The type IC hsd loci of the enterobacteria are flanked by DNA with high homology to the phage P1 genome: implications for the evolution and spread of DNA restriction systems

Eco R124I, Eco DXXI and Eco prrI are the known members of the type IC family of DNA restriction and modification systems. The first three are carried on large, conjugative plasmids, while Eco prrI is chromosomally encoded. The enzymes are coded by three genes, hsdR , hsdM and hsdS . Analysis of the DNA sequences upstream and downstream of the type IC hsd loci shows that all are highly homologous to each other and also to sequences present in the bacteriophage P1 genome. The upstream sequences include functional phd and doc genes, which encode an addiction system that stabilizes the P1 prophage state, and extend to and beyond pac , the site at which phage DNA packaging begins. Downstream of the hsd loci, P1 DNA sequences begin at exactly the same place for all of the systems. For Eco DXXI and Eco prrI the P1 homology extends for thousands of base pairs while for Eco R124I an IS 1 insertion and an associated deletion have removed most of the P1-homologous sequences. The significance of these results for the evolution of DNA restriction and modification systems is discussed.     

Epidemic management and control through risk-dependent individual contact interventions

Testing, contact tracing, and isolation (TTI) is an epidemic management and control approach that is difficult to implement at scale because it relies on manual tracing of contacts. Exposure notification apps have been developed to digitally scale up TTI by harnessing contact data obtained from mobile devices; however, exposure notification apps provide users only with limited binary information when they have been directly exposed to a known infection source. Here we demonstrate a scalable improvement to TTI and exposure notification apps that uses data assimilation (DA) on a contact network. Network DA exploits diverse sources of health data together with the proximity data from mobile devices that exposure notification apps rely upon. It provides users with continuously assessed individual risks of exposure and infection, which can form the basis for targeting individual contact interventions. Simulations of the early COVID-19 epidemic in New York City are used to establish proof-of-concept. In the simulations, network DA identifies up to a factor 2 more infections than contact tracing when both harness the same contact data and diagnostic test data. This remains true even when only a relatively small fraction of the population uses network DA. When a sufficiently large fraction of the population (≳ 75%) uses network DA and complies with individual contact interventions, targeting contact interventions with network DA reduces deaths by up to a factor 4 relative to TTI. Network DA can be implemented by expanding the computational backend of existing exposure notification apps, thus greatly enhancing their capabilities. Implemented at scale, it has the potential to precisely and effectively control future epidemics while minimizing economic disruption.     

Biosensors and molecular imaging

The increase in the understanding of the physical and functional properties of the biological material, from the cellular level down to single molecules, owes its success to the development of suitable high-sensitivity platforms to image the biomaterial and analyze its response to specific stimuli. Imaging has indeed reached molecular capabilities, thanks to optical or magnetic markers [1], to the atomic force microscopy (AFM) in surface reconstruction [2], and is nearing success in three-dimensional (3-D) reconstruction thanks to X-ray holography [3].     

Multiple effects of paclitaxel are modulated by a high c-myc amplification level

Paclitaxel affects microtubule stability by binding to beta-tubulin, thus leading to cell accumulation in the G(2)/M phase, polyploidization, and apoptosis. Because both cell proliferation and apoptosis could be somehow regulated by the protooncogene c-myc, in this work we have investigated whether the c-myc amplification level could modulate the multiple effects of paclitaxel. To this aim, paclitaxel was administered to SW613-12A1 and -B3 human colon carcinoma cell lines (which are characterized by a high and low c-myc endogenous amplification level, respectively), and to the B3mycC5 cell line, with an enforced exogenous expression of c-myc copies. In this experimental system, we previously demonstrated that a high endogenous/exogenous level of amplification of c-myc enhances serum deprivation- and DNA damage-induced apoptosis. Accordingly, the present results indicate that a high c-myc amplification level potentiates paclitaxel cytotoxicity, confers a multinucleated phenotype, and promotes apoptosis to a great extent, thus suggesting that c-myc expression level is relevant in modulating the cellular responses to paclitaxel. We have recently shown in HeLa cells that the phosphorylated form of c-Myc accumulates in the nucleus, as distinct nucleolar and extranucleolar spots; here, we demonstrated that, after the treatment with paclitaxel, phosphorylated c-Myc undergoes redistribution, becoming diffused in the nucleoplasm.     

