L15 cell cycle methods (CC1)

• when cells are in G0 or interphase, they can do their regular cell business. when they are undergoing the cell cycle, they cannot
• mitosis is the quickest phase of the cell cycle with it being one hour long
• G1 and G2 are gap phases where the cell rests. it has nutrition, growth factors, survival factors
• if a cell stays in G1 for too longer, it will go into G0 and differentiate into other cells. this specialized phase is called quiescence
• fission yeast has nuclear segregation (G1 and G2), while budding yeast has no G2. yeast is used for its quick duplication time (20 minutes). yeast is also used because it has a haploid genome.
• the benefits of yeast having a haploid genome means that when a single gene is deleted, it will effect the cell immediately (there is no second gene on a second chromatid). it's much easier to detect genetic mutants using yeast
• if the yeast has been mutated and but through at 37 degrees, some of the mutants will arrest in G1, S, G2 and M. mutated cells lead to much higher protein aggregations.
• genes called cdc genes (cell division cycle genes) help to regulate these cell cycles in yeast.
• general cell cycle order is G0, G1, S, G2, M
• frog eggs are often used for development studies, the version of these cells only have s phase and m phase. E3 ubiquitin ligase was separated this way.
• in mamilian cells, to measure how many cells are proliferating and how many are not is dependent on the modifications of specific nucleotides. 3H thymidine will be incorporated into DNA when cells go through S phase, allowing someone to see which cells are proliferating or not. bromodine on a ureal group with flourescent antibodies is another way to see which cells are proliferating.
• another method to see where the cell is in the phases is through a flow cytometry. this is a flow of cells that goes through fine needles, where a laser beam detects flouresence.
• we find through flow cytometry that most cells are in G1, some are in S and a medium amount are in G2 and M. if a cell has cancer, on the flow cytometry graph, there will be peaks past the G2 and M phases. if a cell has no survival or growth factors, the cell will go into apoptosis, resulting in a sub1 peak.

  
• Phases of the cell cycle:

  • S phase (synthesis phase) when DNA duplicates, occupies half of the cell-cycle time.
  • M phase (mitosis phase) which is less time in the cell-cycle time
• Mitosis involves:

  1. Chromosome condensation → duplicated DNA strands packaged into elongated chromosomes and condensed for segregation.

  2. Nuclear envelope breakdown.

  3. Replicated chromosomes, each consisting of a pair of sister chromatids, become attached to the microtubules of the mitotic spindle
• Function of the cell cycle:

  • Duplicates DNA accurately.
  • Segregates DNA copies precisely into two genetically identical daughter cells
• Gap phases of the cell cycle:

  • G1 Phase: Between M phase and S phase.
  • G2 Phase: Between S phase and mitosis.
  • Serve as periods for cell growth and allow the cell to monitor internal and external environments
• Cell cycle in yeast:

  • Yeasts are tiny, single-celled fungi with mechanisms of cell-cycle control similar to human cells.
  • Schizosaccharomyces pombe and Saccharomyces cerevisiae are yeast species used in studies
• Genetic studies in yeasts:

  • Yeasts reproduce rapidly, akin to bacteria, and have a genome size less than 1% of a mammal's.
  • Mutations in yeasts have led to the discovery of cell-cycle control mechanisms
• Cell cycle of mammalian cells:

  • In early embryos, cleavage divisions involve synchronous divisions without detectable G1 or G2 phases, each cycle comprising S and M phases lasting approximately 15 minutes each
• Studying the cell cycle in cultures:

  • Studies typically use isolated cells from tissues or tumors grown in culture dishes.
  • Immortalized cell lines are widely used in cell-cycle studies due to their unlimited source of genetically homogeneous cells
• Two main processes in the cell cycle are DNA replication and cell growth, which includes the duplication of cell content aside from the nucleus
• Chromosomes separate during mitosis, followed by cytokinesis where the cell divides into two
• Under a microscope, during interphase, little is visible as cells are in DNA replication, G1, or G2 phases
• Mitosis is the quickest phase of the cell cycle, lasting about 1 hour, involving nuclear division followed by cytoplasmic division
• The longer periods of the cell cycle are G1, S, and G2, with S phase being lengthy due to the need to duplicate a complex genome
• G1 and G2 are gap phases where the cell rests and checks for proper conditions before progressing
• G1 is a specialized phase where cells can differentiate into various types if they stay too long, known as the quiescent stage or G0
• Cell-cycle progression is regulated by intracellular and extracellular signals during G1 and G2 phases
• Cell-cycle organization and control are highly conserved during evolution, studied across various systems like yeasts, frog embryos, and mammalian cells in culture
• Yeast cells are commonly used as models due to their small genome, rapid duplication (every 20 minutes), and haploid genome, making it easier to study mutants
• Mutant cell division cycle genes in yeast can be isolated by mutating yeast with a carcinogen, observing cell cycle arrest at high temperatures, and rescuing mutants with wild type genes
• The frog egg method allows studying the cell cycle in a test tube using the large frog egg, which grows rapidly due to specialized development
• Mammalian cells in culture can be studied using methods like radioactive thymidine incorporation, BrdU staining, and flow cytometry to determine cell cycle phases and proliferation rates