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Replication Activity (replication + activity)
Selected AbstractsVirological analysis, genotypes and mutational patterns of the HBV precore/core gene in HBV/HCV-related hepatocellular carcinomaJOURNAL OF VIRAL HEPATITIS, Issue 9 2006M. S. De Mitri Summary., We investigated the replicative profile of hepatitis B (HBV) and hepatitis C (HCV) viruses and the mutational pattern of the HBV precore/core (pre-C/C) domain in hepatocellular carcinoma (HCC). Thirty-eight consecutive patients with HCC were included in the study , 18 of them with HBV/HCV co-infection and 20 with HBV single infection. Twenty-three additional patients with co-infection, without HCC were recruited as the control group. Replication activity was evaluated by detecting and quantitating both HBV and HCV genomes. The HBV pre-C/C region, encompassing the pregenome encapsidation signal involved in viral replication, was analysed by direct sequencing. HBV viraemia levels were significantly lower (P = 0.04) in patients with co-infection in comparison with single-infected HCC, whereas two different HBV viraemia profiles were detected in co-infection with or without circulating HCV. HBV genotype D was prevalent in the three groups and HCV genotype 1b was found to be the infecting strain in all patients. Lower variability in the pre-C/C region was found in co-infection in comparison with HBV single infection (P = 0.0004). A synonymous T1936C mutation was found in all co-infected HCC cases not related to the presence or absence of circulating HCV, and a hypermutated pre-C strain, characterized by the same mutational pattern, was identified in three HCC cases. The mutational pattern of the pre-C/C region was closely related to HBV replication efficiency, and specific HBV mutations selectively associated with HCV co-infection could be linked with accelerated HBV/HCV-related disease progression. [source] Cancer-associated missplicing of exon 4 influences the subnuclear distribution of the DNA replication factor CIZ1,HUMAN MUTATION, Issue 10 2007Faisal Abdel Rahman Abstract Cip1-interacting zinc finger protein 1 (CIZ1, also known as CDKN1A-interacting zinc finger protein 1) stimulates initiation of mammalian DNA replication and is normally tethered to the nuclear matrix within DNA replication foci. Here, we show that an alternatively spliced human CIZ1 variant, lacking exon 4 (, E4), is misexpressed as a consequence of intronic mutation in Ewing tumor (ET) cell lines. In all ET lines tested, exon 4 is skipped and an upstream mononucleotide repeat element is expanded to contain up to 28 thymidines, compared to 16 in controls. In exon-trap experiments, a 24T variant produced three-fold more exon skipping than a 16T variant, demonstrating a direct effect on splicing. In functional assays, , E4 protein retains replication activity, but fails to form subnuclear foci. Furthermore, coexpression of mouse , E4 with Ciz1 prevents Ciz1 from localizing appropriately, having a dominant negative effect on foci formation. The data show that conditional exclusion of exon 4 influences the spatial distribution of the Ciz1 protein within the nucleus, and raise the possibility that CIZ1 alternative splicing could influence organized patterns of DNA replication. Hum Mutat 28(10), 993,1004, 2007. © 2007 Wiley-Liss, Inc. [source] Functional analysis of polyomavirus BK non-coding control region quasispecies from kidney transplant recipientsJOURNAL OF MEDICAL VIROLOGY, Issue 11 2009Gunn-Hege Olsen Abstract Replication of the human polyomavirus BK (BKV) in renal tubular epithelial cells causes viruria and BKV-nephropathy in kidney transplant recipients. Following prolonged high-level BKV replication, rearrangement of the archetype non-coding control region (NCCR) leads to a mixture of BKV variants. The aim of this study was to compare potential functional differences of 12 rearranged (rr)-NCCR variants with the archetype (ww)-NCCR (WWT) found in allograft biopsies or urine from three kidney transplant recipients including two with BKV-nephropathy. Twelve different rr-NCCRs and one archetype ww-NCCR were inserted between the early and late protein coding region of BKV(Dunlop) to make recombinant BKV genomes for transfection into Vero cells. Immunoblotting, immunofluorescence staining, and quantitative PCR demonstrated that viral protein expression and extracellular BKV loads of 10 rr-NCCR variants were similar or higher than observed for the ww-NCCR BKV. Two rr-NCCR variants (RH-2 and RH-19) were non-functional. The functional rr-NCCRs produced infectious progeny successfully infecting primary renal proximal tubular epithelial cells. The number of infected cells and extracellular BKV loads corresponded to the activity seen in Vero cells. Three rr-NCCR variants (RH-1, RH-10, RH-13) only gave rise to a few infected cells similar to ww-NCCR, whereas seven variants had intermediate activity (RH-5, RH-6, RH-8, RH-9, RH-11) or high replication activity (RH-7 and RH-18) with several hundred infected cells per well. The results indicate that both functional and non-functional BKV rr-NCCR variants arise during BKV replication in kidney transplant recipients and that most functional rr-NCCR variants confer a higher replication capacity than archetype ww-NCCR. J. Med. Virol. 81:1959,1967, 2009. © 2009 Wiley-Liss, Inc. [source] Positioning and the specific sequence of each 13-mer motif are critical for activity of the plasmid RK2 replication originMOLECULAR MICROBIOLOGY, Issue 5 2005Lukasz Kowalczyk Summary The minimal replication origin of the broad-host-range plasmid RK2, oriV, contains five iterons which are binding sites for the plasmid-encoded replication initiation protein TrfA, four DnaA boxes, which bind the host DnaA protein, and an AT-rich region containing four 13-mer sequences. In this study, 26 mutants with altered sequence and/or spacing of 13-mer motifs have been constructed and analysed for replication activity in vivo and in vitro. The data show that the replacement of oriV 13-mers by similar but not identical 13-mer sequences from Escherichia coli oriC inactivates the origin. In addition, interchanging the positions of the oriV 13-mers results in greatly reduced activity. Mutants with T/A substitutions are also inactive. Furthermore, introduction of single-nucleotide substitutions demonstrates very restricted sequence requirements depending on the 13-mer position. Only two of the mutants are host specific, functional in Pseudomonas aeruginosa but not in E. coli. Our experiments demonstrate considerable complexity in the plasmid AT-rich region architecture required for functionality. It is evident that low internal stability of this region is not the only feature contributing to origin activity. Our studies suggest a requirement for sequence-specific protein interactions within the 13-mers during assembly of replication complexes at the plasmid origin. [source] | ||||||||||