Biomedical subjects

T Timchenko

Publications and source records attributed to T Timchenko.

8 recordsLinked to original sources

PubMed2006

Infectivity of nanovirus DNAs: induction of disease by cloned genome components of Faba bean necrotic yellows virus.

Circumstantial evidence suggests that the genome of Faba bean necrotic yellows virus (FBNYV), a nanovirus, consists of eight distinct, circular, single-stranded DNAs, each of about 1 kb and encoding only one protein. Here, the use of cloned full-length FBNYV DNAs for reproducing FBNYV-like symptoms in Vicia faba, the principal natural host of FBNYV, is reported. Characteristic symptoms of FBNYV infection were obtained in faba bean plants following biolistic DNA delivery or agroinoculation with all eight FBNYV DNAs. Although the eight different DNAs have been invariably detected in field samples infected with the various geographical FBNYV isolates, experimental infection with different combinations of fewer than eight DNAs also led to typical FBNYV symptoms. Even only five genome components, DNA-R, DNA-S, DNA-M, DNA-U1 and DNA-U2, were sufficient for inducing disease symptoms in V. faba upon agroinoculation. Symptomatic plants agroinoculated or bombarded with eight DNAs contained typical FBNYV virions; however, the virus was not transmitted by Aphis craccivora or Acyrthosiphon pisum, two efficient aphid vectors of FBNYV.

Animals↗

PubMed2000

The master rep concept in nanovirus replication: identification of missing genome components and potential for natural genetic reassortment.

Faba bean necrotic yellows virus (FBYNV), Milk vetch dwarf virus (MDV), and Subterranean clover stunt virus (SCSV) are nanoviruses that infect leguminous plants. From MDV- and SCSV-infected tissue we identified viral DNAs that encode a replication initiator protein (Rep), essential for replication of the multiple circular single-stranded DNAs of these viruses. These previously undescribed Rep proteins of MDV and SCSV are strikingly similar in sequence and functionally equivalent to the master Rep protein of FBYNV. Moreover, we demonstrated that the master Rep proteins of the three viruses are able to trigger replication of heterologous nanovirus DNAs. Such cross-species replication may reflect a considerable potential for genetic reassortment among nanoviruses in nature and be of significance for their evolution.

Amino Acid Sequence↗

PubMed2000

Clink, a nanovirus-encoded protein, binds both pRB and SKP1.

Clink, a 20-kDa protein of faba bean necrotic yellows virus, a single-stranded DNA plant virus, interacts with pRB family members and a SKP1 homologue from Medicago sativa. An LxCxE motif and an F-box of Clink mediate the interactions with the respective proteins. The capacity of Clink to bind pRB correlates with its ability to stimulate viral replication. Interaction of a single protein with the cell cycle regulator pRB and SKP1, a constituent of the ubiquitin-protein turnover pathway, appears to be a novel feature. Hence, Clink may represent a new class of viral cell cycle modulators.

Amino Acid Motifs↗

PubMed1999

A single rep protein initiates replication of multiple genome components of faba bean necrotic yellows virus, a single-stranded DNA virus of plants.

Faba bean necrotic yellows virus (FBNYV) belongs to the nanoviruses, plant viruses whose genome consists of multiple circular single-stranded DNA components. Eleven distinct DNAs, 5 of which encode different replication initiator (Rep) proteins, have been identified in two FBNYV isolates. Origin-specific DNA cleavage and nucleotidyl transfer activities were shown for Rep1 and Rep2 proteins in vitro, and their essential tyrosine residues that catalyze these reactions were identified by site-directed mutagenesis. In addition, we showed that Rep1 and Rep2 proteins hydrolyze ATP, and by changing the key lysine residue in the proteins' nucleoside triphosphate binding sites, demonstrated that this ATPase activity is essential for multiplication of virus DNA in vivo. Each of the five FBNYV Rep proteins initiated replication of the DNA molecule by which it was encoded, but only Rep2 was able to initiate replication of all the six other genome components. Furthermore, of the five rep components, only the Rep2-encoding DNA was always detected in 55 FBNYV samples from eight countries. These data provide experimental evidence for a master replication protein encoded by a multicomponent single-stranded DNA virus.

Adenosine Triphosphatases↗

PubMed1998

Ten distinct circular ssDNA components, four of which encode putative replication-associated proteins, are associated with the faba bean necrotic yellows virus genome.

