Novel cell permeable motif derived from the PreS2-domain of hepatitis-B virus surface antigens

Abstract

Efficient transfer of proteins or nucleic acids across cellular membranes is a major problem in cell biology. Recently the existence of a fusogenic sequence was predicted in the junction area of the PreS2-and S-domain of the hepatitis-B virus surface antigens. We have identified cell permeability as a novel property of the PreS2-domain. Cell permeability of PreS2 is not restricted to hepatocytes. PreS2 translocates in an energy-independent manner into cells and is evenly distributed over the cytosol. Detailed analysis revealed that cell-permeability is mediated by an amphipatic ␣-helix between amino acids 41 and 52 of PreS2. Destruction of this translocation motif (PreS2-TLM) abolishes cell permeability. PreS2-TLM per se can act as a shuttle for peptides and functional proteins (such as EGFP). This permits the highly

A novel system for efficient gene transfer into primary human hepatocytes via cell‐permeable hepatitis B virus–like particle†

“…Previous work has shown that TLM mediates an energyand receptor-independent transport across the membrane. 5 In accordance with this, neither depletion of adenosine triphosphate nor inhibition of the endocytotic machinery affected TLM-mediated membrane translocation of nucleocapsids. Moreover, no vesicle formation or plasma membrane invagination was detected by electron microscopy, confirming that the uptake is not mediated by endocytosis.…”

“…5,13 Moreover, the cargo is transported directly into the cytoplasm: the endosomal compartment is not involved. 5 To investigate whether this holds true as well for the TLM-mediated transfer of nucleocapsids, the endocytotic machinery was blocked by ammonium chloride. Energy dependence was analyzed by adenosine triphosphate depetion using sodium azide.…”

“…5 The TLM mediates receptor-and energy-independent transfer of peptides, proteins, and nucleic acids across membranes, when fused to them, without affecting cellular integrity or interfering with intracellular signal transduction cascades. 5,13,23,24 We describe the potential of the TLM to mediate the transport of fully assembled particles into the cytoplasm of target cells in a receptor-and endocytosis-independent manner. Based on this system, we established cell-permeable VLPs harboring marker genes enabling an efficient gene transfer into primary hepatocytes.…”

“…[1][2][3][4][5][6][7][8][9] PTDs have been used to successfully treat preclinical models of human disease such as cancer, psoriasis, and stroke. [10][11][12][13] Transfer of nano-particular structures across cellular membranes is of increasing importance for the development of novel diagnostic and therapeutic tools.…”

Hypoxia-induced human endonuclease G expression suppresses tumor growth in a xenograft model

“…The N-terminus has a short 24 TLM from the herpes simplex virus, which is fused in-frame to a linker (L). The TLM is 14 amino acids long whereas the linker is 19 amino acids long with a high content of G, S and T. The linker is adapted from a linker-spacer sequence used in the construction of tdTomato.…”