Human respiratory syncytial virus matrix protein is an RNA-binding protein: binding properties, location and identity of the RNA contact residues.
The human respiratory syncytial virus (HRSV) matrix (M) protein is a structural internal membrane protein. Here we have shown that, like its orthomyxovirus and rhabdovirus counterparts, it has RNA-binding capacity, as determined by retardation of (32)P-labelled riboprobes in gel electrophoresis, cross-linking with UV light and Northern-Western assays. Its binding to RNA was neither sequence-specific nor showed a length requirement, although it had cooperative kinetics with a K(d) of 25 nM and probably two different types of RNA-binding sites. After preparative cross-linking of (32)P-labelled riboprobes with purified, renatured HRSV Long strain M protein (256 residues), the residues in contact with RNA were located between amino acids 120 and 170, in the central part of the molecule. Lysine (positions 121, 130, 156 and 157) and arginine (position 170) residues located within this region and conserved among pneumovirus M proteins of different origins were found to be essential for RNA contact. M protein expression did not affect the replication and transcription of HRSV RNA analogues in vivo (except when expressed in large amounts), in contrast to the in vitro transcription inhibition described previously. In addition, M protein was found to aggregate into high-molecular-mass oligomers, both in the presence and absence of its RNA-binding activity. The formation of these structures has been related in other viruses to either viral or host-cell RNA metabolism.
Rodríguez
Lorena
L
Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra Majadahonda-Pozuelo Km 2, Majadahonda, Madrid 28220, Spain.
Cuesta
Isabel
I
Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra Majadahonda-Pozuelo Km 2, Majadahonda, Madrid 28220, Spain.
Asenjo
Ana
A
Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra Majadahonda-Pozuelo Km 2, Majadahonda, Madrid 28220, Spain.
Villanueva
Nieves
N
Centro Nacional de Microbiología, Instituto de Salud Carlos III, Ctra Majadahonda-Pozuelo Km 2, Majadahonda, Madrid 28220, Spain.
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D017124:Conserved Sequence; D006801:Humans; D008969:Molecular Sequence Data; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012136:Respiratory Syncytial Viruses / Q000302:isolation & purification / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D014162:Transfection; D014763:Viral Matrix Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D014763:Viral Matrix Proteins