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BIOL 1090
Wright& Newmaster

BIO 1090 EXAM REVIEW Lecture 9 Lysosomes/ Microtubules I Cytoskeleton - Dynamic network of filaments and tubules that extend through the cytosol - Functions:  Structural support  Intracellular transport  Contractility and motility  Spatial organization within cell Microtubules (MT) - Largest cytoskeletal element - 2 types:  Axonemal o Organized and stable  Cytoplasmic o Dynamic o Located in cytosol - Structure:  Structural polarity is important for MT growth/shrinkage direction of movement of material along MT o Shrinkage can occur very rapidly at the ‘plus’ end (catastrophe) o Dynamic instability  Formation is controlled  Microtubule-organizing center-MTOC Central site of MT assembly) Microtubule-associated Proteins (MAPs) - Stabilize MTs or stimulate assembly - 2 classes:  Non-motor o Control MT organization in cytosol  Motor o Two types (can power intracellular transport):  Kinesin (plus end directed) and dynein (minus end directed) o Use ATP to generate force o Can move material along MT ‘track’ o Can generate sliding force between MTs Lecture 10 Microtubules II/ Intermediate Filaments Intermediate Filaments - Exclusive to multicellular animals - Provide structural support and mechanical strength - Stable - Structure:  Contains a-helical domains o Wrap around each other forming a coiled dimer  Monomers are aligned in parallel if dimers are polar  Aligned anti-parallel if filaments are non-polar Microfilaments - Polymer of protein actin - actin = action - Individual molecules: G-actin - Polymerized microfilament: F-actin - Several functions:  Maintenance of cell shape  Cell movement  Cytokinesis  Muscle contraction - F-actin microfilament:  Polar structure  Highly branched networks  Assembly: o G-actin polymerizes reversibly o Nucleation (slow):  G-actindimerstrimersshort filaments o Elongation (fast):  Monomers add to both ends  Faster at + end  Polymerization/depolymerization and structure/organization of F-actin filaments are regulated by actin-binding proteins o Filaments can be lose or tight  Myosin is a motor protein associated with it o Large family of proteins o Most move its toward + end o Divide into 2 groups:  Conventional (type II)  Unconventional (all other types)  Generate force and contribute to mobility in non-muscle cells Lecture 11 Actin Microfilaments/Nucleus I - The coordinate activity of actin-binding proteins controls microfilament formation in a lamellipodium to allow directed movement of cells Nucleus - Functions:  Storage, replication, and repair of genetic material  Transcription (mRNA, tRNA, rRNA) and splicing - Envelope:  2 parallel phospholipid bilayers  outer membrane binds ribosomes and is continuous with RER  Inner membrane: o Bears integral proteins, which connect to nuclear lamina - Structure:  Envelope: o Nuclear membrane o Nuclear pores o Nuclear lamina - Contents:  Chromatin  Nucleoplasm  Nuclear matrix  Nucleolus - Function:  Envelope (NE): o Separates nuclear content from cytoplasm  Separates transcription and translation o Selective barrier  Allows limited movement of molecules between nucleus and cytoplasm o Supported by nuclear lamina - Pores:  Gateways between cytoplasm and nucleoplasm  Inner and outer membranes fusepores  Pores contain a complex protein structure o Nuclear pore complex (NPC)  Composed of nucleoporins (NUPs)  Octagonal symmetry  Fits into the pore  Projects cytoplasm and nucleoplasm  Large, super-molecule structure  Functions:  Passive diffusion of smaller molecules  Regulated movement of large molecules  Regulated movement of proteins into the nucleus requires a nuclear localization signal (NLS) o Short stretch of positively charged amino acids within the protein - Lamina:  Thin meshwork of filamentous proteins o Lamin  Bound to inner surface of NE  Provides structural support for NE  Attachment sites for chromatin Lecture 12 Nucleus II - Cellular function is acutely dependent upon nuclear import and export (nucleocytoplasmic trafficking)  Nucleotides  Structural proteins  DNA packaging proteins o Histones  Proteins for DNA replication, repair, transcription  Proteins for RNA processing and export  Proteins for ribosome synthesis and export Nucleolus: - Ribosome biogenesis - Synthesis of RNA - rRNA processing - Assembly of subunits - tRNA synthesis Nuclear Matrix: - Network of insoluble protein fibres - Maintains shape and organization of the nucleus - Organizes chromosomes into discrete and specific regions within the nucleus Anchor machinery required for nuclear process Lecture 13 DNA and Chromosomes - Ribonucleic acid (RNA) and deoxyribonucleic acid (DNA) are polymers o Each have nucleotides comprised of:  A phosphate group  A sugar  Ribose (RNA) / Deoxyribose (DNA) (doesn’t have a hydroxyl group)  Base  Connected by phosphodiester bonds  Pyrimidines (U/C/T)  Purines (A/G) DNA - Double stranded - Strands are anti-parallel - Strands are held together by hydrogen bonds between bases and on opposing strands o Opposing strands are complementary - Hydrophobic bonds between adjacent stacked bases - Double helix is right handed - Strands are polar o 5’ end has free phosphate group o 3’ end had free hydroxyl group - Condensation o 1 level  Packaging DNA as a negative supercoil into nucleosomes  The linker region (which connects the histones) is susceptible to digestion by an endonuclease  DNA is wrapped around a nuclethome core of 8 histones proteins and anchored by