The bunyavirus nucleocapsid protein is an RNA chaperone: Possible roles in viral RNA panhandle formation and genome replication

Abstract

Cellular RNA chaperones are crucial for the genesis of correctly folded functional RNAs. Using several complementary in vitro assays we find that the bunyavirus nucleocapsid protein (N) is an RNA chaperone. In the Bunyaviridae genomic RNA is in stable ''panhandle'' formation that arises through the hydrogen bonding of the terminal nucleotides of the RNA. The RNA chaperone function of N facilitates panhandle formation even though the termini are separated by >2 kb. RNA panhandle formation is likely driven by the exceptionally high base-pairing specificity of the terminal nucleotides as evidenced by P-num analysis. N protein can nonspecifically dissociate RNA duplexes. In addition, following panhandle formation, the RNA chaperone activity of N also appears to be involved in dissociation of the RNA panhandle and remains in association with the 5¢ terminus of the viral RNA following dissociation. Thus, N likely functions in the initiation of genome replication to allow efficient initiation of RNA synthesis by the viral polymerase. The RNA chaperone activity of N may be facilitated by an intrinsically disordered domain that catalyzes RNA unfolding driven by reciprocal entropy transfer. These observations highlight the essential features that are probably common to all RNA chaperones in which the role of the chaperone is to nonspecifically dissociate higher order structure and formation of functional higher order structure may often be predicted by RNA P-num value. The data also highlight features of N that are probably specifically important during replication of bunyavirus RNA.

The 3′ Untranslated Region of the Andes Hantavirus Small mRNA Functionally Replaces the Poly(A) Tail and Stimulates Cap-Dependent Translation Initiation from the Viral mRNA

“…Using this approach, we found that a 50-nt truncation in the ScRNA 3ЈNCR impeded efficient 3Ј-5Ј base pairing. This prediction is in accordance with previous reports for other hantaviruses; a 42-nt truncation in the SgRNA 3ЈNCR of the Sin Nombre virus, for instance, hinders panhandle formation (30). Although the degree of ANDV SmRNA truncation is unknown and we cannot ascertain from currently published information whether a 50-nt truncation truly corresponds to the SmRNA 3ЈUTR, we took the view that the 3ЈNCR featuring the 50-nt truncation (herein named 3ЈUTR) better represents the viral SmRNA 3ЈUTR than the full ScRNA 3ЈNCR.…”

Hantavirus Nucleocapsid Protein Has Distinct m7G Cap- and RNA-binding Sites

“…Our previous studies demonstrated that after specific recognition, N unwinds the panhandle and remains attached with the 5Ј terminus leaving the 3Ј terminus single stranded. This renders the 3Ј terminus accessible for both annealing of the primer and transcription initiation by the RdRp (15). In the current model (Fig.…”

“…The characteristic feature of the hantaviral genome is the partially complementary sequence at the 5Ј and 3Ј termini of each of the three genome segments that undergo base pairing and form panhandle structures (4 -6). N is a multifunctional protein playing a vital role in multiple processes of virus replication cycle and has been found to undergo trimerization both in vivo and in vitro (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19). During encapsidation, the three viral RNA (vRNA) molecules are specifically recognized by N inside the host cell and targeted for packaging.…”

Bunyavirus N: eIF4F surrogate and cap-guardian

“…N is a member of a class of proteins designated as, "RNA chaperones". 10 Since these peptides transiently dissociate higher order RNA structure, 11 the intrinsic chaperone activity of N is likely to substitute for the helicase activity of eIF4A.…”

“…(B) N dissociates the terminal viral RNA panhandle and remains associated with the 5' end of the viral RNA. 10 Dissociation facilitates access of the 3' end of the template for transcription initiation. The capped oligonucleotide acquired by N, is used for transcription initiation and elongation by the polymerase.…”