Binding of recombinant but not endogenous prion protein to DNA causes DNA internalization and expression in mammalian cells.
Recombinant prion protein, rPrP, binds DNA. Both the KKRPK motif and the octapeptide repeat region of rPrP are essential for maximal binding. rPrP with pathogenic insertional mutations binds more DNA than wild-type rPrP. DNA promotes the aggregation of rPrP and protects its N terminus from proteinase K digestion. When rPrP is mixed with an expression plasmid and Ca(2+), the rPrP.DNA complex is taken up by mammalian cells leading to gene expression. In the presence of Ca(2+), rPrP by itself is also taken up by cells in a temperature- and pinocytosis-dependent manner. Cells do not take up rPrP(DeltaKKRPK), which lacks the KKRPK motif. Thus, rPrP is the carrier for DNA and the KKRPK motif is essential for its uptake. When mixed with DNA, a pentapeptide KKRPK, but not KKKKK, is sufficient for DNA internalization and expression. In contrast, whereas the normal cellular prion protein, PrP(C), on the cell surface can also internalize DNA, the imported DNA is not expressed. These findings may have relevance to the normal functions of PrP(C) and the pathogenic mechanisms of human prion disease.
Yin
Shaoman
S
Department of Pathology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44120.
Fan
Xingjun
X
Department of Pathology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44120.
Yu
Shuiliang
S
Department of Pathology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44120.
Li
Chaoyang
C
Department of Pathology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44120.
Sy
Man-Sun
MS
Department of Pathology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44120. Electronic address: mxs92@po.cwru.edu.
D020816:Amino Acid Motifs; D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA / Q000737:chemistry*; D019286:Endopeptidase K / Q000737:chemistry; D005786:Gene Expression Regulation; D006801:Humans; D007700:Kinetics; D010873:Pinocytosis; D011328:Prions / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry*
D011328:Prions; D011994:Recombinant Proteins; D004247:DNA; D019286:Endopeptidase K; D002118:Calcium