E2 protein is the major determinant of specificity at the human papillomavirus origin of replication

The replication of human papillomavirus (HPV) genomes requires E1 and E2 proteins as the viral trans-factors and the replication origin, located in the URR, as a cis-element. The minimal requirements for an HPV replication origin vary among different virus types but always include one or more binding sites for the E2 protein. The requirements for an E1 binding site seem to vary among different HPV genera, with alpha-HPV11 and -18 minimal origins able to replicate without E1 binding site in contrast to beta-HPV8. In the present article, we analysed the sequence requirements for the beta-HPV5 minimal origin of replication. We show that the HPV5 URR is able to replicate in U2OS cells without the sequence proposed as an E1 binding site, albeit at lower levels than wt URR, given that three E2 binding sites are intact and both viral replication proteins are present. The lack of an absolute requirement of the E1 binding site for the origin of replication of HPV5 led us to analyse whether the viral E1 and E2 proteins from other HPV types are competent to support replication from this origin. Surprisingly, the E1 and E2 proteins from beta-HPV types support replication from the origin in contrast to proteins from alpha-HPV types 11, -16, or -18. Furthermore, the replication proteins E1 and E2 of these alpha-HPV types are unable to support the replication of HPV5 URR, even if the E1 binding site is intact. In light of these results, we performed a detailed analysis of the ability of different combinations of E1 and E2 proteins from various alpha- and beta-HPV types to support the replication of URR sequences from the respective HPV types in the U2OS cell line

Assessment of the olfactory function in Italian patients with type 3 von Willebrand disease caused by a homozygous 253 Kb deletion involving VWF and TMEM16B/ANO2.

Type 3 Von Willebrand disease is an autosomal recessive disease caused by the virtual absence of the von Willebrand factor (VWF). A rare 253 kb gene deletion on chromosome 12, identified only in Italian and German families, involves both the VWF gene and the N-terminus of the neighbouring TMEM16B/ANO2 gene, a member of the family named transmembrane 16 (TMEM16) or anoctamin (ANO). TMEM16B is a calcium-activated chloride channel expressed in the olfactory epithelium. As a patient homozygous for the 253 kb deletion has been reported to have an olfactory impairment possibly related to the partial deletion of TMEM16B, we assessed the olfactory function in other patients using the University of Pennsylvania Smell Identification Test (UPSIT). The average UPSIT score of 4 homozygous patients was significantly lower than that of 5 healthy subjects with similar sex, age and education. However, 4 other members of the same family, 3 heterozygous for the deletion and 1 wild type, had a slightly reduced olfactory function indicating that socio-cultural or other factors were likely to be responsible for the observed difference. These results show that the ability to identify odorants of the homozygous patients for the deletion was not significantly different from that of the other members of the family, showing that the 253 kb deletion does not affect the olfactory performance. As other genes may compensate for the lack of TMEM16B, we identified some predicted functional partners from in silico studies of the protein-protein network of TMEM16B. Calculation of diversity for the corresponding genes for individuals of the 1000 Genomes Project showed that TMEM16B has the highest level of diversity among all genes of the network, indicating that TMEM16B may not be under purifying selection and suggesting that other genes in the network could compensate for its function for olfactory ability

Cis and trans requirements for stable episomal maintenance of the BPV-1 replicator.

Papillomavirus genomes are maintained as multicopy nuclear plasmids in transformed cells. To address the mechanisms by which the viral DNA is stably propagated in the transformed cells, we have constructed a cell line CH04.15 expressing constitutively the viral proteins E1 and E2, that are required for initiation of viral DNA replication. We show that these viral proteins are necessary and sufficient for stable extrachromosomal replication. Using the cell line CH04.15, we have shown that the bovine papillomavirus-1 (BPV-1) minimal origin of replication (MO) is absolutely necessary, but is not sufficient for stable extrachromosomal replication of viral plasmids. By deletion and insertion analysis, we identified an additional element (minichromosome maintenance element, MME) in the upstream regulatory region of BPV-1 which assures stable replication of the MO-containing plasmids. This element is composed of multiple binding sites for the transcription activator E2. MME appears to function in the absence of replication but requires E1 and E2 proteins for activity. In contrast to, for example, Epstein-Barr virus oriP, stably maintained BPV-1 plasmids are not subject to once-per-cell cycle replication as determined by density labelling experiments. These results indicate that papillomavirus episomal replicators replicate independently of the chromosomal DNA of their hosts

The structure–activity interactions of Cu/Zn, In/Pd and Fe/K catalysts supported on mesoporous SBA-15 for carbon dioxide hydrogenation at low pressure

To minimize greenhouse gas emissions, efficient carbon dioxide capture and utilization need to be addressed. In this study, to determine the structure–activity interplay, three different promising catalytic systems for the CO2 hydrogenation process were synthesized using mesoporous silica SBA-15 as a support material: copper-based catalyst with zinc, indium-based catalyst with palladium and iron-based catalyst with potassium. The role of metal–metal oxide interaction has been showed. The use of Cu/Zn catalytic system and SBA-15 allowed to obtain very small crystallite size of tenorite and zinc oxide, good dispersion of active phases with strong basic sites. In order to find the most effective catalyst providing the maximal methanol yield and selectivity, these catalytic systems were compared under the same reaction conditions (250 °C, 20 bar, H2 to CO2 molar ratio 4 to 1) using fixed-bed tubular micro-activity reactor. Results showed that the highest methanol yield can be obtained with Cu/Zn/SBA-15 catalyst as might be expected according to obtained characterization