Positively charged DNA-binding proteins cause apparent cell membrane translocation.
Several positively charged DNA-binding proteins such as the human immunodeficiency virus Tat protein, the Antennapedia (Antp) homeobox protein, and the herpes simplex virus VP22 protein have been reported to translocate across cell membranes and accumulate in cell nuclei. The import occurs by a poorly understood mechanism that appears to be receptor- and energy-independent. We showed that both VP22 and the positively charged histone H1 adhered to the cell membrane of living cells and were not removed by extensive washing. However, after fixation the proteins relocated to the cell nucleus. The nuclear accumulation of VP22 and histone H1 after fixation shows that positively charged proteins may appear to translocate across the cell membrane because of a fixation artifact. The majority of studies on "membrane permeable" proteins and peptides have been performed using fixation techniques, and our study shows that influx of these proteins may occur during fixation rather than in living cells.
Lundberg
Mathias
M
Medical Nobel Institute for Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden.
Johansson
Magnus
M
D021581:Active Transport, Cell Nucleus; D000818:Animals; D016477:Artifacts; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005404:Fixatives / Q000737:chemistry; D049452:Green Fluorescent Proteins; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics; D000432:Methanol / Q000737:chemistry; D008856:Microscopy, Fluorescence; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D016707:Tissue Fixation; D015678:Viral Structural Proteins / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D005404:Fixatives; D006657:Histones; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D015678:Viral Structural Proteins; C073413:herpes simplex virus type 1 protein VP22; D049452:Green Fluorescent Proteins; D000432:Methanol