Central Dogma, DNA Replication, and Cell Cycle Flashcards

Flow of Genetic Information

• The Central Dogma of Molecular Biology: Describes the detailed pathway and directional flow of genetic information within a biological system.     

  

• Pathways of Genetic Information Flow:

  • DNA Replication: Synthesis of identical DNA copies catalyzed by $\text{DNA Polymerase}$.

  • Transcription: Transfer of genetic information from DNA to sense RNA catalyzed by $\text{RNA Polymerase}$.

  • Translation: Decoding of sense RNA into a specific functional protein sequence catalyzed by $\text{Ribosomes}$.

  • Reverse Transcription: Synthesis of complementary DNA from a sense RNA template catalyzed by $ rối\text{Reverse Transcriptase}.\n * **RNA Replication**: Replication of sense RNA to form antisense RNA and vice-versa catalyzed by \text{RNA Dependent RNA Polymerase (RDRP)}.\n    \n\n* **Cellular Compartmentalization**:\n * Replication and Transcription occur within the cell nucleus of eukaryotic cells.\n * Processed mRNA is transported across the nuclear membrane into the cytoplasm.\n * Translation of mRNA into functional protein takes place on ribosomes located within the cytoplasm.\n\n# DNA Structure and Chromatin Organization\n\n* **Double Helix Geometry and Grooves**:\n    \n * **Major Groove**:\n * Exposes nitrogenous base sequences directly to the external environment.\n * Facilitates sequence-specific binding of regulatory proteins, including transcription factors, polymerases, and nucleases.\n * Helical dimensions: A full helical turn spans \text{3.4}\,\text{nm}$(containing approximately$\text{10}$base pairs), distance between adjacent base pairs is$\text{0.34}\,\text{nm}$, and helix diameter measures$\text{2}\,\text{nm}.\n * **Minor Groove**:\n * Narrower groove that primarily binds non-sequence-specific molecular agents, including specific antibiotics and fluorescent dyes or stains.\n\n* **Law of Complementary Base Pairing**:\n    \n * Operates on a strict lock-and-key chemical principle in DNA and RNA where a purine base selectively binds a specific pyrimidine base through hydrogen bonding.\n * **Purines**: Double-ring nitrogenous bases consisting of Adenine (\text{A}$) and Guanine ($\text{G}).\n * **Pyrimidines**: Single-ring nitrogenous bases consisting of Thymine (\text{T}$) (in DNA), Uracil ($\text{U}$) (in RNA), and Cytosine ($\text{C}).\n * **Hydrogen Bonding Specificity**:\n * Adenine (\text{A}$) pairs exclusively with Thymine ($\text{T}$) via$\text{2} hydrogen bonds.\n * Guanine (\text{G}$) pairs exclusively with Cytosine ($\text{C}$) via$\text{3} hydrogen bonds.\n * **Chargaff's Rule**: Total purines equal total pyrimidines (\text{Purines} = \text{Pyrimidines}$), such that$\text{A} + \text{G} = \text{T} + \text{C}.\n\n* **Antiparallel Strand Orientation**:\n    \n * The two complementary DNA strands run parallel to one another but in opposite chemical directions (\text{5'} \rightarrow \text{3'}$versus$\text{3'} \rightarrow \text{5'}).\n * Chemical end designations are derived from carbon numbering on the deoxyribose sugar backbone:\n * \text{5'}$end terminates with a free phosphate group attached to the$\text{5'} carbon of deoxyribose.\n * \text{3'}$end terminates with a free hydroxyl ($\text{-OH}$) group on the$\text{3'} carbon of deoxyribose.\n * Enzymes responsible for nucleic acid synthesis (replication and transcription polymerases) exhibit mandatory directionality, appending nucleotides exclusively to the free \text{3'-OH} terminus.\n\n* **Chromatin Hierarchy and Nucleosome Structure**:\n    \n * **Chromatin**: Dynamic nucleoprotein complex composed of DNA, histone proteins, and RNA that performs four core functions:\n 1. Packages genomic DNA into a small nuclear volume.\n 2. Structurally reinforces DNA to facilitate organized segregation during mitosis.\n 3. Prevents chemical and physical DNA damage.\n 4. Controls transcription and translation accessibility.\n * **Nucleosome Structure**:\n        \n * The fundamental structural unit of chromatin.\n * **Histone Octamer**: Consists of a protein core containing two copies each of core histones \text{H2A}$,$\text{H2B}$,$\text{H3}$, and$\text{H4}.