Meiosis: Cell Division and Genetic Diversity

• Meiosis, important for sexual reproduction, is when specialized cell division results in creation of haploid cells (or gametes)
• Errors in the meiosis process can lead to birth defects and and infertility in humans in the form of aneuploid gametes
• Recombination, or gene shuffling, is when homologous chromosomes redistribute genetic information, which is an important source of genetic diveristy for evolution and species diversification

Contents Overview

• Meiosis as a step in the cell cycle
• Steps in meiosis
• Genetic recombination
• Meiosis and diseases
• Recombination and gene mapping (lab)

Learning Outcomes

• Understanding of keywords: Homologs, sister chromatids, recombination, spindle checkpoint, recombination hotspots, chiasma, ploidy, inducer of meiosis transcription factor, tetrad, homologues, sister chromatids, chromosome segregation, caryotype, centimorgan/map unit, gene mapping
• Able to make draw illustrations showing the similarities and differences between mitosis and meiosis cycles
• Understanding of how to differentiate between prophase and anaphase during mitosis and meiosis
• Able to describe what makes meiosis essential to sexual reproduction
• Can discuss mechanisms of meiosis that results in genetic diversity
• Understanding of organisms that exhibit alternation of generations
• Able to create illustrations outlining the molecular principles for the retention of homologous chromosomes and/or sister chromatids during stages of mitosis and meiosis
• Able to Identify the molecular switch that commits replication of DNA to meiosis rather than mitosis.
• Able to illustrate variation profiles of the anaphase-promoting complex in mitosis and meiosis
• Able to explain the functions the spindle checkpoint oversees
• Able to draw the steps of genetic recombination
• Able to explain how meiotic defects relates to diseases, like cancer
• Able to explain the relationship between genetic distance and frequency of genetic recombination

Mitosis

• During mitosis, diploid cells replicate chromosomes during S phase and then separate sister chromatids during M phase, resulting in diploid daughter cells

Meiosis

• In meiosis, two chromosome-segregation phases take place in Meiosis I and Meiosis II. These follow one round of DNA replication in the pre-meiotic S phase
• During meiosis I, homologous chromosomes segregate to opposite poles, but resulting daughter cells are still diploid
• During meiosis II, sister chromatids segregate to opposite poles. This results in non-identical haploid gametes being formed and four haploid daughter cells

Meiosis as Part of the Cell Cycle

• Meiosis is not part of the cell cycle from a technical point of view, even though it is often regarded as such
• This is because the decision to enter the meiotic program is influenced by key regulators of the cell cycle
• Meiosis is the reductive phase, which halves the amount of chromosomes. This counteracts the doubling that happens during fertilization, making it a key part of the sexual life cycle
• There are two rounds of chromosome segregation after DNA replication
  • The first meiotic round segregates the homologues, leaving cells in a diploid state
  • The second meiotic round leads to segregation

Meiosis' Impact on Life Cycle

• Haploid organisms (some protists and fungi) are characterized by zygotic meiosis, where the zygote skips the generation of diploid cells by immediately undergoing meiosis. These also see mitosis exclusively higher in the cell cycle
• Diploid organisms (humans) are characterized by gametic meiosis, thus skipping the generation of haploid cells. These organisms form gametes directly via meiosis, and see mitosis exclusively lower in the cell cycle
• Organisms that live as haploid and diploid (plants) are charcterized by two multicellular generations, or alternation of generations. Mitosis can be found in both the upper and lower regions of the cell cycle

Ploidy Impact on Cell Physiology

• Comparison of haploid and diploid human embyronic stem cells (ESCs)
  • Haploid ESC volume is smaller than diploid ESCs
  • Haploid cells have one X chromosome that remains active, while diploid cells may inactive one of two X chromosomes.
  • Because of the above point, haploid cells see higher gene expression between autosomes and the X chromosome compared to diploid cells
  • Haploid human ESCs are not restricted simply to the undifferentiated state since they can differentiate into embryoid bodies and express specific markers for ectodermal (brain and skin), mesodermal (muscle and kidney), and endodermal tissues (liver, pancreas, lung, and intestine)

