Publications by authors named "Lajos Mates"

Cervical intraepithelial neoplasia (CIN) represents a spectrum of preinvasive squamous lesions within the cervical epithelium, whose identification is a diagnostic challenge due to subtle histomorphological differences among its categories. This study explores ORF1p, a nucleic acid-binding protein derived from long interspersed nuclear element-1 (LINE-1), as a potential biomarker for enhancing CIN diagnosis. A comprehensive analysis of 143 cervical specimens, encompassing CIN I (n=20), CIN II (n=46), CIN III (n=14), invasive cancer (n=32), and nondysplastic cases (normal cervical epithelia (n=24) and atrophy (n=7) were conducted.

View Article and Find Full Text PDF

DNA transposon-based gene delivery vectors represent a promising new branch of randomly integrating vector development for gene therapy. For the side-by-side evaluation of the c and systems-the only DNA transposons currently employed in clinical trials-during therapeutic intervention, we treated the mouse model of tyrosinemia type I with liver-targeted gene delivery using both transposon vectors. For genome-wide mapping of transposon insertion sites we developed a new next-generation sequencing procedure called streptavidin-based enrichment sequencing, which allowed us to identify approximately one million integration sites for both systems.

View Article and Find Full Text PDF

Background: Understanding the contribution of gene function in distinct organ systems to the pathogenesis of human diseases in biomedical research requires modifying gene expression through the generation of gain- and loss-of-function phenotypes in model organisms, for instance, the mouse. However, methods to modify both germline and somatic genomes have important limitations that prevent easy, strong, and stable expression of transgenes. For instance, while the liver is remarkably easy to target, nucleic acids introduced to modify the genome of hepatocytes are rapidly lost, or the transgene expression they mediate becomes inhibited due to the action of effector pathways for the elimination of exogenous DNA.

View Article and Find Full Text PDF

Inefficient drug delivery across the blood-brain barrier (BBB) and into target cells in the brain hinders the treatment of neurological diseases. One strategy to increase the brain penetration of drugs is to use vesicular nanoparticles functionalized with multiple ligands of BBB transporters as vehicles. Once within the brain, however, drugs must also be able to reach their therapeutic targets in the different cell types.

View Article and Find Full Text PDF

Transposases are important tools in genome engineering, and there is considerable interest in engineering more efficient ones. Here, we seek to understand the factors determining their activity using the Sleeping Beauty transposase. Recent work suggests that protein coevolutionary information can be used to classify groups of physically connected, coevolving residues into elements called "sectors", which have proven useful for understanding the folding, allosteric interactions, and enzymatic activity of proteins.

View Article and Find Full Text PDF

Sleeping Beauty (SB) is a prominent Tc1/mariner superfamily DNA transposon that provides a popular genome engineering tool in a broad range of organisms. It is mobilized by a transposase enzyme that catalyses DNA cleavage and integration at short specific sequences at the transposon ends. To facilitate SB's applications, here we determine the crystal structure of the transposase catalytic domain and use it to model the SB transposase/transposon end/target DNA complex.

View Article and Find Full Text PDF

Here, we identify a role for the matrilin-2 (Matn2) extracellular matrix protein in controlling the early stages of myogenic differentiation. We observed Matn2 deposition around proliferating, differentiating and fusing myoblasts in culture and during muscle regeneration in vivo. Silencing of Matn2 delayed the expression of the Cdk inhibitor p21 and of the myogenic genes Nfix, MyoD and Myog, explaining the retarded cell cycle exit and myoblast differentiation.

View Article and Find Full Text PDF

The pig has emerged as an important large animal model in biomedical and pharmaceutical research. We describe a protocol for high-efficiency germline transgenesis and sustained transgene expression in pigs by using the Sleeping Beauty (SB) transposon system. The protocol is based on co-injection of a plasmid encoding the SB100X hyperactive transposase, together with a second plasmid carrying a transgene flanked by binding sites for the transposase, into the cytoplasm of porcine zygotes.

View Article and Find Full Text PDF

The laboratory rabbit (Oryctolagus cuniculus) is widely used as a model for a variety of inherited and acquired human diseases. In addition, the rabbit is the smallest livestock animal that is used to transgenically produce pharmaceutical proteins in its milk. Here we describe a protocol for high-efficiency germline transgenesis and sustained transgene expression in rabbits by using the Sleeping Beauty (SB) transposon system.

View Article and Find Full Text PDF

We describe a protocol for high-efficiency germline transgenesis and sustained transgene expression in two important biomedical models, the mouse and the rat, by using the Sleeping Beauty transposon system. The procedure is based on co-injection of synthetic mRNA encoding the SB100X hyperactive transposase, together with circular plasmid DNA carrying a transgene construct flanked by binding sites for the transposase, into the pronuclei of fertilized oocytes. Upon translation of the transposase mRNA, enzyme-mediated excision of the transgene cassettes from the injected plasmids followed by permanent genomic insertion produces stable transgenic animals.

View Article and Find Full Text PDF

Eukaryotic cells exhibit a characteristic response to hyperthermic treatment, involving morphological and cytoskeletal alterations and the induction of heat shock protein synthesis. Small GTPases of the Ras superfamily are known to serve as molecular switches which mediate responses to extracellular stimuli. We addressed here how small GTPase Rac1 integrates signals from heat stress and simultaneously induces various cellular changes in mammalian cells.

View Article and Find Full Text PDF

Background: Hepatocellular carcinoma (HCC) is the most frequent and aggressive primary tumor of the liver and it has limited treatment options.