\n * Exactly \text{146}\,\text{bp}$of double-stranded DNA wraps$\text{1.65} turns around the histone octamer core particle.\n * **Linker DNA**: Spans \text{54}\,\text{bp}$of DNA between adjacent nucleosomes, bound and stabilized by Histone 1 ($\text{H1}).\n * Histones are highly conserved across eukaryotic evolution.\n * Flexible N-terminal **histone tails** extend outward from the core and undergo post-translational modifications that regulate transcription and translation access.\n * **Levels of Structural Compaction**:\n 1. Short region of double-stranded DNA helix (\text{2}\,\text{nm} diameter).\n 2. "Beads-on-a-string" nucleosome chain (\text{11}\,\text{nm} diameter).\n 3. Packed nucleosome chromatin fiber (\text{30}\,\text{nm} diameter).\n 4. Extended chromosome loops (\text{300}\,\text{nm} diameter).\n 5. Condensed section of chromosome (\text{700}\,\text{nm} diameter).\n 6. Entire condensed mitotic chromosome (\text{1,400}\,\text{nm} total width across sister chromatids).\n\n# Protein-Coding Gene Structure and Transcription Mechanics\n\n* **Structure of a Protein-Coding Gene**:\n    \n * **Enhancers (Distal Control Elements)**: Regulatory DNA sequences located upstream or downstream of the promoter that bind activator proteins to stimulate transcription initiation.\n * **Proximal Control Elements**: Control elements positioned adjacent to the promoter that modulate transcription initiation efficiency.\n * **Promoter Region**: Binds \text{RNA Polymerase}$and general transcription factors; contains a conserved TATA box sequence ($\text{TATAAA}) where local unwinding of DNA occurs.\n * **Exons**: DNA coding regions that are preserved in mature mRNA and translated into protein sequences.\n * **Introns**: Non-coding DNA sequences intervening between exons that are excised during post-transcriptional RNA processing.\n * **Poly-A Signal Sequence**: Sequence (\text{AATAAA}) providing the signal for transcript cleavage and transcription termination.\n\n* **Transcription Initiation and Chromatin Remodeling**:\n    \n * Transcription initiates when the cell receives specific signaling indicating a required protein.\n * Chromatin must decondense to permit transcription factor binding and enzyme access:\n * **Decondensed Chromatin (Transcriptional Activation)**:\n * DNA demethylation catalyzed by demethylase enzymes.\n * Histone acetylation catalyzed by Histone Acetyltransferases (\text{HATs}), neutralizing positive histone charges to loosen DNA wrapping.\n * Activating histone methylation marks (e.g., \text{H3K4me3}$,$\text{H3K36me3}$) catalyzed by Histone Methyltransferases ($\text{HMTs}).\n * **Condensed Chromatin (Transcriptional Repression)**:\n * DNA methylation at cytosines catalyzed by methyltransferase enzymes.\n * Histone deacetylation catalyzed by Histone Deacetylases (\text{HDACs}).\n * Repressive histone methylation marks (e.g., \text{H3K9me3}$,$\text{H3K27me3}$) maintained by$\text{HMTs}$and Histone Demethylases ($\text{HDMs}).\n * **Assembly of Pre-Initiation Complex**:\n        \n 1. Activator proteins bind specific enhancer sequences, triggering DNA bending.\n 2. Activators interact with coactivators (\text{SWI/SNF}$chromatin remodeling complexes and$\text{HATs}) to decondense local chromatin.\n 3. Activators bind the Mediator complex, triggering assembly of \text{RNA Polymerase}$and general transcription factors ($\text{TFIID}$,$\text{TFIIB}$,$\text{TFIIF}$,$\text{TFIIE}$,$\text{TFIIH}) at the core promoter.\n\n* **Transcription Elongation**:\n    \n * Double-stranded DNA enters \text{RNA Polymerase} through the DNA entry channel and unwinds inside the enzyme.\n * \text{RNA Polymerase}$reads the template DNA strand strictly in the$\text{3'} \rightarrow \text{5'} direction.\n * The complementary \text{5'} \rightarrow \text{3'} strand is termed the coding strand because it carries the identical sequence as the synthesized pre-mRNA (except Uracil replaces Thymine).\n * Ribonucleotide triphosphates (rNTPs) enter via the rNTP entry channel and supply energy for synthesis.\n * rNTPs are incorporated into the transcript through synthesis reactions releasing pyrophosphate (\text{PP}_i).\n * Uracil (\text{U}$) base-pairs with Adenine ($\text{A}$) in place of Thymine ($\text{T}).