Alternative splicing of pre-mRNAs encoding the nonstructural proteins of minute virus of mice is facilitated by sequences within the downstream intron.

ABSTRACT The production of wild-type-free stocks of recombinant parvovirus minute virus of mice [MVM(p)] is difficult due to the presence of homologous sequences in vector and helper genomes that cannot easily be eliminated from the overlapping coding sequences. We have therefore cloned and sequenced spontaneously occurring defective particles of MVM(p) with very small genomes to identify the minimalcis-acting sequences required for DNA amplification and virus production. One of them has lost all capsid-coding sequences but is still able to replicate in permissive cells when nonstructural proteins are provided in trans by a helper plasmid. Vectors derived from this particle produce stocks with no detectable wild-type MVM after cotransfection with new, matched, helper plasmids that present no homology downstream from the transgene.

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The Expression Strategy of Goose Parvovirus Exhibits Features of both the Dependovirus and Parvovirus Genera

Journal of Virology ◽

10.1128/jvi.79.17.11035-11044.2005 ◽

2005 ◽

Vol 79 (17) ◽

pp. 11035-11044 ◽

Cited By ~ 35

Author(s):

Jianming Qiu ◽

Fang Cheng ◽

Yuko Yoto ◽

Zoltán Zádori ◽

David Pintel

Keyword(s):

High Efficiency ◽

Nonstructural Protein ◽

Nonstructural Proteins ◽

Coding Region ◽

Minute Virus Of Mice ◽

Adeno Associated Virus ◽

Goose Parvovirus ◽

Minute Virus ◽

The Right ◽

A Site

ABSTRACT The RNA transcription profile of the goose parvovirus (GPV) was determined, and it is a surprising hybrid of features of the Parvovirus and Dependovirus genera of the Parvovirinae subfamily of the Parvoviridae. Similar to the Dependovirus adeno-associated virus type 5, RNAs transcribed from the GPV upstream P9 promoter, which encode the viral nonstructural proteins, were polyadenylated at a high efficiency at a polyadenylation site [(pA)p] located within an intron in the center of the genome. Efficient usage of (pA)p required a downstream element that overlaps with the polypyrimidine tract of the A2 3′ splice site of the central intron. An upstream element required for efficient use of (pA)p was also identified. RNAs transcribed from the P42 promoter, presumed to encode the viral capsid proteins, primarily extended through (pA)p and were polyadenylated at a site, (pA)d, located at the right end of the genome and ultimately spliced at a high efficiency. No promoter analogous to the Dependovirus P19 promoter was detected; however, similar to minute virus of mice and other members of the Parvovirus genus, a significant portion of pre-mRNAs generated from the P9 promoter were additionally spliced within the putative GPV Rep1 coding region and likely encode an additional, smaller, nonstructural protein. Also similar to members of the Parvovirus genus, detectable activity of the GPV P42 promoter was highly dependent on transactivation by the GPV Rep1 protein in a manner dependent on binding to a cis-element located in the P42 promoter.

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The cytotoxicity of the autonomous parvovirus minute virus of mice nonstructural proteins in FR3T3 rat cells depends on oncogene expression.

Journal of Virology ◽

10.1128/jvi.68.10.6446-6453.1994 ◽

1994 ◽

Vol 68 (10) ◽

pp. 6446-6453 ◽

Cited By ~ 45

Author(s):

S Mousset ◽

Y Ouadrhiri ◽

P Caillet-Fauquet ◽

J Rommelaere

Keyword(s):

Nonstructural Proteins ◽

Minute Virus Of Mice ◽

Oncogene Expression ◽

Minute Virus

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Limitations to the expression of parvoviral nonstructural proteins may determine the extent of sensitization of EJ-ras-transformed rat cells to minute virus of mice

Virology ◽

10.1016/0042-6822(89)90514-x ◽

1989 ◽

Vol 171 (1) ◽

pp. 89-97 ◽

Cited By ~ 21

Author(s):

Benoît Van Hille ◽

Nadine Duponchel ◽

Nathalie Salomé ◽

Nathalie Spruyt ◽

Sue F. Cotmore ◽

...

Keyword(s):

Nonstructural Proteins ◽

Minute Virus Of Mice ◽

Minute Virus

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Intron Definition Is Required for Excision of the Minute Virus of Mice Small Intron and Definition of the Upstream Exon

Journal of Virology ◽

10.1128/jvi.72.3.1834-1843.1998 ◽

1998 ◽

Vol 72 (3) ◽

pp. 1834-1843 ◽

Cited By ~ 16

Author(s):

Donald D. Haut ◽

D. J. Pintel

Keyword(s):

Alternative Splicing ◽

Consensus Sequence ◽

Critical Role ◽

Viral Mrnas ◽

Minute Virus Of Mice ◽

Natural Context ◽

Minute Virus ◽

Definition Of ◽

Intron Definition ◽

Upstream Exon

ABSTRACT Alternative splicing of pre-mRNAs plays a critical role in maximizing the coding capacity of the small parvovirus genome. The small-intron region of minute virus of mice (MVM) pre-mRNAs undergoes an unusual pattern of overlapping alternative splicing—using two donors (D1 and D2) and two acceptors (A1 and A2) within a region of 120 nucleotides—that determines the steady-state ratios of the various viral mRNAs. In this report, we show that the determinants that govern excision of the small intron are complex and are also required for efficient definition of the upstream exon. For the MVM small intron in its natural context, the two donors appear to compete for the splicing machinery: the position of D1 favors its usage, while the primary sequence of D2 must be more like the consensus sequence than is D1 to be used efficiently. We have genetically defined the branch points that are used for generation of the major and minor spliced forms and show that recognition of components of the small-intron acceptors is likely to be the dominant determinant in alternative small-intron excision. We have also identified a G-rich intronic enhancer sequence within the small intron that is essential for splicing of the minor form (D2 to A2) but not the major form (D1 to A1) of MVM mRNAs and is required for efficient definition of the upstream NS2-specific exon. In its natural context, the small intron appears to be excised by a mechanism consistent with intron definition. When the MVM small intron is expanded, various parameters of its excision are altered, indicating that critical cis-acting signals are context dependent. Relative use of the donors and acceptors is altered, and the upstream NS2-specific exon is no longer efficiently defined. The fact that definition of the upstream NS2-specific exon can be achieved by the MVM small intron in its natural context, but not when it is expanded, suggests that the multiple determinants that govern definition and excision of the small intron are required, in concert, for upstream exon definition. Our data are consistent with a model in which alternative splicing of the MVM P4-generated pre-mRNAs is governed by a hybrid of intron- and exon-defining mechanisms.

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