title: Characterization of a novel cell penetrating peptide derived from human Oct4.
abstract: BACKGROUND
  Oct4 is a transcription factor that plays a major role for the preservation of the pluripotent state in embryonic stem cells as well as for efficient reprogramming of somatic cells to induced pluripotent stem cells (iPSC) or other progenitors. Protein-based reprogramming methods mainly rely on the addition of a fused cell penetrating peptide. This study describes that Oct4 inherently carries a protein transduction domain, which can translocate into human and mouse cells.
  
  
  RESULTS
  A 16 amino acid peptide representing the third helix of the human Oct4 homeodomain, referred to as Oct4 protein transduction domain (Oct4-PTD), can internalize in mammalian cells upon conjugation to a fluorescence moiety thereby acting as a cell penetrating peptide (CPP). The cellular distribution of Oct4-PTD shows diffuse cytosolic and nuclear staining, whereas penetratin is strictly localized to a punctuate pattern in the cytoplasm. By using a Cre/loxP-based reporter system, we show that this peptide also drives translocation of a functionally active Oct4-PTD-Cre-fusion protein. We further provide evidence for translocation of full length Oct4 into human and mouse cell lines without the addition of any kind of cationic fusion tag. Finally, physico-chemical properties of the novel CPP are characterized, showing that in contrast to penetratin a helical structure of Oct4-PTD is only observed if the FITC label is present on the N-terminus of the peptide.
  
  
  CONCLUSIONS
  Oct4 is a key transcription factor in stem cell research and cellular reprogramming. Since it has been shown that recombinant Oct4 fused to a cationic fusion tag can drive generation of iPSCs, our finding might contribute to further development of protein-based methods to generate iPSCs. Moreover, our data support the idea that transcription factors might be part of an alternative paracrine signalling pathway, where the proteins are transferred to neighbouring cells thereby actively changing the behaviour of the recipient cell.
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.
authors.forename: Eva
authors.initials: E
authors.lastname: Harreither
authors.affiliation: Department of Chemical and Biological Engineering/Physical Chemistry, Chalmers University of Technology, Kemivägen 10, S-412 96 Gothenburg, Sweden.
authors.forename: Hanna A
authors.initials: HA
authors.lastname: Rydberg
authors.affiliation: Department of Chemical and Biological Engineering/Physical Chemistry, Chalmers University of Technology, Kemivägen 10, S-412 96 Gothenburg, Sweden.
authors.forename: Helene L
authors.initials: HL
authors.lastname: Amand
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.
authors.forename: Vaibhav
authors.initials: V
authors.lastname: Jadhav
authors.affiliation: ACIB GmbH, Austrian Center of Industrial Biotechnology, Petersgasse 14, 8010 Graz, Austria.
authors.forename: Lukas
authors.initials: L
authors.lastname: Fliedl
authors.affiliation: Key Laboratory of Regenerative Biology, Chinese Academy of Sciences, and Guangdong Provincial Key Laboratory of Stem Cells and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, 510530 Guangzhou, China.
authors.forename: Christina
authors.initials: C
authors.lastname: Benda
authors.affiliation: Key Laboratory of Regenerative Biology, Chinese Academy of Sciences, and Guangdong Provincial Key Laboratory of Stem Cells and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, 510530 Guangzhou, China.
authors.forename: Miguel A
authors.initials: MA
authors.lastname: Esteban
authors.affiliation: Key Laboratory of Regenerative Biology, Chinese Academy of Sciences, and Guangdong Provincial Key Laboratory of Stem Cells and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, 510530 Guangzhou, China.
authors.forename: Duanqing
authors.initials: D
authors.lastname: Pei
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria ; ACIB GmbH, Austrian Center of Industrial Biotechnology, Petersgasse 14, 8010 Graz, Austria.
authors.forename: Nicole
authors.initials: N
authors.lastname: Borth
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria ; Evercyte GmbH, Muthgasse 18, 1190 Vienna, Austria ; ACIB GmbH, Austrian Center of Industrial Biotechnology, Petersgasse 14, 8010 Graz, Austria.
authors.forename: Regina
authors.initials: R
authors.lastname: Grillari-Voglauer
authors.affiliation: Stem Cell Engineering Group, Institute of Reconstructive Neurobiology, University of Bonn - Life & Brain Center and Hertie Foundation, Sigmund-Freud Strasse 25, D-53105 Bonn, Germany.
authors.forename: Oliver
authors.initials: O
authors.lastname: Hommerding
authors.affiliation: Stem Cell Engineering Group, Institute of Reconstructive Neurobiology, University of Bonn - Life & Brain Center and Hertie Foundation, Sigmund-Freud Strasse 25, D-53105 Bonn, Germany ; Stem Cell and Regenerative Medicine Group, Institute of Anatomy and Cell Biology, Julius-Maximilians-University Würzburg, Koellikerstrasse 6, D-97070 Würzburg, Germany.
authors.forename: Frank
authors.initials: F
authors.lastname: Edenhofer
authors.affiliation: Department of Chemical and Biological Engineering/Physical Chemistry, Chalmers University of Technology, Kemivägen 10, S-412 96 Gothenburg, Sweden.
authors.forename: Bengt
authors.initials: B
authors.lastname: Nordén
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria ; Evercyte GmbH, Muthgasse 18, 1190 Vienna, Austria ; Christian Doppler Laboratory on Biotechnology of Skin Aging, University of Natural Resources and Life Sciences, Muthgasse 18, 1190 Vienna, Austria.
authors.forename: Johannes
authors.initials: J
authors.lastname: Grillari
keywords: Cell penetrating peptides; Cellular internalization; Homeodomain transcription factors; Oct4; Penetratin; Reprogramming