Projects Proposed by Faculty of Science for the 8th call
Metabolism of host glycogen by human pathogen Trichomonas vaginalis
Trichomonas vaginalis is the causal agent of the most prevalent non-viral sexually transmitted disease in humans worldwide and it affects about 170 million people annually, concentrated particularly in developing countries. Trichomonas vaginalis infection is primarily considered as a women disease, although it occurs also in men. The symptoms in women mainly include vaginal discharge, malodour, edema and colpitis, known as strawberry cervix. In addition, trichomoniasis has been associated with adverse pregnancy outcome including low birth weight, premature rupture of membranes, and preterm delivery. Accumulating number of studies report an increased risk of HIV infection associated with trichomoniasis. Trichomonosis in men is largely asymptomatic, which naturally favours the transmission of the parasite.
Glycogen in vaginal fluid is believed to be the main carbohydrate substrate for energy metabolism of T. vaginalis. Glycogen content increases in vaginal epithelial cells upon stimulation with estrogen and its level undergoes cyclic changes during the menstrual cycle, while glycogen content is low during childhood and menopause. This is consistent with female susceptibility to T. vaginalis infection during ontogenesis, as trichomoniasis appears in individuals during reproductive years, rarely in newborn infants of infected mothers. It was also observed that the amount of glycogen in vaginal liquids decreases when trichomonads are present. Moreover, relationship between glycogen content stimulated by estradiol and susceptibility to T. vaginalis infection was suggested based on experimental infections of animals. Alternatively, T. vaginalis can utilize arginine for ATP synthesis by mean of arginine dehydrolase pathways, which led to the production of polyamines.
Although mechanisms of intracellular glycogen synthesis as well as catabolism are well known, little information is available concerning the mechanisms of external glycogen acquisition and degradation by pathogens such as trichomonads. We hypothesize that free glycogen is first degraded by isoamylases producing disacharide maltose, subsequently maltose is metabolized by α-glucosidases producing glucose, which is then imported by facilitated diffusion across T. vaginalis membrane. This hypothesis is supported by identification of multiple genes coding for isoamylases and α-glucosidases in T. vaginalis genome, identification of isoamylase in the proteome of T. vaginalis surface, and by observation that glucosidase activity is exported by T. vaginalis to its environment. In this project we would like to investigate (i) mechanisms of external glycogen degradation, (ii) whether enzymes involved in glycogen degradation are exported to the cell surface, released to the cell environment or targeted to lysosomes (as observed in the case of isoamylases of Entamoeba histolytica), and (iii) to investigate expression of genes coding for isoamylases and α-glucosidases when alternative sources of energy such as arginine are present.
Methods that will be employed in the proposed project include measurements of corresponding enzymatic activities, expression of tagged recombinant enzymes in T. vaginalis to investigate the mechanisms of their transport within T. vaginalis cell to their final destinations using advance bioimaging methods (fluorescence lifetime imaging, fluorescence confocal super-resolution microscope based on STED) and proteomic analysis of cellular fractions (particularly isolated lysozomes), and investigation of changes in transcriptome of T. vaginalis grown with various sources for their energy metabolism. The knowledge gained by this study will uncover one of the key steps that are required for establishment of T. vaginalis infection: utilization of host glycogen as a main source of energy for the parasite. The unique pathways for glycogen degradation by T. vaginalis may represent interesting target for development of new therapies against this important but underestimated pathogen.
Research group: Laboratory of Molecular and Biochemical Parasitology, Department of Parasitology, Faculty of Science, Charles University in Prague.
Contact: Prof. Jan Tachezy, Ph.D.
Title of the research project:
Computational modelling of weak polyelectrolytes
¨The core of the project is the study of acid-base equilibria in solutions of weak polyelectrolytes - charged polymers whose ionization depends on external conditions, such as pH. Their behaviour is a result of complex interplay of ionization of the polymer, its conformation and external conditions, such as pH and the presence of other ions. This stimuli-responsive behaviour has promising potential applications. In the proposed project we use simulations to gain molecular-level understanding of the involved processes, which should enable better exploitation of the application potential.