\n * The newly synthesized pre-mRNA transcript exits through the RNA exit channel in the \text{5'} \rightarrow \text{3'} direction.\n\n* **Transcription Termination**:\n * The Poly-A Signal Sequence directs \text{RNA Polymerase} to halt transcription and release from DNA.\n * The primary transcript undergoes cleavage downstream of the polyadenylation signal.\n\n* **Post-Transcriptional RNA Processing**:\n * **5' Capping**: Addition of a \text{7-methylguanosine}$cap ($\text{m}^7\text{G}$cap) to the$\text{5'} end.\n * **3' Polyadenylation**: Enzymatic addition of a Poly-A tail comprising \text{50}$to$\text{250}$adenine nucleotides to the cleaved$\text{3'} end.\n * **Cap and Tail Functions**: Regulate mRNA export from the nucleus, prevent enzymatic degradation, and promote ribosome binding during translation initiation.\n * **Splicing and Alternative Splicing**:\n        \n * Non-coding intron sequences are excised, and coding exons are joined together.\n * Alternative Splicing allows a single gene to code for multiple unique protein isoforms by varying which exons are included in the mature mRNA.\n\n# Translation and Protein Synthesis\n\n* **Ribosomes and Transfer RNA (tRNA)**:\n    \n * **Ribosomes**:\n * Ribonucleoprotein structures composed of ribosomal RNA (rRNA) and structural proteins.\n * Designated as ribozymes because peptidyl transferase activity resides in catalytic rRNA.\n * Composed of a large subunit and a small subunit.\n * Contains three functional tRNA binding sites:\n * \text{A} site (aminoacyl site): Accepts incoming aminoacyl-tRNA.\n * \text{P} site (peptidyl site): Holds peptidyl-tRNA attached to the growing polypeptide chain.\n * \text{E} site (exit site): Releases uncharged tRNA.\n * **tRNA Structure**:\n * Folded RNA molecule conjugated to a specific amino acid via an ester bond at its \text{3'}$acceptor arm ($\text{CCA-3'} sequence).\n * Contains distinct functional loops: Acceptor arm, \text{D}$arm,$\text{T}\Psi\text{C} arm, and Anticodon arm.\n * Contains a 3-nucleotide anticodon sequence that forms complementary hydrogen bonds with mRNA codons.\n\n* **The Genetic Code**:\n    \n * Arranged in triplet codons: sequence of \text{3}$nucleotides specifies$\text{1} amino acid.\n * **Degeneracy**: Multiple distinct triplet codons can encode the exact same amino acid (e.g., UCU, UCC, UCA, and UCG all encode Serine).\n * **Start Codon**: \text{AUG} (codes for Methionine).\n * **Stop Codons**: \text{UAA}$,$\text{UAG}$, and$\text{UGA}.\n\n* **Translation Initiation Modes**:\n    \n * **Cap-Dependent Initiation**:\n 1. Small ribosomal subunit (\text{40S}$) complexes with eukaryotic initiation factors ($\text{eIF1}$,$\text{eIF1A}$,$\text{eIF3}$,$\text{eIF5}$) and a ternary complex ($\text{eIF2-GTP}$bound to$\text{Met-tRNA}_i^{\text{Met}}$) to form the$\text{43S} pre-initiation complex.\n 2. Cap-binding complex (\text{eIF4E}$,$\text{eIF4G}$,$\text{eIF4A}$,$\text{eIF4B}$) binds the$\text{5'}$$\text{m}^7\text{G} cap of mRNA.\n 3. The \text{43S}$complex scans along mRNA$\text{5'} \rightarrow \text{3'}$powered by ATP hydrolysis until locating the start codon ($\text{AUG}).\n 4. Initiation factors dissociate, and the large ribosomal subunit (\text{60S}$) joins to complete the$\text{80S}$initiation complex with$\text{Met-tRNA}$positioned in the$\text{P} site.\n * **Cap-Independent Initiation**:\n * Protein complexes bind directly to an Internal Ribosome Entry Site (IRES) near the start codon without \text{5'} cap scanning.\n * Operates under cellular stress conditions (e.g., starvation, viral infection, hypoxia) that normally suppress cap-dependent translation.\n * Mechanisms involve direct RNA binding, ITAF-mediated recruitment, or modified initiation factor combinations (\text{eIF2/5B}$,$\text{Ligatin}$,$\text{MCT-1/DENR}).\n\n* **Translation Elongation Mechanics**:\n    \n * Translation proceeds along mRNA strictly from the \text{5'}$end to the$\text{3'} end.\n * **Step 1**: Initiator tRNA carrying Methionine (\text{Met}$) occupies the$\text{P}$site. Incoming aminoacyl-tRNA bound to elongation factor$\text{eEF1}\alpha\text{-GTP}$enters the$\text{A}$site. GTP hydrolysis releases$\text{eEF1}\alpha\text{-GDP} + \text{P}_i$.