Results: In this study, we report the in vitro and in vivo effects of two novel amino-trifluoro-phtalimide analogs, Ac-915 and Ac-2010. Both compounds bind lipid droplets and endoplasmic reticulum membrane, and interact with several proteins with chaperone functions (HSP60, HSP70, HSP90, and protein disulfide isomerase) as determined by affinity chromatography and resonant waveguide optical biosensor technology.

View Article and Find Full Text PDF

The architectural high mobility group box 1 (Hmgb1) protein acts as both a nuclear and an extracellular regulator of various biological processes, including skeletogenesis. Here we report its contribution to the evolutionarily conserved, distinctive regulation of the matrilin-1 gene (Matn1) expression in amniotes. We previously demonstrated that uniquely assembled proximal promoter elements restrict Matn1 expression to specific growth plate cartilage zones by allowing varying doses of L-Sox5/Sox6 and Nfi proteins to fine-tune their Sox9-mediated transactivation.

View Article and Find Full Text PDF

Background: Long interspersed element type one (L1) actively modifies the human genome by inserting new copies of itself. This process, termed retrotransposition, requires the formation of an L1 ribonucleoprotein (RNP) complex, which must enter the nucleus before retrotransposition can proceed. Thus, the nuclear import of L1 RNP presents an opportunity for cells to regulate L1 retrotransposition post-translationally.

View Article and Find Full Text PDF

Germline transgenesis is an important procedure for functional investigation of biological pathways, as well as for animal biotechnology. We have established a simple, nonviral protocol in three important biomedical model organisms frequently used in physiological studies. The protocol is based on the hyperactive Sleeping Beauty transposon system, SB100X, which reproducibly promoted generation of transgenic founders at frequencies of 50-64, 14-72, and 15% in mice, rats, and rabbits, respectively.

View Article and Find Full Text PDF

Background: Transposon-mediated transgenesis is an excellent method for creating stable transgenic lines and insertional mutants. In the chordate Ciona intestinalis, Minos is the only transposon that has been used as the tool for germline transformation. Adding another transposon system in this organism enables us to conduct genetic techniques which can only be realized with the use of two transposons.

View Article and Find Full Text PDF

The Gateway technology cloning system and transposon technology represent state-of-the-art laboratory techniques. Combination of these molecular tools allows rapid cloning of target genes into expression vectors. Here, we describe a novel Gateway technology-compatible transposon plasmid that combines the advantages of Gateway recombination cloning with the Sleeping Beauty (SB) transposon-mediated transgene integrations.

View Article and Find Full Text PDF

Genetic engineering can expand the utility of pigs for modeling human diseases, and for developing advanced therapeutic approaches. However, the inefficient production of transgenic pigs represents a technological bottleneck. Here, we assessed the hyperactive Sleeping Beauty (SB100X) transposon system for enzyme-catalyzed transgene integration into the embryonic porcine genome.

View Article and Find Full Text PDF

It has been previously shown that integrase-defective HIV-1-based gene vectors can serve, with moderate efficiency, as substrate for DNA transposition by a transiently expressed Sleeping Beauty (SB) transposase. Here, we describe the enhanced gene transfer properties of a HIV-1/SB hybrid vector that allows efficient DNA transposition, facilitated by the hyperactive SB100X transposase, from vector DNA intermediates in primary human cells. Potent transposase-dependent integration of genetic cargo carried by the hybrid HIV-1/SB vector (up to 160-fold above background) is reported in human cell lines as well as in primary human fibroblasts and keratinocytes.

View Article and Find Full Text PDF

DNA-based transposons are natural gene delivery vehicles. Similarly to retroviruses, these elements -integrate into the chromosomes of host cells, but their life-cycle does not involve reverse transcription and they are not infectious. Transposon-based gene delivery has several advantageous features compared to viral methods; however, its efficacy has been the bottleneck of transposon utilization.

View Article and Find Full Text PDF

Background: The transposon-based gene delivery technique is emerging as a method of choice for gene therapy. The Sleeping Beauty (SB) system has become one of the most favored methods, because of its efficiency and its random integration profile. Copy-number determination of the delivered transgene is a crucial task, but a universal method for measuring this is lacking.

View Article and Find Full Text PDF

Adult stem cells and induced pluripotent stem cells (iPS) hold great promise for regenerative medicine. The development of robust nonviral approaches for stem cell gene transfer would facilitate functional studies and potential clinical applications. We have previously generated hyperactive transposases derived from Sleeping Beauty, using an in vitro molecular evolution and selection paradigm.

View Article and Find Full Text PDF

Transposon-based gene vectors have become indispensable tools in vertebrate genetics for applications ranging from insertional mutagenesis and transgenesis in model species to gene therapy in humans. The transposon toolkit is expanding, but a careful, side-by-side characterization of the diverse transposon systems has been lacking. Here we compared the Sleeping Beauty (SB), piggyBac (PB), and Tol2 transposons with respect to overall activity, overproduction inhibition (OPI), target site selection, transgene copy number as well as long-term expression in human cells.

View Article and Find Full Text PDF

Transposable elements are DNA segments with the unique ability to move about in the genome. This inherent feature can be exploited to harness these elements as gene vectors for genome manipulation. Transposon-based genetic strategies have been established in vertebrate species over the last decade, and current progress in this field suggests that transposable elements will serve as indispensable tools.

View Article and Find Full Text PDF