Expression of Mammalian Membrane Proteins in Mammalian Cells Using Semliki Forest Virus Vectors

2009 ◽  
pp. 149-163 ◽  
Author(s):  
Kenneth Lundstrom
Keyword(s):  
Membrane Proteins ◽  
Mammalian Cells ◽  
Semliki Forest Virus ◽  
Virus Vectors

TMCC3 localizes at the three-way junctions for the proper tubular network of the endoplasmic reticulum

2019 ◽  
Vol 476 (21) ◽  
pp. 3241-3260
Author(s):  
Sindhu Wisesa ◽  
Yasunori Yamamoto ◽  
Toshiaki Sakisaka
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Mammalian Cells ◽  
Coiled Coil ◽  
Transmembrane Domains ◽  
Domain Family ◽  
Coiled Coil Domain ◽  
U2os Cells ◽  
Er Membrane

The tubular network of the endoplasmic reticulum (ER) is formed by connecting ER tubules through three-way junctions. Two classes of the conserved ER membrane proteins, atlastins and lunapark, have been shown to reside at the three-way junctions so far and be involved in the generation and stabilization of the three-way junctions. In this study, we report TMCC3 (transmembrane and coiled-coil domain family 3), a member of the TEX28 family, as another ER membrane protein that resides at the three-way junctions in mammalian cells. When the TEX28 family members were transfected into U2OS cells, TMCC3 specifically localized at the three-way junctions in the peripheral ER. TMCC3 bound to atlastins through the C-terminal transmembrane domains. A TMCC3 mutant lacking the N-terminal coiled-coil domain abolished localization to the three-way junctions, suggesting that TMCC3 localized independently of binding to atlastins. TMCC3 knockdown caused a decrease in the number of three-way junctions and expansion of ER sheets, leading to a reduction of the tubular ER network in U2OS cells. The TMCC3 knockdown phenotype was partially rescued by the overexpression of atlastin-2, suggesting that TMCC3 knockdown would decrease the activity of atlastins. These results indicate that TMCC3 localizes at the three-way junctions for the proper tubular ER network.

scholarly journals 472. Biodistribution of Semliki Forest Virus Vectors and Effect of Preimmunizations in Mice

2006 ◽  
Vol 13 ◽  
pp. S183
Author(s):  
Juan R. Rodriguez ◽  
Erkuden Casales ◽  
Jesus Prieto ◽  
Cristian Smerdou
Keyword(s):  
Semliki Forest Virus ◽  
Virus Vectors

scholarly journals Semliki Forest virus vectors: efficient vehicles for in vitro and in vivo gene delivery

FEBS Letters ◽  
2001 ◽  
Vol 504 (3) ◽  
pp. 99-103 ◽  
Author(s):  
Kenneth Lundstrom ◽  
Christophe Schweitzer ◽  
Daniel Rotmann ◽  
Danielle Hermann ◽  
Edith M. Schneider ◽  
...  
Keyword(s):  
Gene Delivery ◽  
Semliki Forest Virus ◽  
Virus Vectors ◽  
In Vivo Gene Delivery

A modification of the split-tobacco etch virus method for monitoring interactions between membrane proteins in mammalian cells

2012 ◽  
Vol 423 (1) ◽  
pp. 109-118 ◽  
Author(s):  
Xavier Capdevila-Nortes ◽  
Tania López-Hernández ◽  
Francisco Ciruela ◽  
Raúl Estévez
Keyword(s):  
Membrane Proteins ◽  
Mammalian Cells ◽  
Tobacco Etch Virus

Individual and bivalent vaccines against botulinum neurotoxin serotypes A and B using DNA-based Semliki Forest virus vectors

Vaccine ◽  
2009 ◽  
Vol 27 (44) ◽  
pp. 6148-6153 ◽  
Author(s):  
Yunzhou Yu ◽  
Jiyu Yu ◽  
Na Li ◽  
Shuang Wang ◽  
Weiyuan Yu ◽  
...  
Keyword(s):  
Botulinum Neurotoxin ◽  
Semliki Forest Virus ◽  
Virus Vectors

scholarly journals Proteolytic processing of Semliki Forest virus-specific non-structural polyprotein by nsP2 protease

2001 ◽  
Vol 82 (4) ◽  
pp. 765-773 ◽  
Author(s):  
Andres Merits ◽  
Lidia Vasiljeva ◽  
Tero Ahola ◽  
Leevi Kääriäinen ◽  
Petri Auvinen
Keyword(s):  
Mammalian Cells ◽  
Active Sites ◽  
Insect Cells ◽  
Semliki Forest Virus ◽  
Point Mutations ◽  
Proteolytic Processing ◽  
Wild Type ◽  
In Trans ◽  
Polyprotein Precursor

The RNA replicase proteins of Semliki Forest virus (SFV) are translated as a P1234 polyprotein precursor that contains two putative autoproteases. Point mutations introduced into the predicted active sites of both proteases nsP2 (P2) and nsP4 (P4), separately or in combination, completely abolished virus replication in mammalian cells. The effects of these mutations on polyprotein processing were studied by in vitro translation and by expression of wild-type polyproteins P1234, P123, P23, P34 and their mutated counterparts in insect cells using recombinant baculoviruses. A mutation in the catalytic site of the P2 protease, C478A, (P2CA) completely abolished the processing of P12CA34, P12CA3 and P2CA3. Co-expression of P23 and P12CA34 in insect cells resulted in in trans cleavages at the P2/3 and P3/4 sites. Co-expression of P23 and P34 resulted in cleavage at the P3/4 site. In contrast, a construct with a mutation in the active site of the putative P4 protease, D6A, (P1234DA) was processed like the wild-type protein. P34 or its truncated forms were not processed when expressed alone. In insect cells, P4 was rapidly destroyed unless an inhibitor of proteosomal degradation was used. It is concluded that P2 is the only protease needed for the processing of SFV polyprotein P1234. Analysis of the cleavage products revealed that P23 or P2 could not cleave the P1/2 site in trans.

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