OR Vývojové a buněčné biologie
Vypsané doktorské práce pro akademický rok 2024/2025

 

Projekty vypsané pro prezenční formu studia

 

Engineering of E.coli essential metabolic pathways to reduced amino acid repertoire


Mgr. Klára Hlouchová, Ph.D.
ID 277194 – Tento projekt je otevřen pro prezenční formu studia


The ultimate goal of this project will be to edit selected essential metabolic pathways of E.coli with the aim to substitute all tryptophan (Trp) residues in their proteins. This will test the in/dispensability of Trp in protein structure/function throughout different parts of the metabolism (such as cofactor biosynthesis, glycolysis and Krebs cycle).
The first part of the PhD project will focus on adaptation of recombineering methodology for genome engineering, to incorporate parallel variant substitutions and optimize co-/counter-selection strategies. An optimized protocol will be used to modify the selected proteins essential for cell survival in vivo, using degenerate recombineering oligonucleotide pools. The most viable variants will be selected by outgrowth of the bacteria and identified using next-generation sequencing methodology.
A parallel approach will involve in vitro engineering of those pathway proteins which are semi-essential or non-essential, to select the most viable Trp-less variants. The best variants will then be incorporated into the E. coli genome to assemble complete Trp-less metabolic pathways.
This project will thus provide a case-study of Trp indispensability in the proteome, using a number exemplary metabolic pathways. It will demonstrate the potential cost of Trp removal. The experimental data will be connected with structural bioinformatics analysing the structural context and consequences of Trp removal. The PhD student will collaborate with the team’s bioinformatician to develop potential rescue strategies if Trp substitution will require further compensatory mutations.
The candidate should have basic molecular biology, biochemistry and bioinformatic skills. Previous experience with genome editing will be an advantage for the project. Systematic approach to work, good communication and collaborative skills, resilience and creativity are essential qualities of the candidate.

 

 

 

Regulation of intracellular transport by interactions between intermediate filaments and
microtubules


RNDr. Zdeněk Lánský, Ph.D.
ID 262400 – Tento projekt je otevřen pro prezenční formu studia


Long-range cargo transport is essential, especially in highly elongated cells, such as neurons. The transport is typically driven by molecular motors which translocate along microtubules in a directed manner. Motors are regulated by a plethora of auxiliary factors presumably determining the cargo trajectory and final destination. Although crucial for the functioning of the cell, very little is known about this network of regulatory factors. Preliminary results suggest that intermediate filaments abundant in the neurons might play a role in the regulation of these transport processes. We will take advantage of a panel of microtubule-associated proteins that we have recently established in the lab to probe the interaction of these proteins with intermediate filaments and their effect on microtubule-related molecular motors. We will thus elucidate the regulatory roles of intermediate filaments in microtubule-based cargo transport. Candidate profile: We are looking for an enthusiastic PhD student motivated to work on cross-disciplinary projects. The candidate should hold a master´s degree in (bio)chemistry, (bio)physics, molecular/cellular biology or an equivalent field.

 

 

 

Understanding the Jagged1 role within the liver periportal mesenchyme


Mgr. Jan Mašek, Ph.D.
ID277112 – Tento projekt je otevřen pro prezenční formu studia


The mesenchymal tissue is a critical compartment of the liver, playing an integral role in liver development and injury response. The critical importance of Notch ligand Jagged1, expressed by the periportal mesenchyme, in hepatoblasts fate determination and bile duct development, is indisputable. However, unexplained detrimental effects of disrupted Jagged1 signaling on the mesenchymal tissue and hepatic arteries, point to an overlooked role of the Jagged1 role within the periportal mesenchyme. The proposed project aims to test Jagged1 effects on the embryonic and neonatal periportal mesenchyme in mice. To tackle this complex question, we developed unique experimental mouse models with disease-associated missense mutations in Jag1, that we aim to analyze using i) lineage tracing and 3D microscopy, ii) advanced transcriptomic approaches, and iii) mesenchymal in vitro culture assays. Insights into mesenchymal Jagged1-mediated signaling will enhance our understanding of liver morphogenesis and advance the development of targeted therapeutics for Jagged1-driven liver diseases.

