Cellular manipulation of human embryonic stem cells by TAT-PDX1 protein transduction.
Human embryonic stem cells (hESCs) are an in vitro model system for the study of human early development and a potential source for cell-based therapies. An efficient strategy for cellular manipulation of hESCs may be highly valuable for the analysis of gene function involved in human embryogenesis and the development of cell-based therapies via induced differentiation into particular cell types. However, plasmid transfection of hESCs has low efficiency and viral transduction may not be the method of choice for cell-based therapies due to genome integration. To overcome these limitations, we applied protein transduction technology that can transfer proteins into cells via direct penetration across the lipid bilayer. Here, we show that the FITC dye fused to the TAT protein transduction domain (PTD) was efficiently transferred into hESCs. In addition, the PDX1 transcription factor, which plays a central role in pancreatic development, was transferred into hESCs as a fusion form of TAT PTD. The transduced TAT-PDX1 activated its downstream target genes and induced insulin protein production in hESCs. These results demonstrate that protein transduction could be used in the cellular manipulation of hESCs and would provide a significant breakthrough for basic and therapeutic research in hESCs.
Kwon
Young Do
YD
Institute of Reproductive Medicine and Population, Medical Research Center, Korea.
Oh
Sun Kyung
SK
Kim
Hee Sun
HS
Ku
Seung-Yup
SY
Kim
Seok Hyun
SH
Choi
Young Min
YM
Moon
Shin Yong
SY
D000818:Animals; D002460:Cell Line; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D015696:Gene Products, tat / Q000235:genetics* / Q000378:metabolism; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology; D014161:Transduction, Genetic
D015696:Gene Products, tat; D018398:Homeodomain Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C084010:pancreatic and duodenal homeobox 1 protein; D016650:Fluorescein-5-isothiocyanate