Muscle differentiation in a colonial ascidian: organisation, gene expression and evolutionary considerations

Background  

Ascidians are tunicates, the taxon recently proposed as sister group to the vertebrates. They possess a chordate-like swimming larva, which metamorphoses into a sessile adult. Several ascidian species form colonies of clonal individuals by asexual reproduction. During their life cycle, ascidians present three muscle types: striated in larval tail, striated in the heart, and unstriated in the adult body-wall.     

Human Papillomavirus Type 16 (HPV-16) Virus-Like Particle L1-Specific CD8+ Cytotoxic T Lymphocytes (CTLs) Are Equally Effective as E7-Specific CD8+ CTLs in Killing Autologous HPV-16-Positive Tumor Cells in Cervical Cancer Patients: Implications for L1 Dendritic Cell-Based Therapeutic Vaccines

Papillomavirus-like particles (VLPs) based on L1 capsid protein represent a promising prophylactic vaccine against human papillomavirus (HPV) infections. However, cell-mediated immune responses against this antigen are believed to be of limited therapeutic value in established HPV-infected cervical lesions and, for this reason, have not been intensively investigated in cervical cancer patients. In this study we analyzed and quantified by real-time PCR (RT-PCR) the RNA expression levels of E6, E7, and L1 genes in flash-frozen HPV-16 cervical carcinomas. In addition, the kinetics of expression of E6, E7, and L1 in HPV-16-infected primary cell lines established as long-term cultures in vitro was also evaluated at RNA and protein levels. Finally, in order to evaluate the therapeutic potential of L1-specific CD4⁺ and CD8⁺ T lymphocytes responses in cervical cancer patients, L1 VLP-loaded dendritic cells (DCs) were used to stimulate peripheral blood lymphocytes from cervical cancer patients and such responses were compared to those elicited by the E7 oncoprotein. We show that 22 of 22 (100%) flash-frozen cervical biopsy samples collected from HPV-16-positive cervical cancer patients harbor L1, in addition to E6 and E7 RNA, as detected by RT-PCR. E7 RNA copy number (mean, 176.2) was significantly higher in HPV-16-positive cervical cancers compared to the E6 RNA copy number (mean, 47.3) and the L1 copy number (mean, 58.3) (P < 0.0001 and P < 0.001, respectively). However, no significant differences in expression levels between E6 and L1 were found. Kinetic studies of E6, E7, and L1 RNA and protein expression levels in primary tumors showed a sharp reduction in L1 expression after multiple in vitro passages compared to E6 and E7. Autologous DCs pulsed with HPV-16 VLPs or recombinant full-length E7 elicited strong type 1 L1- and E7-specific responses in CD4⁺ and CD8⁺ T cells from cervical cancer patients. Importantly, L1 VLP-specific CD8⁺ T lymphocytes expressed strong cytolytic activity against autologous tumor cells and were as effective as E7-specific cytotoxic T lymphocytes in lysing naturally HPV-16-infected autologous tumor cells. Taken together, these data demonstrate a consistent expression of L1 in primary cervical tumors and the possibility of inducing effective L1/tumor-specific CD4⁺ and CD8⁺ T-lymphocyte responses in patients harboring HPV-infected cervical cancer. These results may have important implications for the treatment of patients harboring established HPV-infected lesions with L1 VLPs or combined E7/L1 DC-based vaccinations.Human papillomavirus (HPV) infection represents the most important risk factor for the development of cervical cancer. Although more than 100 distinct HPV genotypes have been described, and at least 20 are associated with cervical cancer, HPV type 16 (HPV-16) is by far the most frequently detected in cervical neoplasia regardless of the geographical origin of the patients (4). In the last few years significant advances have been made in the development of candidate prophylactic vaccine against cervical cancer and HPV-related infections. In several large prospective randomized studies, virus-like particles consisting of the HPV-16 and HPV-18 major capsid protein L1 (L1-VLPs) have shown promise in protecting young healthy females against persistent infection with HPV-16 and HPV-18 and their associated cervical intraepithelial neoplasia (reviewed in reference 12). These data strongly suggest that the implementation of large-scale L1-VLP-based prophylactic vaccinations have the potential to dramatically reduce worldwide cervical cancer rates in the years to come.Unfortunately, because HPV infection is endemic in humans and there is a long latency from HPV infection to the development of invasive cervical cancer in women, even if prophylactic L1-based vaccinations are implemented on a worldwide scale today it would take decades to perceive any significant benefit. Consistent with this view, an estimated 5 million cervical cancer deaths will occur in the next 20 years due to existing HPV infections (4, 12). Thus, the current development of therapeutic vaccines for protection against persistent HPV infections, cervical cancer, and its precursor lesions remains an area of great interest.Although the interactions between the host immune system and HPV-infected cells are still not completely understood, several lines of evidence suggest that protection against HPV-related infections by L1-VLP-based vaccines is likely conferred by the generation of high levels of neutralizing antibodies (12, 38). Nevertheless, a potential crucial role of L1-specific T-cell responses and the involvement of T cells in mediating the production of neutralizing antibodies and antiviral effect in infected hosts has been previously hypothesized (8, 24). This point may be particularly noteworthy in patients harboring HPV-infected cervical lesions because several studies have demonstrated the critical importance of both cytotoxic (CD8⁺) and helper (CD4⁺) T cells in achieving clinical responses (1, 5, 16-18, 20, 23). However, limited information is currently available to evaluate whether cell-mediated immune responses to L1-VLP may have any significant therapeutic effect in cervical cancer patients harboring HPV-16 positive tumors. Furthermore, to our knowledge, no direct comparison of the therapeutic efficacy of L1 and E7-specific immune responses against naturally HPV-16-infected cervical cancer have been yet reported in human patients.In the present study we have analyzed and quantified by highly sensitive real-time PCR (RT-PCR) the RNA levels of E6, E7, and L1 in flash-frozen biopsy specimens obtained from HPV-16-infected cervical carcinomas and in short- and long-term primary cultures of HPV-16-positive cervical tumors. In addition, we have studied the kinetics of expression of these genes and proteins during the establishment of HPV-16-positive primary tumors in vitro. Finally, using completely autologous systems of naturally infected HPV-16-positive human tumors, we have carefully studied the phenotype and function of L1-specific CD4⁺ and CD8⁺ T-lymphocyte responses generated by VLP-loaded dendritic cells (DCs) and compared their therapeutic potential to those elicited by DC loaded with the E7 oncoprotein.     

