Further investigation indicated that most of the protein was tied up in the endosomes and was unavailable for reprogramming. Once this problem has been solved it seems likely that protein reprogramming will be the method of choice for clinical applications.
Pan
Chuanying
C
College of Animal Science and Technology, Shaanxi Key Laboratory of Molecular Biology for Agriculture, Northwest A&F University, No. 22 Xinong Road, Yangling, 712100, Shaanxi, China. cpan@wfubmc.edu
Lu
Baisong
B
Chen
Hong
H
Bishop
Colin E
CE
D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D065150:Cellular Reprogramming / Q000235:genetics*; D005347:Fibroblasts / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D015497:HIV-1 / Q000378:metabolism*; D006801:Humans; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism; D050814:Octamer Transcription Factor-3 / Q000378:metabolism; D011489:Protein Denaturation; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism*; D012394:Rosaniline Dyes; D055748:SOXB1 Transcription Factors / Q000378:metabolism; D013194:Staining and Labeling; D013997:Time Factors; D014157:Transcription Factors / Q000378:metabolism*; D014161:Transduction, Genetic; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000378:metabolism*
C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D050814:Octamer Transcription Factor-3; D016271:Proto-Oncogene Proteins c-myc; D011993:Recombinant Fusion Proteins; D012394:Rosaniline Dyes; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; D014157:Transcription Factors; D054322:tat Gene Products, Human Immunodeficiency Virus; C048139:Coomassie blue