Studium organizace a dynamiky bezmembránových buněčných kompartmentů

doktorské práce Název práce: Studium organizace a dynamiky bezmembránových buněčných kompartmentů Autor: Michaela Blažíková Ústav: Univerzita Karlova v Praze, Matematicko-fyzikální fakulta, Fyzikální ústav UK Vedoucí práce: Doc. RNDr. Petr Heřman, CSc., Univerzita Karlova v Praze, Matematicko-fyzikální fakulta, Fyzikální ústav UK Abstrakt Eukaryotické buňky obsahují množství organel a specifických tělísek. Kromě organel ohraničených membránou jako je např. buněčné jádro, mitochondrie nebo Golgiho aparát, jsou v buňkách i strukturně a funkčně rozlišené bezmembránové struktury. Tato práce se zabývá samo- organizačními procesy, tj. procesy nevyžadujícími specifické interakce, bezmembránových struktur v jádře, cytoplazmě a plasmatické membráně savčích buněk a kvasinek. Konkrétně se jedná o výzkum formace jadérek a Cajalových tělísek v savčím buněčném jádře a processing bodies (P-bodies) v cytoplasmě savčích buněk. Byla též studována organizace MCC domén v plasmatické membráně kvasinek (Membrane compartment of Can1). Bylo ukázáno, že nespecifické interakce v důsledku molekulárního crowdingu mohou být jednou z hlavních hnacích sil formování a stabilizace těchto vysoce dynamických struktur.of Doctoral Thesis Title: Study of the organization and dynamics of the membraneless cell compartments Author: Michaela Blažíková Institute: Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Supervisor: Doc. RNDr. Petr Heřman, CSc., Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Abstract Eukaryotic cells contain many organelles and specific bodies. Beside the membrane delimited organelles such as nucleus, mitochondria or Golgi apparatus there are other structurally and functionally distinct membraneless structures in the cells. In this work we studied the self-organization processes, i.e. the processes that do not require specific interactions, of membraneless structures in nuclei, cytoplasm and plasma membrane of mammalian cells and yeast. The research was focused on the formation of nucleoli and Cajal bodies in mammalian cell nulei and processing bodies (P- bodies) in the cytoplasm of mammalian cells. The organization of MCC domains in the yeast plasma membrane (Membrane compartment of Can1) was studied as well. It was shown that nonspecific interactions as the result of macromolecular crowding could be one of the main driving forces in formation and stabilization of these...Fyzikální ústav UKInstitute of Physics of Charles UniversityFaculty of Mathematics and PhysicsMatematicko-fyzikální fakult

Cationic Porphyrins as Transport Agents for Antisense Oligonucleotides

Fyzikální ústav UKInstitute of Physics of Charles UniversityMatematicko-fyzikální fakultaFaculty of Mathematics and Physic

Studium organizace a dynamiky bezmembránových buněčných kompartmentů

doktorské práce Název práce: Studium organizace a dynamiky bezmembránových buněčných kompartmentů Autor: Michaela Blažíková Ústav: Univerzita Karlova v Praze, Matematicko-fyzikální fakulta, Fyzikální ústav UK Vedoucí práce: Doc. RNDr. Petr Heřman, CSc., Univerzita Karlova v Praze, Matematicko-fyzikální fakulta, Fyzikální ústav UK Abstrakt Eukaryotické buňky obsahují množství organel a specifických tělísek. Kromě organel ohraničených membránou jako je např. buněčné jádro, mitochondrie nebo Golgiho aparát, jsou v buňkách i strukturně a funkčně rozlišené bezmembránové struktury. Tato práce se zabývá samo- organizačními procesy, tj. procesy nevyžadujícími specifické interakce, bezmembránových struktur v jádře, cytoplazmě a plasmatické membráně savčích buněk a kvasinek. Konkrétně se jedná o výzkum formace jadérek a Cajalových tělísek v savčím buněčném jádře a processing bodies (P-bodies) v cytoplasmě savčích buněk. Byla též studována organizace MCC domén v plasmatické membráně kvasinek (Membrane compartment of Can1). Bylo ukázáno, že nespecifické interakce v důsledku molekulárního crowdingu mohou být jednou z hlavních hnacích sil formování a stabilizace těchto vysoce dynamických struktur.of Doctoral Thesis Title: Study of the organization and dynamics of the membraneless cell compartments Author: Michaela Blažíková Institute: Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Supervisor: Doc. RNDr. Petr Heřman, CSc., Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Abstract Eukaryotic cells contain many organelles and specific bodies. Beside the membrane delimited organelles such as nucleus, mitochondria or Golgi apparatus there are other structurally and functionally distinct membraneless structures in the cells. In this work we studied the self-organization processes, i.e. the processes that do not require specific interactions, of membraneless structures in nuclei, cytoplasm and plasma membrane of mammalian cells and yeast. The research was focused on the formation of nucleoli and Cajal bodies in mammalian cell nulei and processing bodies (P- bodies) in the cytoplasm of mammalian cells. The organization of MCC domains in the yeast plasma membrane (Membrane compartment of Can1) was studied as well. It was shown that nonspecific interactions as the result of macromolecular crowding could be one of the main driving forces in formation and stabilization of these...Institute of Physics of Charles UniversityFyzikální ústav UKFaculty of Mathematics and PhysicsMatematicko-fyzikální fakult