Genetic mapping and QTL analysis in European hazelnut (<Emphasis Type="Italic">Corylus avellana</Emphasis> L.)

The European hazelnut (Corylus avellana L.) is the most economically important nut species in the Betulaceae family. Despite the need for new improved hazelnut cultivars, few breeding programs are carried out because of the large plant size, the long lifecycle of the plant, and the expense and time required. To date, there are no reports of maps with quantitative trait loci (QTL) in hazelnut. Our objective in the present study was to identify QTL associated with vegetative traits to allow marker-assisted selection (MAS). An F1 progeny (275 plants) of Tonda Gentile delle Langhe × Merveille de Bollwiller obtained in 2009 was used to develop a QTL linkage map for vigour, sucker habit, and time of bud burst, after three years of observations. A set of 163 plants were analysed with 152 microsatellite markers. A map of 11 linkage groups was obtained, covering 663.1 cM, and 15 QTLs were identified and mapped for the traits examined. Of them, 10 were ‘major’ QTL, including a stably expressed region on LG_02 for leaf bud burst. At least one major QTL for each year underlies the variation in each trait and a clustering of QTL for trunk circumference and suckers/trunk circumference ratio with high inter-trait correlations was observed on LG_05, suggesting a single pleiotropic locus. This research represents an initial step in the future identification of chromosomal regions carrying genes of interest, important for breeding programs and MAS.     

A Robust Method of Measuring Other-Race and Other-Ethnicity Effects: The Cambridge Face Memory Test Format

Other-race and other-ethnicity effects on face memory have remained a topic of consistent research interest over several decades, across fields including face perception, social psychology, and forensic psychology (eyewitness testimony). Here we demonstrate that the Cambridge Face Memory Test format provides a robust method for measuring these effects. Testing the Cambridge Face Memory Test original version (CFMT-original; European-ancestry faces from Boston USA) and a new Cambridge Face Memory Test Chinese (CFMT-Chinese), with European and Asian observers, we report a race-of-face by race-of-observer interaction that was highly significant despite modest sample size and despite observers who had quite high exposure to the other race. We attribute this to high statistical power arising from the very high internal reliability of the tasks. This power also allows us to demonstrate a much smaller within-race other ethnicity effect, based on differences in European physiognomy between Boston faces/observers and Australian faces/observers (using the CFMT-Australian).