The candidate will be expected to carry out coarse-grained simulations of weak polyelectrolytes in solution or polyelectrolyte networks (hydrogels) in various environments (pH, salt, other ions of complex structure). For this purpose, the group provides a specialized in-house software. Alternatively, an open-source package ESPResSo can be used. Optionally, the candidate may also participate in method and software development. The project involves international collaboration with the group of prof. C. Holm (University of Stuttgart, DE), and Dr. O. Borisov (University of Pau, FR).
Profile of an ideal candidate:
Completed PhD at the time of application, but not more than 10 years since its completion, fulfilment of other conditions prescribed by the University (required)
Good knowledge of English (FCE equivalent or better)
Background in soft matter and statistical mechanics
Experience with molecular simulation, programming and Linux OS
Contact: Dr. Peter Košovan
Department of Physical and Macromolecular Chemistry
Faculty of Science, Charles University in Prague
Hlavova 8, 128 43 Prague, Czech Republic
Title of the research project:
Genomics of speciation in nightingales
Research group: Population and evolutionary genetics
Funding of the project: Czech Science Foundation (15-10884Y). Evolution of reproductive isolation in two songbird species, The Common Nightingale and the Thrush nightingale: genomic and ecological perspective (2015-2017, PI – R. Reifová)
Understanding the mechanisms of species origin is a major goal of evolutionary biology. Until recently, studies of genetic basis of species formation has been restricted to model organisms amenable to laboratory crosses. Development of next-generation sequencing methods along with the emergence of novel population genomic approaches brings a great opportunity to study genetic underpinnings of speciation even in wild populations. This can broaden our knowledge of mechanisms of speciation and allow us to make more general conclusions about selective forces driving the origins of biodiversity.
The aim of this project is to elucidate the genetic architecture of reproductive isolation between two closely related and still hybridizing song bird species, the Common Nightingale (Luscinia megarhynchos) and the Thrush Nightingale (L. luscinia). Using novel genotyping approaches based on next generation sequencing we will genotype several hundreds of DNA samples from naturally occurring hybrid populations, already collected during our previous research. The obtained data will allow us to use two complementary approaches for studying genetic basis of reproductive isolation. First, we will use genomic cline analysis to identify candidate reproductive isolation loci with low levels of introgression relative to most of the genome. Second, we will use admixture mapping to examine genetic basis of traits that contribute to reproductive isolation. Combination of both approaches will enable us to infer the importance of different reproductive barriers in nightingale speciation, identify candidate speciation genes and elucidate the mechanisms responsible for the large effect of the Z chromosome in reproductive isolation, a hallmark of avian speciation. We are looking for a post-doc trained in bioinformatics and/or population genetics.
Supervisor: RNDr. Radka Reifová, Ph.D.
Department of Zoology, Faculty of Science, Charles University in Prague
Title of the research project:
Analysis of Src kinase activation using a novel FRET-based biosensor (Analýza aktivace kinázy Src pomocí nového FRET biosenzoru)
Research group: Laboratory of cancer cell invasion
Src kinase is a well-known proto‑oncogene that plays crucial roles in migration, mechanotransduction, mitogenic and anti-apoptotic signaling, and is often found deregulated in tumors. One of the most evident phenotypes of Src-transformed cells is the formation of invadopodia, the actin-rich protrusive structures of cancer cells that mediate extracellular matrix degradation, allowing cancer cells to invade surrounding tissue.
We have prepared a novel genetically encoded FRET‑based Src biosensor and have shown that the biosensor is sensitive to various physiological and artificial stimuli. The project will cover three aims: i) analysis of Src activation mechanisms; ii) analysis of Src activation in mechanotransduction; iii) analysis of Src activation in invadopodia.
The novel FRET-based biosensor represents a unique tool for unravelling the mechanisms of Src proto-oncogene activation and will help us to ascertain the role of Src in mechanotransduction and cancer cell invasiveness and invadopodia formation. Better understanding of Src activation could ultimately lead to increased effectiveness of anti-metastatic treatment.
The project expenses and in part (50%) the applicant salary will be covered by Czech Science Foundation grant 15-07321S (2015-2017).
Supervisor: Doc. RNDr. Daniel Rösel, PhD.