Expression of SLAM (CDw150) on Sf9 cell surface using recombinant baculovirus mediates measles virus infection in the nonpermissive cells.
Signaling lymphocytic activation molecule (SLAM; also known as CDw150) has been reported as the receptor of measles virus (MV) interacting with MV hemagglutinin (MVH). In this study, we developed a baculovirus-derived vector, the Bacmid-egfp, containing a reporter gene encoding the enhanced green fluorescent protein (EGFP) under the control of the promoter of very late polyhedrin gene from Autographa californica multiple nucleopolyhedrovirus (AcMNPV), and employed the recombinant baculovirus to express SLAM in Sf9 (Spodoptera frugiperda) cells and investigate SLAM function. The result showed that the integration of the EGFP expression cassette in the Bac-to-Bac system facilitated research with the system without introducing compromises due to its use. SLAM protein fused to His-tag was expressed in Sf9 cells through the modified Bac-to-Bac system. The expressed SLAM was identified as approximately 46 kDa, and it presented on the cell surface, as revealed by fluorescent immunochemical staining and confocal microscopic analysis. The pull-down assay proved that SLAM protein expressed in this system could interact with MVH protein. After incubating with MV vaccine strain S191, cell fusion was only observed in the Sf9 cells expressing both EGFP and SLAM from recombinant baculovirus rather than those expressing EGFP only from the modified viral vector. Furthermore, MV replicated and induced apoptosis in the Sf9 cells with SLAM expression.
Liu
Xin
X
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072, China.
Zhou
Wenke
W
Zhang
Peng
P
Xu
Qin
Q
Hu
Chunlin
C
Chen
Xi
X
Yao
Lunguang
L
Li
Lingyun
L
Qi
Yipeng
Y
D000818:Animals; D015703:Antigens, CD; D017209:Apoptosis; D016367:Baculoviridae / Q000235:genetics; D002459:Cell Fusion; D002460:Cell Line; D017930:Genes, Reporter; D006023:Glycoproteins / Q000096:biosynthesis* / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D006389:Hemagglutinins, Viral / Q000378:metabolism; D007120:Immunochemistry; D007136:Immunoglobulins / Q000096:biosynthesis* / Q000235:genetics*; D008459:Measles virus / Q000502:physiology*; D008565:Membrane Proteins / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D018613:Microscopy, Confocal; D011485:Protein Binding; D011956:Receptors, Cell Surface; D011991:Receptors, Virus / Q000235:genetics* / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D000071177:Signaling Lymphocytic Activation Molecule Family Member 1; D018411:Spodoptera; D013194:Staining and Labeling; D014779:Virus Replication
D015703:Antigens, CD; D006023:Glycoproteins; D006389:Hemagglutinins, Viral; D007136:Immunoglobulins; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; D011991:Receptors, Virus; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; C032397:hemagglutinin protein G, measles virus; D049452:Green Fluorescent Proteins; D000071177:Signaling Lymphocytic Activation Molecule Family Member 1