  • Step 2 (Peptide Bond Formation):         

    

    • Peptide bonds are synthesized via dehydration synthesis (condensation reaction releasing $\text{H}_2\text{O}$).

    • A peptide bond ($\text{-C(=O)-NH-}$) forms between the carboxyl group ($\text{-COO}^-$) of the amino acid in the $\text{P}$ site and the amino group ($\text{-NH}_3^+$) of the amino acid in the $\text{A}$ site.

  • Step 3 (Translocation):

    • Elongation factor $\text{eEF2}$ (driven by GTP hydrolysis) advances the ribosome one codon forward.

    • Uncharged tRNA moves from $\text{P}$ site to $\text{E}$ site and exits; peptidyl-tRNA moves from $\text{A}$ site to $\text{P}$ site.

• Translation Termination:

  • Occurs when a stop codon ($\text{UAA}$, $\text{UAG}$, or $\text{UGA}$) enters the $\text{A}$ site.

  • Eukaryotic release factor 1 ($\text{eRF1}$) binds directly to the stop codon.

  • Release factor $\text{eRF3-GTP}$ promotes ester bond hydrolysis, releasing the completed polypeptide chain.

  • Ribosomal subunits, mRNA, and uncharged tRNAs dissociate completely.

• Post-Translational Modifications (PTM):

  • Covalent structural modifications appended to cellular proteins following translation:

    • Acetylation ($\text{Ac}$): Addition of acetyl groups.

    • Methylation ($\text{Me}$): Addition of methyl groups.

    • Phosphorylation ($\text{Ph}$): Addition of phosphate groups to amino acid side chains.