a 9 o 2 level  More supercoiling  Driven by supercoiling interactions  Solenoid and zip-zag rd o 3 level  Attachment of fibre at many positions to a (non-histone_ protein scaffold  Histones removed - Chromosome ends are protected by telomeres o Functions  Resist degradation by DNases  Prevent fusion of chromosomal ends  Facilitate replication of the ends of the linear DNA - Centromeres o Provide the point of attachment of chromosomes to microtubules in the mitotic spindle o Regions I and III are conserved sequences that bind proteins involved in spindle attachment o Region II ~90 base pairs, >90% A/T o In multicellular eukaryotes are much larger and more complex Lecture 14 DNA is the Genetic Material - Type IIS o Bad o Virulent - Type IIR o Not so bad o Avirulent - The “transforming principle” - The transfer of information from DNA to protein is a two step process in all organisms o The central dogma of molecular biology o DNA with gene in it  transcription  RNA transcript  RNA processing + stability mRNA  translation  protein - RNA uses uracil instead of thymine - RNA synthesis o Involves template strand and non-template strand o Step One: RNA chain initiation  Localized unwinding o Step Two: RNA chain elongation  Genes closely spaced  Several genes can be encoded on a single RNA molecule o Step Three: RNA chain termination  Transcription terminator sequence (ρ rho- independent) Lecture 15 Transcription and Translation - Eukaryotes are more complex o The primary transcript is process and exported to the cytoplasm for translation o There is a promoter  Position the RNA polymerase for accurate initiation of transcription  TATA box o Step One: Cleavage by endonuclease o Step Two: Addition of poly(A) tail by poly(A) polymerase o Step Three: Removal of introns o Final mRNA transcript still contains coding but also non-coding sequence at both ends o Non-coding will be un-translated o UTRs; un-translated regions - Introns o Non-coding sequences located between coding sequences o Removed from the pre-mRNA and are not present in the mature mRNA o Vary in size and may be very large - Exons o Both coding and non-coding sequences o Composed of sequences that remain in the mature mRNA after splicing - In prokaryotes an RNA sequence positions the ribosome to begin translation at the beginning of a coding sequence or open reading frame o Transcription and translation occur simultaneously - In eukaryotes the RNA sequence around the AUG influences where translation begins and the ribosome scans from the 5’ end until a suitable start codon is found - Proteins are assembled on the ribosome according to the mRNA sequence (genetic code) - Codons are adjacent there is no spaces - The genetic code is non-overlapping; each nucleotide is part of one codon - The genetic code is degenerate; most amino acids are specified by more than one codon - The genetic code is ordered; amino acids with similar properties are specified by related codons - The genetic code is universal; each triplet /codon has the same meaning - Translation terminates at specific codons o Step One: enzyme binds to the UAG termination codon in the A site of the ribosome and the tRNA leaves the E site o Step Two: Release the nascent polypeptide and transfer of tRNA from the P site to the E site o Step Three: Dissociation of the mRNA/tRNA ribosome complex Lecture 16 Cell cycle and Mitosis - Each mitotic chromosome is comprised of a pair of sister chromatids o Called identical sister chromatids - Mitosis o What happens? (see PMAT)  Cellular contents (mitochondria, chloroplasts, other organelles) get divided between the daughter cells  Chromation duplicates and condenses  Centrosome duplicates  Nuclear envelope dissolves  Nucleolus dissolves  ER and Golgi are broken down  Micro tubules become reorganized via MTOC  Nuclear chromosomes must be duplicated exactly and distributed equally to the daughter cells o Cell cycle  Can take 30mins-several years  G 1hase - growth/cellular metabolism  S phase- DNA replication (chromosome duplication)  G 2hase - preparation for mitosis  M (mitosis) phase – chromosomal separation and cytokinesis  Duplicated chromosomes condense under the influence of condensing  Cells that exit the cell from G and enter the G stage are said to 1 0 be quiescent  No invariant clock that regulates cell cycle o Interphase  The time between successive mitosis (G 1 S + G )2  Chromosomes duplicate to produce sister chromatids o Prophase  Duplicated chromosomes condense  Initiation of spindle formation  Fragmentation of ER and Golgi  Nucleolus disappears  Nuclear membrane starts to break down  Spindle microtubules invade the nuclear space o Pro-metaphase  Microtubules attach to the kinetochores (which are on the outer surface of centromeres)  Chromosomes move towards the equator of the spindle o Metaphase  Duplicated chromosome migrate to the equatorial plane (metaphase plate) of the cell  Nuclear membrane breaks down o Anaphase  Sister chromatids of each duplicated chromosome move to opposite poles of the cell  Spindle poles move further apart o Telophase  Chromosomes de-condense and new nuclear membranes form  Nuclear envelope assembles around chromosomes  Golgi and ER reform  Daughter cells form cytokinesis  Membrane forms around daughter cells o Centrosome cycle  Centrioles are duplicated  Progresses with the cell cycle Lecture 17 Meiosis and Gametogenesis - Meiosis o Begins with a duplicated chrom
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