Multifunctional Polyarginine Tag Antibody: protein transduction, iPS generation, and beyond


The Multifunctional Polyarginine Tag

iPS Generation and Beyond

Polyarginine Tag

Figure 1: Application of the polyarginine-tag in iPS technology

Polyarginine Western Blot

Figure 2: Left: Western Blot detection of 11R-tagged reprogramming factors, including human c-Myc, KLF4, Oct4, and Sox2 proteins, using Polyarginine (9R) Antibody (Cat# CY1114). Middle & Right: This antibody also stains Human Dermal Fibroblasts (HDF)^ (Cat# 106-05a) transduced with the 11R-tagged reprogramming factors, human c-Myc, KLF4, Oct4, and Sox2 proteins (Right), but not untreated HDF (Middle), in immunofluorescent analysis. (^This cell was also used by Dr. Yamanaka's lab to generate the world's first iPS cells. See: Takahashi, K. et al: Cell 131:861-72, 2007.)

The polyarginine tag, typically consisting of nine to eleven arginine residues, is a widely used cell-penetrating peptide (CPP), that improves cellular uptake of linked cargo such as proteins, nucleic acids (oligonucleotides, peptide-nucleic acids, siRNAs), nanoparticles and liposomes in a wide range of cells.1 The polyarginine tag has been shown to be 20-fold more efficient in cellular uptake than classic CPPs,2 and can be a powerful tool for cell biology and clinical translational studies. For example, polyarginine-fused p53 protein (p53-11R) can be delivered into cancer cells and reduce proliferation.3

Polyarginine protein transduction has also been used to generate induced pluripotent stem (iPS) cells. Adult Human Dermal Fibroblasts (HDF) (Cat.# 106-05a, Cell Applications, Inc.) were first reprogrammed by Dr. Yamanaka and colleagues using the four transcription factor genes: c-Myc, KLF4, Oct4, and Sox2, under ES cell culture conditions.4 However, clinical application of iPS cells faces many obstacles, including the risk of tumor formation when introducing inducing genes. In order to reduce such risk, recombinant fusion proteins of the four reprogramming factors (c-Myc, KLF4, Oct4, and Sox2) tagged with polyarginine were created by Dr. Ding and colleagues.5 These polyarginine-tagged proteins entered Mouse Embryonic Fibroblasts (MEF) and reprogrammed them into iPS cells. Therefore, polyarginine protein transduction provides a novel method of producing iPS cells and paves the way for powerful cell-based therapies, as seen in Figure 1.

Given the ability of CPPs such as polyarginine and HIV-1 Tat tag sequence to promote cellular uptake, it is important to monitor the amount and lifespan of CPP-tagged cargo in cells as a reference for the success of uptake. To address this need, Cell Applications, Inc. has developed a polyarginine-specific rabbit polyclonal antibody (Cat# CY1114) using proprietary *NatureTope™ technology. This antibody recognizes both 9R and 11R polyarginine, and specifically detects polyarginine-tagged proteins in a variety of assays, including Western Blotting, immunofluorescent staining, and immunoprecipitation. As shown in Figure 2, this antibody detects the 11R-tagged reprogramming transcriptional factors c-Myc, KLF4, Oct4, and Sox2. Thus, polyarginine antibody facilitates and extends the applications of polyarginine-tags.

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1. Tunnemann, G. et al: J. Peptide Sci. 14:469–76, 2008.
2. Wender, P.A. et al: Proc. Natl. Acad. Sci. USA 97:13003-8, 2000.
3. Takenobu, T. et al: Mol. Cancer Ther. 1:1043-9, 2002.

4. Takahashi, K. et al: Cell 131:861-72, 2007
5. Zhou, H. et al: Cell Stem Cell 4:381-4, 2009