    • Sulfation ($\text{Su}$): Addition of sulfate groups.

    • Glycosylation ($\text{Gy}$): Attachment of carbohydrate oligosaccharide chains.

DNA Replication Mechanics

• Overview and Origin Recognition:     

  

  • DNA replication initiates at specific AT-rich sequences called replication origins (which unwind easily due to $\text{2}$ hydrogen bonds per $\text{A=T}$ pair versus $\text{3}$ per $\text{G}\equiv\text{C}$ pair).

  • Initiator proteins separate double strands, creating replication bubbles flanked by active replication forks.

• Replication Initiation and Assembly:

  • The Pre-Replication Complex (pre-RC) assembles at origins during $\text{G}_1$ phase:

    • Origin Recognition Complex (ORC): Hexameric protein complex that recognizes origin sequences.

    • Cdc6 and Cdt1: Recruited by ORC to load helicases.

    • Minichromosome Maintenance Complex (MCM2-7): Hexameric helicase loaded onto DNA with unwinding activity.

• Replisome Architecture and Elongation:     

  

  • Active replisomes assemble at each fork containing MCM helicases, $\text{Cdc45}$, $\text{GINS}$, and specialized DNA polymerases ($\text{Pol }\alpha$, $\text{Pol }\epsilon$, $\text{Pol }\delta$).

  • Single-stranded binding protein (RPA) coats unwound single strands to prevent re-annealing.

  • Chemical Step:         

    

    • $\text{DNA Polymerase}$ catalyzes nucleophilic attack of the $\text{3'-OH}$ group on the incoming deoxyribonucleoside triphosphate (dNTP), attaching the nucleotide and releasing pyrophosphate ($\text{PP}_i$), which is hydrolyzed to \text{2P}_i$.\n * **Asymmetric Fork Polymerization**:\n        \n * \text{DNA Polymerase}$reads template DNA strictly$\text{3'} \rightarrow \text{5'}$and synthesizes new DNA$\text{5'} \rightarrow \text{3'}.\n * **Leading Strand**: Template runs \text{3'} \rightarrow \text{5'}$toward the fork; synthesized continuously by$\text{DNA Polymerase }\epsilon$.

    • Lagging Strand: Template runs $\text{5'} \rightarrow \text{3'}$ toward the fork; synthesized discontinuously away from the fork by $\text{DNA Polymerase }\delta$ in short Okazaki fragments.

    • Lagging Strand Mechanism: Template DNA loops around so polymerases read $\text{3'} \rightarrow \text{5'}$. Primase inserts short RNA primers; $\text{DNA Polymerase }\delta$ extends DNA from primers. RNA primers are removed, gaps filled, and nicks sealed by DNA ligase.

• Replication Termination and Telomere Maintenance:     

  

  • End-Replication Problem: Removal of the terminal RNA primer on the lagging strand leaves an un-replicated gap at the $\text{5'}$ end of newly synthesized DNA because polymerases require a $\text{3'-OH}$ primer. Without intervention, linear chromosomes shorten progressively with each cell division cycle.

  • Telomerase Action:         

    

    • Telomerase (a specialized reverse transcriptase carrying its own integral RNA template) binds repetitive telomeric sequences on the $\text{3'}$ overhang of the template strand.

    • Telomerase synthesizes additional repetitive DNA repeats on the template strand end, expanding it outward.

    • This extension allows Primase and $\text{DNA Polymerase}$ to complete lagging strand synthesis without deleting vital genomic coding sequences.

Genetic Mutation Categories and Effects

• Mutational Inheritance:     

  

  • Germline Mutations:

    • Occur in germ cells/gametes (sperm or egg).

    • Transmitted to offspring; present in every cell of the resulting organism.

    • Passes mutation to $\text{50}\%$ of the organism's gametes.

  • Somatic Cell Mutations:

    • Occur in non-germline somatic body cells.

    • Affects only a localized patch of tissue in the individual organism.