Regulatory Control of Meiosis in Yeast

• For yeast, the decision to enter meiosis takes place in the G1 phase and occurs due to external factors, like limited nutrients
• The two main regulators of meiotic initiation, IME1 and IME2, promote pre-meiotic S phase entry
• IME2 codes a meiosis-specific CDK-like protein kinase (G1-like CDKs, bottom), which in turn regulates IME1 transcription, which acts as the main transcription factor for the meiotic program
• Mitosis see one peak for G1CDKs, but IME2 (G1-like CDKs) see a second, higher peak that overlaps the two meiotic divisions
• Meiotic CDKs (blue) are responsible for directing chromosome segregation during meiosis I and are partially inactive between meiosis I and II, to prevent DNA replication
  • Activity of meiotic CDKs increases again to allow progress through meiosis II, and complete inactivation triggers exit from meiosis II in a manner similar to the drop in mitotic CDKs at anaphase

Regulatory Control of Meiosis in Mice

• Mammals use meiosis the last time the cells will replicate until fertlization
• The timing of meiotic division between sexes can vary
  • For male mice, meiotic prophase begins at over ten days and has two divisions to create haploid spermatids
  • Females have shorter prophase (~4 days), and can arrest oocytes for months/decades until the first meiotic division by ovulation
• In mammals, the transition from mitotic to meiotic cell cycle is programmed developmentally through endogenous events
• Retinoic acid (RA), or vitamin A, diffuse into germ cells and stimulates key target genes. This includes the gene Stra8 (stimulated by retinoic acid 8). Retinoic acid 8's upregulation is critical for meiosis initiation, and RA also causes the production of meiosis, a gene that is important for the mitosis to meiosis transition
• Key epigenetic modifiers used in chromatin remodeling coordinate with STRA8 and meiosis to activate downstream targets to initiate/complete meiosis

Genetic Approaches to Study Meiotic Gene Regulation

• In the Molecular Biology Era (1980s), screening mutants with meiotic defects took place to study meiotic gene regulation
• The Genomic Era (2000s) brought on transcriptone profiling of mutants
• Single-cell-based, with genome-wide epigenomic assays (2020s), brought regulatory mechanisms governing gene expression, including the pause-release regulatory control of RNA polymerase II (Pol II)

Overview of the Meiosis Extended Prophase I

• Meiosis prophase I is a multistep process that generates genetic diversity
  • At the Leptotene (thin) stage chromosomes begin to condense and vaguely become visible with a microscope
  • At the Zygotene (pairing) stage homologous chromosome pair up via synapsis and form tetrads
  • The Pachytene (large, thick) stage is when chromosomes thicken and homologous chromosomes exchange genetic material via crossover
  • The Diplotene (double ribbon) stage is when homologous chromosomes begin separation while remaining attached at crossover points, called chiasmata
  • The Diakinesis (movement through) is when chromosomes condense further and the nuclear envelope breaks down, setting he stage for the next stage

Recombination of the Meiosis Extended Prophase I

• Homologous recombination (HR) is activated by a DNA damage response to programmed double strand beaks (DSBs)
• Homologous enzymes can participate in both in DNA repair and HR
  • The first process is crucial to preventing changes to DNA to ensure stability, while the latter process contributes to genetic diversity
• Crossovers, a subset of HR events, remain in an intermediate state before first meiotic division
• Many of the genes involved in these processes are unique to meiosis

