Expression and trafficking of fluorescent viral membrane proteins in baculovirus-transduced BHK cells

Autor: Kirsi Ojala, Päivi J. Tikka, Pirjo Käpylä, Varpu Marjomäki, Christian Oker-Blom, Liisa Kautto
Rok vydání: 2004
Předmět:
Zdroj: Journal of Biotechnology. 114:165-175
ISSN: 0168-1656
DOI: 10.1016/j.jbiotec.2004.06.015
Popis: Baculovirus vectors show promise as a novel tool for gene delivery into mammalian cells and gene transfer with wild-type baculovirus has been demonstrated both in vitro and in vivo. To study expression and intracellular trafficking of foreign viral membrane proteins in baculovirus-transduced mammalian cells, the envelope proteins, E1 and E2, of rubella virus (RV) were chosen as a model. The enhanced green fluorescent protein (EGFP) and a red fluorescent protein (RFP) were fused to the C-terminus of E1 and E2, respectively. The proteins were cloned under a cytomegalovirus (CMV) promoter and expressed as fluorescent fusion proteins in baculovirus-transduced baby hamster kidney (BHK) cells. Expression of the chimeric proteins in these cells showed that E1 was retained within the ER and cis-Golgi when expressed alone. In contrast, E2 was efficiently transported to the trans-Golgi network (TGN). However, when expressed together, E1 co-localized with E2 in TGN and to some extent in the lysosomes. The recombinant baculovirus vectors were able to transduce the BHK cells efficiently and the fluorescent fusion constructs allowed easy detection of the trafficking events in the transduced mammalian cells. Consequently, this technique should have wide applications when intracellular analysis of protein synthesis and maturation is under study.
Databáze: OpenAIRE
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