title: Evaluating the translocation properties of a new nuclear targeted penetrating peptide using two fluorescent markers.
abstract: Human-derived cell penetrating peptides (CPPs) have attracted much more attentions than other CPPs which are limited by their potential toxicity and immunogenicity. Previously, we identified a novel human-originated CPP (named heparin-binding domain (HBD) in this article), which derived from the C-terminus of human extracellular superoxide dismutase, and demonstrated HBD is an efficient vector for delivering exogenous drug molecules such as apoptin into HeLa cells. In this study, we found this novel CPP showed differentiated efficiency in several tested cell lines. Heparin competitive inhibition experiment and heparanase pre-incubation experiment showed cell surface polysaccharides play an important role for the transmembrane transport. The results of endocytosis inhibitors suggested that HBD penetrates the cell membrane via a direct translocation, which is different from that of TAT, a classical clathrin-mediated endocytosis. HBD could deliver up to 90 kD protein cargoes into cells. Different conjugated modes with cargo molecules greatly affect their translocation efficiency. HBD also showed significant nuclear transport capacity when it was incubated with HeLa cells. Furthermore, the core region for HBD possessing membrane-penetrating ability was identified by deletion analyses. These results would be helpful for developing HBD as a new nuclear delivery tool for therapeutic biomolecules.
authors.affiliation: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology , Shanghai , China .
authors.forename: Rui
authors.initials: R
authors.lastname: Zhang
authors.forename: Xu-Zhong
authors.initials: XZ
authors.lastname: Yang
authors.forename: Jia-Wei
authors.initials: JW
authors.lastname: Wang
authors.forename: Long-Yun
authors.initials: LY
authors.lastname: Fu
authors.forename: Jian
authors.initials: J
authors.lastname: Zhao
authors.forename: Fu-Jun
authors.initials: FJ
authors.lastname: Wang
mesh_terms: D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D057846:Cell-Penetrating Peptides / Q000378:metabolism*; D004337:Drug Carriers / Q000378:metabolism*; D016503:Drug Delivery Systems; D005456:Fluorescent Dyes / Q000378:metabolism; D005966:Glucuronidase / Q000378:metabolism; D006367:HeLa Cells; D006493:Heparin / Q000378:metabolism; D006801:Humans; D011134:Polysaccharides / Q000378:metabolism; D000099175:Heparanase
chemical_list: D057846:Cell-Penetrating Peptides; D004337:Drug Carriers; D005456:Fluorescent Dyes; D005966:Glucuronidase; D006493:Heparin; D011134:Polysaccharides; D000099175:Heparanase
keywords: Characteristic; HBD; human-derived CPP; nuclear targeted delivery