Meiotic Recombination: The Early Crossover Decision (ECD) Model

• Recombination begins with DNA double-strand breaks (DSBs)
• Following formation of of DSB, DNA is resected from the 5' end
• Afterword, 3' single-stranded DNA tails invade intact homologous DNA duplex
• This nascent 3' interaction is unstable, and is where the decision takes place to follow non-crossover (NCO; left) or crossover (CO) pathway (right).
  • In the NCO pathway, the 3' single-stranded tail begins DNA synthesis with the extended end ejected. It will then anneal with its partner. DNA synthesis continues and it is followed by ligation
  • In the CO pathway, the nascent 3’ interaction is stabilized to form a single-end invasion (SEI) intermediate.
• The other 3 is invaded/annealed to the displaced strand in the SEI intermediate DNA synthesis happens from both ends, with ligation happening afterwards to form a double Holliday junction (DHJ)
• The DHJ will then by nicked and resolved to create two recombinant particles

Frequency of Recombination and Gene Mapping

• Frequency of recombination between genes or genetic markers is used to made linkage maps (or genetic maps)
• The frequency of recombination measured in centimorgan is a reasonable estimate of chromosome length
• Recombination frequency between genes can be used to map genes onto their chromosomes
  • If one locus has a frequency of recombination of "X" times higher than another, its distance of locus to the centromere is said to be higher by "X" times compared between the centromere and other gene. -Recombination results in gametes with allele combinations never seen in generations prior -The number of crossover events during meiosis is indicative of the genome/chromosome/genetic distance. Freqeuncy of recombination is found to vary and usually takes place at 1 pair per
• Crossover is thought to be a common event occuring once per chromosome during the meiosis phase

Recombination Hotspots

• Recombination hotspots are regions with greater than normal crossover rate relative to surrounding areas
• These high recombination crossover rates are determined by unique DNA elements that help in driving evolution and promoting gene diversity
• Certain features of human recombination hotspots
  • Recombination rates higher than other area
  • Found close to telomeres and in high gene regions
  • Human genome has around 30,000+
  • Are important foci, with links to cancer and other diseases by genetic instability

Chromosome Segregation: MItotic Anaphase

• (Reminder) The alignment of chromosomes on metaphase plate (establishment and dissolution of cohesion linkages between sister chromatids) and subsequent segreation is dependent on anaphase
• Sister chromatids during anaphase are separated due to cohesins being removed through the protease separate (held inactive until anaphase due to inhibtor protein securin)
• Separase becomes activated when anaphase-promoting complex/cyclosome (APC/C) targets securin for proteolytic cleave
• Cohesins are retained on chromosome segments. This ensures sister chromatids stay together/around centeromeres

Chromosome Segregation: Meiotic Anaphase I and II

• Homologues, shown in blue and red, are linked by at least one chiasma and are liked this way until recombination is finished
• A second consideration shows kinetochores attaching to microtubules
• Afterword, cohesin rings, secured by centromeric cohesin rings, holds sister chromatids together, and ensures rings last through meiosis and sister chromatids can co-segregate
• Centromeric cohesin is taken apart prior to meiosis II to have the remaining cohesin cleaved through separate, resulting in opposite poles being segregated

Diseases and Defective Meiosis

• Mis-segregation causes progeny to undergo aneuploidy (incorrect number of chromosome present), regardless of whether eggs are fertilized or not.
• Studies suggest a recent theory points to defective meiosis is a catalyst factor for many somatic mutation. This leads to disrupt human germlines.
• Aneuploidy can cause miscarriages and inferitlity as well as diseases like down syndrome
• It can result in cancer and may accelerate aging

Genetic Recombination and Gene Mapping

• Genetic mapping of Sordaria Fimicola may result in spore patterns due to the result of the mitosis that comes out of fungi's post-meitoic phase
• Ascospores that are held with in Ascus allow the events associated with meiosis to be known depending on how they are aligned which can be linked to recombination events -Genetic recombinations lead to spore alignment through 4-4 alignment -While alignments without recombination events lead to alignments with 2-2-2-2 or 2-4-2
• Sordaria firmicola has a linear model with respect to its 2 genetic markers as well as the centromere within its own host cells that link with recombination