 

 

Projekty vypsané pro kombinovanou formu studia

 

 

Regulation of growth and metabolism by the mTOR pathway


prof. David Marcelo Sabatini, M.D., Ph.D.
ID 266447 – Tento projekt je otevřen pouze pro kombinovanou formu studia


My lab has a long-standing interest in the regulation of growth and metabolism. This interest stems from our early work on the pathway anchored by mTOR protein kinase, which we now appreciate is a major regulator of growth and anabolism (mass accumulation) in eukaryotes and responds to diverse stimuli, including nutrients. Our lab identified the mTOR-containing protein complexes, mTORC1 and
mTORC2, and their biochemical and in vivo functions, as well as the complicated pathway upstream of mTORC1 that senses nutrients, including the Rag GTPases, GATOR complexes, and sensors for leucine and arginine.
Because our work revealed that lysosomes play a key role in the activation of mTORC1 by nutrients, we began to study lysosomes as well as other organelles, like mitochondria and melanosomes. We developed widely used methods for the rapid isolation and profiling of these organelles (e.g., Lyso-IP and Mito-IP), and used them to deorphan the functions of disease-associated genes. Because mTORC1 senses nutrients, we also became interested in the metabolic pathways that cells to use incorporate biomass and generate energy, particularly in cancer. We are also active in technology development and previously developed methodologies for genome-scale RNAi and CRISPR screening.

These are a few of the thesis projects available for graduate students available:


(1) Nutrient sensing by mTORC1 in vitro and in vivo. There are projects available to: identify the glucose sensor for the mTORC1 pathway; discover nutrient sensors in animals beyond mammals; understand how the known nutrient sensors (Sestrin for leucine, CASTOR for arginine, and SAMTOR for methionine) function in vivo in mice; and elucidate the biochemical function of key components of the nutrient sensing pathway, including GATOR2. These projects will use the tools of biochemistry and/or mouse mutants with specific mutations in nutrient-sensing pathway components.


(2) Lysosomes in normal physiology and disease. Our interest in mTORC1 led is to lysosomes as the activation of mTORC1 requires its translocation to the lysosomal surface. Using the Lyso-IP methodology and CRISPR screening technology there are projects available to: understand how common and rare neurodegenerative diseases impact lysosomal function; characterize and identify the contents of lysosomes in specialized cell types, like immune cells.


(3) Development of drug-like molecules for proteins of interest: In collaboration with medicinal chemists at IOCB and elsewhere, there are projects available to develop drug-like molecules that target mTOR pathway components as well lysosomal and mitochondrial proteins.
I am also open to ideas in the broader area of growth and metabolism from motivated students who are excited to forge a novel thesis project in consultation with me.


Research Group:
David. M. Sabatini, MD/PhD
Senior Group Leader
Institute of Organic Chemistry and Biochemistry
(IOCB)
Flemingovo n. 2
166 10 Praha 6
Czech Republic
david.sabatini@uochb.cas.cz

 

 

 

Regulation of metabolism and tool development


prof. David Marcelo Sabatini, M.D., Ph.D.
ID 277389 – Tento projekt je otevřen pouze pro kombinovanou formu studia


My lab has a long-standing interest in the regulation of growth and metabolism. This interest stems from our early work on the pathway anchored by mTOR protein kinase, which we now appreciate is a major regulator of growth and anabolism (mass accumulation) in eukaryotes and responds to diverse stimuli, including nutrients. Our lab identified the mTOR-containing protein complexes, mTORC1 and
mTORC2, and their biochemical and in vivo functions, as well as the complicated pathway upstream of mTORC1 that senses nutrients, including the Rag GTPases, GATOR complexes, and sensors for leucine and arginine.
Because our work revealed that lysosomes play a key role in the activation of mTORC1 by nutrients, we began to study lysosomes as well as other organelles, like mitochondria and melanosomes. We developed widely used methods for the rapid isolation and profiling of these organelles (e.g., Lyso-IP and Mito-IP), and used them to deorphan the functions of disease-associated genes. Because mTORC1 senses nutrients, we also became interested in the metabolic pathways that cells to use incorporate biomass and generate energy, particularly in cancer. We are also active in technology development and previously developed methodologies for genome-scale RNAi and CRISPR screening.
Beyond our interest in mTOR and nutrient sensing (Project 1), we have widespread interests in the regulation of metabolic pathways, particularly in mitochondria, and in developing new tools to study metabolism.


There are thesis projects for graduate students available in the following areas of interest:


(1)Nutrient transport and metabolism in mitochondria. Using the Mito-IP methodology, CRISPR screening technology, and structural biology there are projects available to de-orphan the functions and solve the structures of mitochondrial proteins of unknown functions and to understand how they connect to cytosolic metabolic pathways.
(2)Methods to study small molecule metabolism in vivo. There are projects available to develop methods to study carbohydrate and amino acid metabolism in cells in vivo in mice. We have novel ideas on how to study cell-type specific metabolism in vivo. These projects will use mass spectrometry-based metabolomics and tracing studies.
I am also open to ideas in the broader area of growth and metabolism from motivated students who are excited to forge a novel thesis project in consultation with me.