Four further circular ssDNA components (C7-C10), about 1 kb in size and structurally similar to the previously described components (C1-C6) found associated with a Syrian (Sy) isolate of faba bean necrotic yellows virus (FBNYV), have been identified. Similar to C1 and C2, two of the new components (C7 and C9) encode putative replication-associated (Rep) proteins of 33.2 and 32.7 kDa, respectively, the former of which is 90% identical to the C10 Rep protein of milk vetch dwarf virus (MDV). C8 encodes a putative protein (17.4 kDa) whose function is unknown, but which is highly conserved between FBNYV and the other nanoviruses MDV, subterranean clover stunt virus and banana bunchy top virus. The putative protein (19.7 kDa) encoded by C10 contains an LXCXE motif, which is also present in the homologues of its relatives, suggesting that they may all interact with plant retinoblastoma-like proteins. Sequence information for seven components of an Egyptian (Eg) FBNYV isolate indicated that six of them share > 96% identity with FBNYV-Sy. However, the C1 Rep protein of FBNYV-Eg was only 63.5% identical to that of FBNYV-Sy, but was 88.3 % identical to the MDV-C2 Rep protein. We conclude that the FBNYV genome consists of seven to ten components, six of which encode non-Rep proteins. All ten components of the FBNYV genome, except the Rep components C2 and C9, had closely related counterparts in the MDV genome. In spite of this similarity, FBNYV and MDV appear to be distinct virus species.

Amino Acid Sequence↗

PubMed1996

Btcd, a mouse protein that binds to curved DNA, can substitute in Escherichia coli for H-NS, a bacterial nucleoid protein.

In an Escherichia coli mutant devoid of H-NS, a bacterial nucleoid protein, mouse protein Btcd was able to substitute for H-NS in two tested functions. It restored cell motility and repression of the expression of the bgl operon. Btcd1, a mutant Btcd protein deleted of its zinc finger and thus having reduced DNA binding, failed to substitute for H-NS. Mouse protein Btcd was shown to repress the bgl operon at the level of transcription initiation and to bind preferentially to a curved DNA fragment encompassing the bgl promoter. These effects of Btcd on bacterial gene transcription can be accounted for by the binding of Btcd or H-NS to a curved DNA sequence near a promoter. A few mammalian proteins have been shown to substitute for their Escherichia prototypes involved in DNA and RNA transactions. The efficiency of Btcd protein in substituting for H-NS in Escherichia suggests its possible involvement in regulating gene expression in mouse cells.

Animals↗

PubMed1995

The mouse Kin-17 gene codes for a new protein involved in DNA transactions and is akin to the bacterial RecA protein.

We have sought to characterize the molecular basis of the sensitivity to ionising radiation and to identify the genes involved in the cellular response of mammalian cells to such radiation. Using the Escherichia coli model, we tested the hypothesis that functional domains of RecA protein are represented in proteins of mammalian cells. We review here the results obtained in the detection of nuclear proteins of mammalian cells that are recognized by anti-RecA antibodies. We have called them kin proteins. Kin proteins likely play a role in DNA metabolism. We summarize the cloning of the mouse Kin-17 cDNA and our work on the identification and preliminary characterisation of the biochemical properties of mouse kin17 protein, a new nuclear protein able to recognize bent DNA and suspected to be involved in illegitimate recombination. We briefly describe our latest experiments on the molecular characterisation of the mouse Kin-17 gene. Finally, we discuss the properties of kin17 protein and the possible participation of kin17 protein in DNA transactions like transcription or recombination.

Animals↗

PubMed1994

Kin17, a mouse nuclear zinc finger protein that binds preferentially to curved DNA.

Kin17 is a 45 kDa protein encoded by the KIN17 gene located on mouse chromosome 2, band A. The kin17 amino acid sequence predicts two domains, which were shown to be functional: (i) a bipartite nuclear localization signal (NLS) that can drive the protein to the cell nucleus, (ii) a bona fide zinc finger of the C2H2 type. The zinc finger is involved in kin17 binding to double-stranded DNA since a mutant deleted of the zinc finger, kin17 delta 1, showed reduced binding. Single-stranded DNA was bound poorly by kin17. Interestingly, we found that kin17 protein showed preferential binding to curved DNA from either pBR322 or synthetic oligonucleotides. Binding of kin17 to a non-curved DNA segment increased after we had inserted into it a short curved synthetic oligonucleotide. Kin17 delta 2, a mutant deleted of 110 amino acids at the C-terminal end, still exhibited preferential binding to curved DNA and so did kin17 delta 1, suggesting that a domain recognizing curved DNA is located in the protein core.

Amino Acid Sequence↗