Two spatially distinct genetic elements constitute a bipartite DNA replication origin in the minute virus of mice genome

In this work, we report the transduction of a chloramphenicol acetyltransferase (CAT) reporter gene into a variety of normal and transformed human cells of various tissue origins. The vector used was MVM/P38cat, a recombinant of the prototype strain of the autonomous parvovirus minute virus of mice (MVMp). The CAT gene was inserted into the capsid-encoding region of the infectious molecular clone of MVMp genome, under the control of the MVM P38 promoter. When used to transfect permissive cells, the MVM/P38cat DNA was efficiently replicated and expressed the foreign CAT gene at high levels. By cotransfecting with a helper plasmid expressing the capsid proteins, it was possible to produce mixed virus stocks containing MVM/P38cat infectious particles and variable amounts of recombinant MVM. MVM/P38cat viral particles were successfully used to transfer the CAT gene and to express it in a variety of human cells. Both viral DNA replication and P38-driven CAT expression were achieved in fibroblasts, epithelial cells, T lymphocytes, and macrophages in a transformation-dependent way, but with an efficiency depending on the cell type. In transformed B

mentioning

confidence: 99%

Use of an autonomous parvovirus vector for selective transfer of a foreign gene into transformed human cells of different tissue origins and its expression therein

Dupont

Tenenbaum

Guo

et al. 1994

“…However, the binding site was very close to the 5Ј nick site of the genome predicted by comparison with the well-defined sequences within the MVM 5Ј-terminal palindrome ( Fig. 7A), suggesting that there may still be some association between binding of the protein and replication at the 5Ј end (9,16,18,43,50).…”

Section: Discussionmentioning

confidence: 64%

A Heterogeneous Nuclear Ribonucleoprotein A/B-Related Protein Binds to Single-Stranded DNA near the 5′ End or within the Genome of Feline Parvovirus and Can Modify Virus Replication

Wang

Parrish

1999

Phage display of cDNA clones prepared from feline cells was used to identify host cell proteins that bound to DNA-containing feline panleukopenia virus (FPV) capsids but not to empty capsids. One gene found in several clones encoded a heterogeneous nuclear ribonucleoprotein (hnRNP)-related protein (DBP40) that was very similar in sequence to the A/B-type hnRNP proteins. DBP40 bound specifically to oligonucleotides representing a sequence near the 5′ end of the genome which is exposed on the outside of the full capsid but did not bind most other terminal sequences. Adding purified DBP40 to an in vitro fill-in reaction using viral DNA as a template inhibited the production of the second strand after nucleotide (nt) 289 but prior to nt 469. DBP40 bound to various regions of the viral genome, including a region between nt 295 and 330 of the viral genome which has been associated with transcriptional attenuation of the parvovirus minute virus of mice, which is mediated by a stem-loop structure of the DNA and cellular proteins. Overexpression of the protein in feline cells from a plasmid vector made them largely resistant to FPV infection. Mutagenesis of the protein binding site within the 5′ end viral genome did not affect replication of the virus.

“…Moreover, the comparison of presently available genomic sequences of several related autonomous parvoviruses of vertebrates revealed 6-to 8-of-8-nt matches of PREs near to the resolution sites of the left-hand palindrome (Fig. 8), in the internal promoter (3,21,43,47), and in the right-hand palindrome (21,30,43,47,50). Among the 3Ј termini listed in Fig.…”

Section: Discussionmentioning

confidence: 91%

“…Minute virus of mice (MVM) is an autonomous parvovirus belonging to the family Parvoviridae, a group of linear singlestranded DNA viruses that parasitize arthropods, birds, and mammals, including humans (54). Their genomes (approximately 5 kb long) comprise terminal palindromic sequences at both ends which contain critical cis-acting elements required for viral DNA replication (15,50). Whereas members of the adeno-associated virus subgroup of parvoviruses mostly rely on helper viruses (adenoviruses and herpesviruses) for their multiplication (6), the autonomous parvoviruses replicate during the S phase in appropriate host cells (17).…”

mentioning

confidence: 99%

NF-Y controls transcription of the minute virus of mice P4 promoter through interaction with an unusual binding site

Plaza

Perros

et al. 1995

Electrophoretic mobility shift assays performed with nuclear extracts from human fibroblasts revealed the formation of two major protein complexes with an oligonucleotide (nucleotides 78 to 107) from the palindromic region located upstream from the minute virus of mice (MVM) P4 promoter. It was shown that this oligonucleotide bound USF at the enhancer E box CACATG. The second complex contained the transcription factor NF-Y, whose association was surprising because its target sequence lacks the canonical CCAAT motif present in all mammalian NF-Y binding sites identified so far. The MVM NF-Y recognition element instead contains the CCAAC sequence. USF and NF-Y had distinct but overlapping sequence requirements for binding, suggesting that their associations with MVM DNA were mutually exclusive. Because of the palindromic nature of MVM DNA terminal sequences, NF-Y associated with the three nucleotide configurations corresponding to the hairpin structure and to the external and internal arms of the extended duplex replication form, respectively. However, owing to the imperfection of the palindrome, the binding of USF was restricted to the internal arm. Point mutations that suppressed the in vitro binding of NF-Y to the internal palindromic arm reduced the activity of the resident P4 promoter, while those preventing complex formation with USF did not, as determined by transient expression assays using the luciferase reporter gene. The data led to the identification of a novel P4 upstream regulatory region capable of interacting with two transcription factors, from which one (NF-Y) appeared to upmodulate the activity of the promoter.