Contribution of type W human endogenous retroviruses to the human genome: Characterization of HERV-W proviral insertions and processed pseudogenes

Background: Human endogenous retroviruses (HERVs) are ancient sequences integrated in the germ line cells and vertically transmitted through the offspring constituting about 8% of our genome. In time, HERVs accumulated mutations that compromised their coding capacity. A prominent exception is HERV-W locus 7q21.2, producing a functional Env protein (Syncytin-1) coopted for placental syncytiotrophoblast formation. While expression of HERV-W sequences has been investigated for their correlation to disease, an exhaustive description of the group composition and characteristics is still not available and current HERV-W group information derive from studies published a few years ago that, of course, used the rough assemblies of the human genome available at that time. This hampers the comparison and correlation with current human genome assemblies. Results: In the present work we identified and described in detail the distribution and genetic composition of 213 HERV-W elements. The bioinformatics analysis led to the characterization of several previously unreported features and provided a phylogenetic classification of two main subgroups with different age and structural characteristics. New facts on HERV-W genomic context of insertion and co-localization with sequences putatively involved in disease development are also reported. Conclusions: The present work is a detailed overview of the HERV-W contribution to the human genome and provides a robust genetic background useful to clarify HERV-W role in pathologies with poorly understood etiology, representing, to our knowledge, the most complete and exhaustive HERV-W dataset up to date

Multiple phases in a generalized Gross-Witten-Wadia matrix model

We study a unitary matrix model of the Gross-Witten-Wadia type, extended with the addition of characteristic polynomial insertions. The model interpolates between solvable unitary matrix models and is the unitary counterpart of a deformed Cauchy ensemble. Exact formulas for the partition function and Wilson loops are given in terms of Toeplitz determinants and minors and large $N$ results are obtained by using Szeg\"o theorem with a Fisher-Hartwig singularity. In the large $N$ (planar) limit with two scaled couplings, the theory exhibits a surprisingly intricate phase structure in the two-dimensional parameter space.Comment: 22 page

Closed combination of context-embedding iterative strategies

This work is motivated by the challenging problem of the computer-aided generation of approximations (viewed as a series of transformations) of partial derivative equations. In this framework, the approximations posed over complex settings are incrementally constructed by extending an approximation posed on a simple setting and combining these extensions. In order to formalize these extensions and their combination, we introduce a class of rewriting strategies, called context-embedding iterative strategies (CE-strategies, for short). Roughly speaking, the class of CE-strategies is constructed by means of adding contexts and an iteration operator allowing the definition of recursive strategies. We show that the class of CE-strategies is closed under combination with respect to a correctness-completeness criterion. It turns out that the class CE-strategies enjoy nice algebraic properties, namely, the combination is associative, has a neutral element, and all the elements are idempotents

Exploiting high-throughput screens to optimize Adeno-Associated Viral Vectors for gene transfer into primary human keratinocytes

Chronic non-healing wounds such as diabetic ulcers or burns represent a devastating health problem with significant clinical, physical and social implications. The healing can be frustrating and painful for patients. The difficult healing process requires advanced therapeutic strategies such as the use of primary human keratinocytes (HK) as autologous transplants, which may be considered for clinical use. To improve engraftment or to introduce therapeutic genes into primary HK, efficient and safe vectors are required. One of the most promising vector systems today is based on the adeno-associated virus (AAV), a member of the parvovirus family. Recombinant AAV (rAAV) vectors possess a number of attractive properties including low immunogenicity, high stability and the potential to integrate site-specifically without known side-effects. Unfortunately, cell entry into primary HK of rAAV2 is barely detectable and consequentially, HK are poor targets of rAAV2-mediated transductions. As demonstrated in this thesis, primary HK do not express AAV2´s primary receptor heparan sulphate proteoglycan (HSPG), the presence of which, however, is required for binding to AAV2´s internalization receptors. Cell surface targeting allows re-directing the viral vector tropism towards a novel receptor mediating thereby transduction of cells in absence of AAV’s natural receptors. These AAV capsid mutants have displayed improved transduction efficiency in wild-type-AAV non-permissive cells and have provided the opportunity of rAAV-mediated, cell-type-specific gene transfer. As documented in this study, new rAAV vectors were developed as promising tools for modifying primary HK. Using an AAV peptide display library that displayed 7mer peptides of random sequence at capsid position 587; three AAV peptide insertion mutants differing in sequence of inserted ligand (Kera1, Kera2 and Kera3) were selected and subsequently analyzed. To select rAAV targeting vectors with a re-directed tropism, the library was optimized by depleting mutants capable of binding to HSPG prior to selection by heparin affinity chromatography. Furthermore, the selection was performed on primary HK obtained from different donors to target a common receptor and the selection pressure was continuously increased by decreasing the vector genomes per cell ratio to select for the fittest variant. The thereby developed rAAV targeting vectors Kera1 (RGDTATL), Kera2 (PRGDLAP) and Kera3 (RGDQQSL) showed a remarkable change in tropism, transducing primary HK with high efficiency and specificity even in mixed cultures of target and non-target cells. In this study, a novel microarray based bioinformatic approach (comparative gene analysis (CGA)), was used for the identification of the receptor that targeted the mutant that showed the most striking change in tropism, Kera2. Briefly, in cooperation with Giovanni Di Pasquale (NCI/NIH, Bethesda, USA), a screening of the NIH cell line panel was performed, pointing towards the involvement of beta8 integrin subunit for cell transduction by Kera2. Beta8 is unique as it is solely described as heterodimer with alpha V and the integrin αVβ8 could be detected on cell surface of primary human keratinocytes. By blocking experiments with blocking αV- or αVβ8-antibodies experimental evidence was provided that the integrin αVβ8 serves as receptor for Kera2. Finally, this study has shown that the targeting vectors Kera1, Kera2 and Kera3 transduced airlifted differentiated keratinocytes in organotypic 3D cultures. In summary, the three rAAV targeting vectors Kera1, Kera2 and Kera3, selected from an optimized library and using a novel selection strategy, are excellent candidates for successful application in clinical use

A structural alignment model of noun-noun compound interpretation

The interpretation of noun-noun compounds is complex, yet compounds such as 'web surfer' and 'beef baron' are generated and interpreted easily by native English speakers. Concept combination is the core process in the generation and interpretation o f noun-noun compounds. Such compounds may be read literally or metaphorically suggesting that the combination process is capable of both literal and metaphoric interpretations. The motivation for this thesis is to tackle three problems which occur in concept combination. These problems are: (1) compounds are often polysemous, (2) compounds often appear to be understood by evoking a context (or world knowledge) and (3) compounds can be interpreted figuratively. We suggest that adopting structural alignment allows us to deal with each of these problems. Structural alignment is a process whereby conceptual structures are placed into correspondence and similarities are found. The structural alignment model proposed in this thesis suggests that there are six core combination types and that an interpretation of a nounnoun compound will fall into one of these combination types. Some of these combination types are figurative and some rely on finding a context. We provide an implementation of the model, the fNCA system. The INCA system is a program where a user can find interpretations for noun-noun compounds. INCA has a knowledge base and attempts to find fixed patterns in a network representation of concepts. Depending on the type of pattern found, several types of interpretation can be generated. The performance of INCA is compared with that of a number of human subjects in a brief evaluation study. The study shows that combination types proposed by our structural alignment model to offer a good coverage of the interpretations that people generate. Finally we set out proposals for developing INCA further and outline directions for future research