Importance of RNA length for in vitro encapsidation by the nucleoprotein of human respiratory syncytial virus

被引:7
|
作者
Gonnin, Lorene [1 ]
Richard, Charles-Adrien [1 ]
Gutsche, Irina [2 ]
Chevret, Didier [1 ]
Troussier, Joris [3 ]
Vasseur, Jean-Jacques
Debart, Francoise [3 ]
Eleoue, Jean-Francois [1 ]
Galloux, Marie [1 ]
机构
[1] Univ Paris Saclay, INRAE, UVSQ, VIM, Jouy En Josas, France
[2] Univ Grenoble Alpes, CEA, CNRS, IBS, Grenoble, France
[3] Univ Montpellier, CNRS, UMR, IBMM,ENSCM, Montpellier, France
关键词
CRYSTAL-STRUCTURE; BINDING-SITE; COMPLEX; PHOSPHOPROTEIN; TRANSCRIPTION; INFECTIONS; TAXONOMY; SEQUENCE; PROTEIN;
D O I
10.1016/j.jbc.2022.102337
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Respiratory syncytial virus has a negative-sense single -stranded RNA genome constitutively encapsidated by the viral nucleoprotein N, forming a helical nucleocapsid which is the template for viral transcription and replication by the viral polymerase L. Recruitment of L onto the nucleocapsid depends on the viral phosphoprotein P, which is an essential L cofactor. A prerequisite for genome and antigenome encapsidation is the presence of the monomeric, RNA-free, neosynthesized N pro-tein, named N0. Stabilization of N0 depends on the binding of the N-terminal residues of P to its surface, which prevents N oligomerization. However, the mechanism involved in the transition from N0-P to nucleocapsid assembly, and thus in the specificity of viral genome encapsidation, is still unknown. Furthermore, the specific role of N oligomerization and RNA in the morphogenesis of viral factories, where viral transcription and replication occur, have not been elucidated although the interaction between P and N complexed to RNA has been shown to be responsible for this process. Here, using a chimeric protein comprising N and the first 40 N-terminal residues of P, we succeeded in purifying a recombinant N0-like protein competent for RNA encapsidation in vitro. Our results showed the importance of RNA length for stable encapsidation and revealed that the nature of the 50 end of RNA does not explain the specificity of encapsidation. Finally, we showed that RNA encapsidation is crucial for the in vitro reconstitution of pseudo-viral factories. Together, our findings provide insight into respiratory syncytial virus viral genome encapsidation specificity.
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页数:14
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