Simultaneous imaging and restoration of cell function using cell permeable peptide probe.
Targeting tissues/cells using probing materials to detect diseases such as cancer and inflammatory disease has been attempted with some success. Most of the molecular targets used in diagnosis and therapy were identified through the discovery of intracellular signaling pathways. Among intracellular signaling processes, the ubiquitination of proteins, and thereby their proteasomal degradation, is important because it plays a role in most diseases involving alterations to a component of the ubiquitination system, particularly E3 ligases, which have selective target-binding affinity and are key to the success of regulating the disorder. The regulation and monitoring of E3 ligases can be achieved using peptides containing protein-protein binding motifs. We generated a human protein-derived peptide that could target Smurf1, a member of the E3 ligase family, by competitively binding to osteo-Smads. To effectively deliver it into cells, the peptide was further modified with a cell-penetrating peptide. The peptide contains two fluorescent dyes: fluorescein isothiocyanate (FITC; absorbance/emission wavelengths: 495/519 nm) as a fluorophore and black hole quencher-1 (BHQ-1) as a fluorescence quencher. When the target Smurf1 combined with complementary sequences in the peptide probe, the distance between the fluorophore and BHQ-1 increased via a conformational change, resulting in the recovery of the fluorescence signal. Simultaneously, the degradation of Smad1/5/8 was blocked by the binding of the peptide probe to Smurf1, leading to the potentiation of the osteogenic pathway, which was reflected by an increase in the expression of osteoinductive genes, such as alkaline phosphatase and osteocalcin. Possible future applications of the peptide probe include its integration into imaging tools for the diagnosis of Smurf1-overexpressing diseases.
Dental Regenerative Biotechnology Major, Dental Research Institute, School of Dentistry, Seoul National University, Seoul, South Korea.
Jin Sook
JS
Suh
Central Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul, South Korea.
Jue Yeon
JY
Lee
Dental Genetics Major, Dental Research Institute, School of Dentistry, Seoul National University, Seoul, South Korea.
Gene
G
Lee
Central Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul, South Korea.
Chong Pyoung
CP
Chung
Dental Regenerative Biotechnology Major, Dental Research Institute, School of Dentistry, Seoul National University, Seoul, South Korea; Central Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul, South Korea. Electronic address: parkyj@snu.ac.kr.
Yoon Jeong
YJ
Park
D000595:Amino Acid Sequence; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002463:Cell Membrane Permeability / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D057846:Cell-Penetrating Peptides / Q000737:chemistry* / Q000494:pharmacology; D004195:Disease Models, Animal; D004705:Endocytosis / Q000187:drug effects; D005260:Female; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008954:Models, Biological; D057054:Molecular Imaging; D015335:Molecular Probes / Q000494:pharmacology*; D008969:Molecular Sequence Data; D010012:Osteogenesis / Q000187:drug effects; D011917:Rats, Inbred Lew; D012213:Rheumatic Fever / Q000378:metabolism / Q000473:pathology; D051785:Smad Proteins / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D054875:Ubiquitination / Q000187:drug effects
D057846:Cell-Penetrating Peptides; D005456:Fluorescent Dyes; D015335:Molecular Probes; D051785:Smad Proteins; C498342:SMURF1 protein, human; D044767:Ubiquitin-Protein Ligases
Cell penetrating peptide probe; Osteogenesis; Stem cell monitoring; Theranosis; Ubiquitin-proteasomal system