Blocking translocation of cell surface molecules from the ER to the cell surface by intracellular antibodies targeted to the ER

Abstract

Intracellular antibodies (intrabodies) constitute a potent tool to neutralize the function of target proteins inside specific cell compartments (cytosol, nucleus, mitochondria and ER). The intrabody technology is an attractive alternative to the generation of gene-targeted knockout animals and complements or replaces knockdown techniques such as antisense-RNA, RNAi and RNA aptamers. This article focuses on intrabodies targeted to the ER. Intracellular anti-bodies expressed and retained inside the ER (ER intrabodies) are shown to be highly efficient in blocking the translocation of secreted and cell surface molecules from the ER to the cell surface.The advantage of ER intrabodies over cytoplasmic intrabodies is that they are correctly folded and easier to select. A particular advantage of the intrabody technology over existing ones is the possibility of inhibiting selectively post-translational modifications of proteins.The main applications of ER intrabodies so far have been (i) inactivation of oncogenic receptors and (ii) functional inhibition of virus envelope proteins and virus-receptor molecules on the surface of host cells.In cancer research, the number of in vivo mouse models for evaluation of the therapeutic potential of intrabodies is increasing.In the future, endosomal localized receptors involved in bacterial and viral infections, intracellular oncogenic receptors and enzymes involved in glycosylation of tumour antigens might be new targets for ER intrabodies.

S156C Mutation in Tissue Inhibitor of Metalloproteinases-3 Induces Increased Angiogenesis

“…These data indicate that mutant TIMP-3 has distinctive effects on the early steps of maturation of the receptor. It has been recently reported that novel highly efficient intrabodies can bind newly synthesized VEGFR-2 in the endoplasmic reticulum and block receptor transport to the cell surface (37,38). It is an interesting possibility that WT and mutant TIMP-3 may differentially affect immature receptor translocation via distinct interactions with newly synthesized VEGFR-2 in the endoplasmic reticulum.…”

Targeting antibodies to the cytoplasm

“…The method has been successfully applied to many targets, including human IL2 receptor, ErbB-2 receptor, b-amyloid precursor protein and VCAM1. 6,[17][18][19][20][21] Other subcellular compartments could be targeted by adding suitable signal sequences. 19,22 Functional studies of membrane receptors or secreted proteins ©2 0 1 1 L a n d e s B i o s c i e n c e .…”

“…In addition, their biochemical properties can be changed at will, e.g., by fusion to other protein domains, or they can even be immediately expressed inside the living cell to induce knockdown phenotypes. 6,7 The usefulness of antibodies in visualization or the knockdown of protein functions in the living cell has been proven by microinjection, [8][9][10][11][12] which demonstrated the stability and function of mature antibodies in the living cytoplasm. However, examples published since the first report of this method are scarce, mainly *Correspondence to: Stefan Dübel; Email: s.duebel@tu-bs.de Submitted: 10/13/10; Accepted: 11/04/10 DOI: 10.4161/mabs.3.…”

Molecular cloning and characterization of a novel anti-TLR9 intrabody

“…To bypass any lack of specificity, we used an intrabody with intrinsic specificity for its target [18,19]. Intrabodies can be targeted to the nucleus, mitochondria or endoplasmic reticulum (ER), where they bind their target proteins and thus persistently inhibit their function.…”

“…Intrabodies can be targeted to the nucleus, mitochondria or endoplasmic reticulum (ER), where they bind their target proteins and thus persistently inhibit their function. For instance, intrabodies targeted to the ER arrest transitory target proteins preventing their further subcellular trafficking [18,19]. Intrabodies are a promising alternative to RNA-based knockdown strategies in cases of failure of the latter [20][21][22].…”