    • Cannot be inherited by offspring (none of the gametes carry the mutation).

• Mutational Origins:

  • Spontaneous Mutations: Result from intrinsic errors, base mispairings, or tautomeric shifts during DNA replication.

  • Induced Mutations: Caused by external environmental mutagens (e.g., radiation or chemical agents).         

    

    • Intercalating chemicals (e.g., Acridine) slip between adjacent base pairs, distorting the double helix and causing insertion or deletion errors during replication.

• Base Substitution Point Mutations:     

  

  • Silent Mutation: Base substitution alters codon sequence, but due to genetic code degeneracy, it codes for the identical amino acid (e.g., TTC $\rightarrow$ TTT; both code for Lysine). Protein structure remains unchanged.

  • Nonsense Mutation: Base substitution converts an amino acid codon into a premature stop codon (e.g., ATC $\rightarrow$ mRNA UAG = STOP). Leads to truncated, typically non-functional proteins.

  • Missense Mutation: Base substitution changes codon to specify a different amino acid.

    • Conservative Missense: Replaces amino acid with one sharing similar chemical properties (e.g., TTC $\rightarrow$ TCC; Lysine [basic] $\rightarrow$ Arginine [basic]). Functional impact is minimal.

    • Non-conservative Missense: Replaces amino acid with one having distinct chemical properties (e.g., TTC $\rightarrow$ TGC; Lysine [basic] $\rightarrow$ Threonine [polar]). Frequently disrupts folding and protein function.

• Frameshift Mutations:

  • Insertion Mutation:         

    

    • Insertion of single or multiple nucleotides (not multiples of $\text{3}$) shifts downstream codon reading frames, altering all subsequent amino acids (e.g., His-His-His $\rightarrow$ His-His-His-Thr-Ser-Ser-Ser).

  • Deletion Mutation:         

    

    • Deletion of nucleotides shifts reading frames downstream, changing amino acid translation (e.g., His-His-His $\rightarrow$ His-His-His-Leu-Ile-Ile-Ile) and generating nonfunctional proteins.

Eukaryotic Cell Cycle and Regulation

• Subphases of the Cell Cycle:     

  

  • Interphase: Comprises three subphases covering cell growth and genome duplication:

    1. $\text{G}_1$ Phase (Gap 1): Duration $\text{6--12}\,\text{hours}$. Cell active in metabolism and growth, accumulating materials, ATP, and enzymes required for DNA replication. DNA content is $\text{2n}$.

    2. $\text{G}_0$ Phase (Resting Phase): Quiescent state entered from $\text{G}_1$ by terminally differentiated or non-dividing cells. Can re-enter $\text{G}_1$ upon mitogenic signals.

    3. $\text{S}$ Phase (Synthesis Phase): Duration $\text{6--8}\,\text{hours}$. DNA replication duplicates entire genome, doubling DNA content from $\text{2n} \rightarrow \text{4n}$. Centrioles are duplicated.

    4. $\text{G}_2$ Phase (Gap 2): Duration $\text{3--4}\,\text{hours}$. Complete centriole maturation, synthesis of structural components, and production of mitotic control proteins. DNA content remains $\text{4n}$.

  • M Phase (Mitosis and Cytokinesis): Duration $\sim \text{1}\,\text{hour}$. Nuclear division (mitosis) and cytoplasmic division (cytokinesis) separate duplicated chromosomes into two daughter cells, returning DNA content from $\text{4n} \rightarrow \text{2n}$.

• Cell Cycle Control and Kinase Regulation:     

  

  • Driven by Cyclin proteins and Cyclin-Dependent Kinases (CDKs):

    • $\text{G}_1$ Phase: $\text{CDK2 / Cyclin E}$ regulates initiation of pre-replication complex assembly and centrosome duplication.

    • $\text{S}$ Phase: $\text{CDK2 / Cyclin A}$ activates origin firing and blocks re-replication.