Modeling of nucleolar self-assembly

Mammalian cell nucleoli disassemble at the beginning of mitosis and reassemble again during telophase and the early G1. Dynamics of this process was studied using a model based on entropy driven self-assembly of pre-ribosomal particles generated in a single biosynthetic source. Monte Carlo simulations revealed that such model can explain formation of a large aggregate, a nucleolus, in the vicinity of the source. We examined influence of nucleoplasm properties on dynamics of the aggregate formation

Study of the organization and dynamics of the membraneless cell compartments

of Doctoral Thesis Title: Study of the organization and dynamics of the membraneless cell compartments Author: Michaela Blažíková Institute: Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Supervisor: Doc. RNDr. Petr Heřman, CSc., Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Abstract Eukaryotic cells contain many organelles and specific bodies. Beside the membrane delimited organelles such as nucleus, mitochondria or Golgi apparatus there are other structurally and functionally distinct membraneless structures in the cells. In this work we studied the self-organization processes, i.e. the processes that do not require specific interactions, of membraneless structures in nuclei, cytoplasm and plasma membrane of mammalian cells and yeast. The research was focused on the formation of nucleoli and Cajal bodies in mammalian cell nulei and processing bodies (P- bodies) in the cytoplasm of mammalian cells. The organization of MCC domains in the yeast plasma membrane (Membrane compartment of Can1) was studied as well. It was shown that nonspecific interactions as the result of macromolecular crowding could be one of the main driving forces in formation and stabilization of these..

Cationic Porphyrins as Transport Agents for Antisense Oligonucleotides

Fyzikální ústav UKInstitute of Physics of Charles UniversityMatematicko-fyzikální fakultaFaculty of Mathematics and Physic

Study of the organization and dynamics of the membraneless cell compartments

of Doctoral Thesis Title: Study of the organization and dynamics of the membraneless cell compartments Author: Michaela Blažíková Institute: Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Supervisor: Doc. RNDr. Petr Heřman, CSc., Charles University in Prague, Faculty of Mathematics and Physics, Institute of Physics of Charles University Abstract Eukaryotic cells contain many organelles and specific bodies. Beside the membrane delimited organelles such as nucleus, mitochondria or Golgi apparatus there are other structurally and functionally distinct membraneless structures in the cells. In this work we studied the self-organization processes, i.e. the processes that do not require specific interactions, of membraneless structures in nuclei, cytoplasm and plasma membrane of mammalian cells and yeast. The research was focused on the formation of nucleoli and Cajal bodies in mammalian cell nulei and processing bodies (P- bodies) in the cytoplasm of mammalian cells. The organization of MCC domains in the yeast plasma membrane (Membrane compartment of Can1) was studied as well. It was shown that nonspecific interactions as the result of macromolecular crowding could be one of the main driving forces in formation and stabilization of these..

The F-Actin-Binding MPRIP Forms Phase-Separated Condensates and Associates with PI(4,5)P2 and Active RNA Polymerase II in the Cell Nucleus

Here, we provide evidence for the presence of Myosin phosphatase rho-interacting protein (MPRIP), an F-actin-binding protein, in the cell nucleus. The MPRIP protein binds to Phosphatidylinositol 4,5-bisphosphate (PIP2) and localizes to the nuclear speckles and nuclear lipid islets which are known to be involved in transcription. We identified MPRIP as a component of RNA Polymerase II/Nuclear Myosin 1 complex and showed that MPRIP forms phase-separated condensates which are able to bind nuclear F-actin fibers. Notably, the fibrous MPRIP preserves its liquid-like properties and reforms the spherical shaped condensates when F-actin is disassembled. Moreover, we show that the phase separation of MPRIP is driven by its long intrinsically disordered region at the C-terminus. We propose that the PIP2/MPRIP association might contribute to the regulation of RNAPII transcription via phase separation and nuclear actin polymerization

Spliceosomal Small Nuclear Ribonucleoprotein Particles Repeatedly Cycle through Cajal Bodies

The Cajal body (CB) is a nuclear structure closely associated with import and biogenesis of small nuclear ribonucleoprotein particles (snRNPs). Here, we tested whether CBs also contain mature snRNPs and whether CB integrity depends on the ongoing snRNP splicing cycle. Sm proteins tagged with photoactivatable and color-maturing variants of fluorescent proteins were used to monitor snRNP behavior in living cells over time; mature snRNPs accumulated in CBs, traveled from one CB to another, and they were not preferentially replaced by newly imported snRNPs. To test whether CB integrity depends on the snRNP splicing cycle, two human orthologues of yeast proteins involved in distinct steps in spliceosome disassembly after splicing, hPrp22 and hNtr1, were depleted by small interfering RNA treatment. Surprisingly, depletion of either protein led to the accumulation of U4/U6 snRNPs in CBs, suggesting that reassembly of the U4/U6·U5 tri-snRNP was delayed. Accordingly, a relative decrease in U5 snRNPs compared with U4/U6 snRNPs was observed in CBs, as well as in nuclear extracts of treated cells. Together, the data show that particular phases of the spliceosome cycle are compartmentalized in living cells, with reassembly of the tri-snRNP occurring in CBs