Minute Virus of Canines NP1 Protein Governs the Expression of a Subset of Essential Nonstructural Proteins via Its Role in RNA Processing

Abstract

Parvoviruses use a variety of means to control the expression of their compact genomes. The bocaparvovirus minute virus of canines (MVC) encodes a small, genus-specific protein, NP1, which governs access to the viral capsid gene via its role in alternative polyadenylation and alternative splicing of the single MVC premRNA. In addition to NP1, MVC encodes five additional nonstructural proteins (NS) that share an initiation codon at the left end of the genome and which are individually encoded by alternative multiply spliced mRNAs. We found that three of these proteins were encoded by mRNAs that excise the NP1-regulated MVC intron immediately upstream of the internal polyadenylation site, (pA)p, and that generation of these proteins was thus regulated by NP1. Splicing of their progenitor mRNAs joined the amino termini of these proteins to the NP1 open reading frame, and splice site mutations that prevented their expression inhibited virus replication in a host celldependent manner. Thus, in addition to controlling capsid gene access, NP1 also controls the expression of three of the five identified NS proteins via its role in governing MVC pre-mRNA splicing. IMPORTANCEThe Parvovirinae are small nonenveloped icosahedral viruses that are important pathogens in many animal species, including humans. Minute virus of canine (MVC) is an autonomous parvovirus in the genus Bocaparvovirus. It has a single promoter that generates a single pre-mRNA. NP1, a small genus-specific MVC protein, participates in the processing of this pre-mRNA and so controls capsid gene access via its role in alternative internal polyadenylation and splicing. We show that NP1 also controls the expression of three of the five identified NS proteins via its role in governing MVC pre-mRNA splicing. These NS proteins together are required for virus replication in a host cell-dependent manner.KEYWORDS gene expression, parvovirus P arvoviruses use multiple mechanisms to maximize the coding potential of their compact genomes, including alternative transcription initiation, alternative splicing, alternative polyadenylation, and alternative translation initiation mechanisms (1-3).Infection with minute virus of canines (MVC), a member of the genus Bocaparvovirus (4), can cause abortion and stillbirth in pregnant dogs, as well as mild gastroenteritis and respiratory disease in puppies (5-8). MVC generates a single pre-mRNA from a promoter at the left-hand end of the genome (P6) that is processed via alternative splicing and alternative polyadenylation into multiple nonstructural and capsidencoding transcripts (9, 10). As with other parvoviruses, an open reading frame (ORF) in the left half of the genome encodes nonstructural (NS) proteins, while an ORF in the right half encodes the capsid proteins VP1 and VP2 (2). The bocaparvoviruses also

The Human Bocavirus 1 NP1 Protein Is a Multifunctional Regulator of Viral RNA Processing

“…It has recently been reported that the C-terminal regions of three of the MVC NS proteins are generated from mRNAs spliced at the third intron (17). Therefore, their expression is facilitated by MVC NP1 (17).…”

“…It has recently been reported that the C-terminal regions of three of the MVC NS proteins are generated from mRNAs spliced at the third intron (17). Therefore, their expression is facilitated by MVC NP1 (17). For HBoV1, the C terminus of the viral NS proteins is determined by splicing of the second intron of their encoding mRNAs, which is independent of NP1 (20).…”

“…HBoV1 can cause mild to severe respiratory tract infections, primarily in children (3)(4)(5)(6). Until its recent cloning, and the development of a useful tissue culture system to grow HBoV1 (7)(8)(9)(10)(11), MVC was often used as a surrogate for the characterization of aspects of bocavirus gene expression and virus-host interactions (12)(13)(14)(15)(16)(17).…”

“…Both of these processes are necessary to gain proper access to the capsid gene ORF (13,14,19). Additionally, the C-terminal region of three of the MVC NS proteins are generated from mRNAs spliced at the third intron; thus, their expression is also facilitated by MVC NP1 (17). RNAs which encode the HBoV1 NS proteins are not spliced at the analogous intron (20).…”

Minute Virus of Canines NP1 Protein Interacts with the Cellular Factor CPSF6 To Regulate Viral Alternative RNA Processing

“…NP1 is required for efficient read-through of the internal polyadenylation site (pA)p and splicing of the adjacent upstream intron. Additionally, three essential nonstructural proteins are encoded by mRNAs that excise the NP1-regulated MVC intron immediately upstream of the internal polyadenylation site (pA)p, and so generation of these proteins is also regulated at the RNA processing level by NP1 (17). A number of other viral proteins, including herpesvirus simplex virus (HSV-1) ICP27 (18)(19)(20) and Kaposi sarcoma-associated herpesvirus (KSHV) ORF57 (21)(22)(23)(24), are known to regulate similar processes.…”