    • $\text{G}_2\text{--}M$ Transition: $\text{CDK1 / Cyclin B}$ (Mitosis Promoting Factor / MPF) triggers entry into mitosis.

• Cell Cycle Checkpoints:     

  

  • $\text{G}_1$ Checkpoint: Blocks entrance into $\text{S}$ phase if DNA damage is detected.

  • $\text{S}$ Checkpoint: Halts DNA replication if genomic damage or stalled forks occur.

  • $\text{G}_2$ Checkpoint: Blocks entrance into $\text{M}$ phase if DNA replication is incomplete or damaged.

  • $\text{M}$ Checkpoint (Spindle Assembly Checkpoint): Blocks anaphase onset if sister chromatids are improperly assembled on the mitotic spindle.

Mitosis Stages and Cytokinesis Mechanics

• Overview of Mitosis Stages:     

  

  • Nuclear division divided into five continuous phases: Prophase, Prometaphase, Metaphase, Anaphase, and Telophase.

• Prophase and Prometaphase:     

  

  • Prophase:

    • Chromatin coils and condenses tightly into distinct, visible chromosomes consisting of two identical sister chromatids joined at centromeres.

    • Nucleolus breaks down and disappears.

    • Centrosomes migrate toward opposite cell poles, radiating aster microtubules and forming the early mitotic spindle.

  • Prometaphase:

    • Nuclear envelope breaks down into membrane fragments.

    • Spindle microtubules enter the nuclear region.

    • Kinetochore protein complexes assemble at chromosome centromeres and attach to kinetochore microtubules.

• Chromosome Compaction Structure:     

  

  • DNA double helix wraps around histones $\rightarrow$ forms chromatin chain $\rightarrow$ coils into supercoiled loops $\rightarrow$ condenses into duplicated mitotic chromosome ($\text{1,400}\,\text{nm}$) flanked by telomeres at end tips and centromere at center junction.

• Mitotic Spindle Architecture:     

  

  • Organized by two centrosome poles containing centriole pairs.

  • Three distinct microtubule sets:

    1. Astral Microtubules: Anchor spindle poles to the cell cortex.

    2. Kinetochore Microtubules: Attach directly to kinetochore protein complexes on chromatids.

    3. Polar (Non-kinetochore) Microtubules: Overlap at the cell equator to push poles apart.

• Metaphase and Spindle Assembly Checkpoint (SAC):     

  

  • Metaphase Alignment: Chromosomes align in single file along the metaphase plate (equatorial plane).

  • Mitotic Checkpoint Regulation:         

    

    • Unattached kinetochores accumulate inhibitory proteins (Mad1, Mad2, Bub1, Bub3, BubR1, CENP-E) that bind and sequester Cdc20, keeping Anaphase Promoting Complex (APC) inactive.

    • When all kinetochores are attached under bipolar tension, Mad2/Bub complexes dissociate, releasing Cdc20 to activate APC ($\text{APC}^{\text{Cdc20}}$).

    • Active APC ubiquitinates securin for degradation, activating separase enzyme to cleave cohesin rings holding sister chromatids together.

• Anaphase:     

  

  • Anaphase A: Cohesin cleavage allows sister chromatids (now individual daughter chromosomes) to separate. Kinetochore microtubules depolymerize at kinetochore ends (losing tubulin subunits), pulling chromosomes toward opposite poles powered by motor proteins.

  • Anaphase B: Polar microtubules slide past each other and elongate, increasing separation between spindle poles.

• Telophase and Cytokinesis:     

  

  • Telophase: Daughter chromosomes reach opposite poles and decondense into diffuse chromatin. Nuclear envelope re-forms around each genome set; nucleoli reappear, and spindle fibers disassemble.

  • Cytokinesis: Cytoplasm divides along the metaphase plate driven by a contractile ring composed of actin and myosin filaments, pinching the parent cell into two genetically identical daughter cells.