Primary cilia are hair-like sensory organelles protruding from the surface of most human cells. As cilia are dynamic, several aspects of their biology can only be revealed by real-time analysis in living cells. Here we describe the generation of primary cilia reporter cell lines. Furthermore, we provide a detailed protocol of how to use the reporter cell lines for live-cell imaging microscopy analysis of primary cilia to study their growth as well as intraciliary transport. For complete details on the use and execution of this protocol, please refer to Bernatik et al. (2020) and Pejskova et al. (2020).
- MeSH
- buněčné linie MeSH
- cilie * metabolismus MeSH
- lidé MeSH
- mikroskopie metody MeSH
- počítačové zpracování obrazu * metody MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Nutrient sensing and metabolic reprogramming are crucial for metazoan cell aging and tumor growth. Here, we identify metabolic and regulatory parallels between a layered, multicellular yeast colony and a tumor-affected organism. During development, a yeast colony stratifies into U and L cells occupying the upper and lower colony regions, respectively. U cells activate a unique metabolism controlled by the glutamine-induced TOR pathway, amino acid-sensing systems (SPS and Gcn4p) and signaling from mitochondria with lowered respiration. These systems jointly modulate U cell physiology, which adapts to nutrient limitations and utilize the nutrients released from L cells. Stress-resistant U cells share metabolic pathways and other similar characteristics with tumor cells, including the ability to proliferate. L cells behave similarly to stressed and starving cells, which activate degradative mechanisms to provide nutrients to U cells. Our data suggest a nutrient flow between both cell types, resembling the Cori cycle and glutamine-NH(4)(+) shuttle between tumor and healthy metazoan cells.
- MeSH
- aminokyseliny metabolismus MeSH
- autofagie MeSH
- biologické modely MeSH
- buněčné dělení MeSH
- fyziologický stres MeSH
- genový knockout MeSH
- geny hub MeSH
- kvartérní amoniové sloučeniny metabolismus MeSH
- lidé MeSH
- metabolické sítě a dráhy MeSH
- nádory metabolismus patologie MeSH
- Saccharomyces cerevisiae - proteiny genetika MeSH
- Saccharomyces cerevisiae cytologie genetika růst a vývoj metabolismus MeSH
- signální transdukce MeSH
- spotřeba kyslíku MeSH
- transkriptom MeSH
- transportní systémy aminokyselin genetika MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Much like other microorganisms, wild yeasts preferentially form surface-associated communities, such as biofilms and colonies, that are well protected against hostile environments and, when growing as pathogens, against the host immune system. However, the molecular mechanisms underlying the spatiotemporal development and environmental resistance of biofilms and colonies remain largely unknown. In this paper, we show that a biofilm yeast colony is a finely tuned, complex multicellular organism in which specialized cells jointly execute multiple protection strategies. These include a Pdr1p-regulated mechanism whereby multidrug resistance transporters Pdr5p and Snq2p expel external compounds solely within the surface cell layers as well as developmentally regulated production by internal cells of a selectively permeable extracellular matrix. The two mechanisms act in concert during colony development, allowing growth of new cell generations in a well-protected internal cavity of the colony. Colony architecture is strengthened by intercellular fiber connections.
- MeSH
- ABC transportéry genetika metabolismus MeSH
- biofilmy růst a vývoj MeSH
- biologické modely MeSH
- delece genu MeSH
- DNA vazebné proteiny genetika metabolismus MeSH
- extracelulární matrix fyziologie MeSH
- galaktokinasa genetika metabolismus MeSH
- galaktosa metabolismus MeSH
- hydroxymethylglutaryl-CoA-reduktasy genetika metabolismus MeSH
- měď metabolismus MeSH
- membránové glykoproteiny genetika metabolismus MeSH
- metalothionein genetika metabolismus MeSH
- oxaziny metabolismus MeSH
- permeabilita MeSH
- profiliny genetika MeSH
- proteiny buněčného cyklu genetika MeSH
- proteiny spojené s mnohočetnou rezistencí k lékům genetika metabolismus MeSH
- rekombinantní fúzní proteiny genetika metabolismus MeSH
- Saccharomyces cerevisiae - proteiny genetika metabolismus MeSH
- Saccharomyces cerevisiae cytologie růst a vývoj metabolismus MeSH
- transkripční faktory genetika metabolismus MeSH
- zelené fluorescenční proteiny genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- arteterapie metody MeSH
- duševně nemocní MeSH
- interprofesionální vztahy MeSH
- komunikace MeSH
- lidé MeSH
- mezioborová komunikace MeSH
- neverbální komunikace MeSH
- pacientův souhlas se zdravotní péčí MeSH
- psychiatrie MeSH
- psychologie aplikovaná MeSH
- psychologie MeSH
- rehabilitační centra MeSH
- spokojenost pacientů MeSH
- spolupráce organizací a občanů MeSH
- ústavy pro duševně nemocné MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- rozhovory MeSH
