Cell Biology


DNA Synthesis and Transcription

  1. Chromosomes
    • interior of the nucleus contains 23 paired chromosomes carrying 25,000 - 30,000 genes
    • each chromosome is composed of nearly equal amounts of deoxyribonucleic acid (DNA) and basic histone proteins
    • DNA component is composed of 2 deoxyribonucleotide polymers coiled into a double helix
    • the basic repeating unit of each polymer is a deoxyribose sugar with a phosphate attached at one end and a nitrogenous base attached to the other
    • the two nucleotide chains are held together by hydrogen bonds which join the purine bases adenine (A) and guanine (G) with the pyrimidine bases thymine (T) and cytosine (C)
    • base pairing is invariant: every A is paired to a T; every G is paired to a C
    • hence, the base sequence of one chain exactly specifies the base sequence of the other chain (the chains are complementary)
    • DNA must be tightly packed to fit inside the nucleus - histone proteins are instrumental in this function
    • the telomere is a specialized DNA cap at the end of the chromosome - its major function appears to be prevention of chromosomal shortening during replication

  2. DNA Replication
    1. DNA Synthesis
      • DNA is duplicated by the unwinding and use of each chain as a template for copying by complementary base pairing
      • this results in semiconservative replication - each DNA molecule is composed of one old (conserved) and one new chain
      • DNA polymerase catalyzes nucleotides into DNA
      • in addition to polymerizing DNA, DNA polymerase is also responsible for editing and proofreading, gap filling, and repair functions
      • two complete DNA sequences are formed that contain identical genetic information
      • a mistake in DNA synthesis is called a mutation, and will result in incorrect DNA sequences being copied to daughter cells
      • a mistake in a single base pair is called a point mutation, potentially leading to either a missense or nonsense mutation
      • missense mutations can result in a single amino acid being changed, which can lead to alterations in protein structure and function
      • nonsense mutations result in replacement of an amino acid with a stop codon, leading to premature termination of protein translation
      • frameshift mutations result from the addition or deletion of several amino acids, leading to the insertion of an unrelated amino acid or stop codon
      • some mutations are silent and do not affect function of the protein

    2. Cell Cycle
      • cells spend most of their time in the G0 state
      • cells stimulated to divide enter the cell cycle: G1==>S==>G2==>M
      • S phase is where DNA synthesis occurs
      • M phase (mitosis) is where the nucleus and cell divide
      • the cell cycle is tightly regulated to ensure that cells divide only when necessary
      • one of the hallmarks of cancer is the loss of cell cycle regulation

      1. Cell Cycle Control
        • progression through the cell cycle phases is governed by the sequential activation and inactivation of a family of regulatory proteins called cyclin-dependent kinases (CDKs)
        • CDK activation requires the binding of a regulatory protein (cyclin)
        • CDK activity is inhibited by CDK inhibitory proteins (CKIs)
        • the active cyclin-CDK complex phosphorylates other cell cycle regulatory proteins
        • multiple cyclin/cyclin-dependent kinase complexes exist and exhibit a cell cycle phase specificity
        • mutations that cause increased activity of the cyclin/CDK complexes are important factors for malignant transformation

        1. p53
          • transcription factor that binds to DNA, activating transcription of p21
          • p21 blocks the activity of a cyclin-dependent kinase required for progression through G1
          • this block allows time for the cell to repair any DNA damage before it is replicated
          • if the genomic damage cannot be repaired, then the progression through the cell cycle is stopped and apoptosis induced
          • functions as a tumor suppressor and is the most frequently mutated gene in human cancers
  3. RNA Transcription
    1. RNA Synthesis
      • nucleolus is the site where most cellular RNA is produced and organized
      • mRNA contains the base sequence that codes for the amino acids of the newly synthesized protein
      • uracil (U) is substituted for thymine, such that U pairs with A as its complementary base
      • RNA transcription is asymmetric because RNA polymerase selectively copies only one of the two DNA strands – the choice of strand is dependent on the gene
      • mRNA synthesis first involves a large precursor molecule that contains both protein coding sequences (exons) and intervening noncoding sequences (introns)
      • the final mRNA molecule is shortened by splicing to contain only exons

      1. Control of RNA Synthesis
        • transcription is highly regulated
        • RNA synthesis begins with binding of RNA polymerase to an upstream promoter region (TATA box)
        • there are thousands of gene regulatory proteins – transcription factors -, which bind to specific DNA sequences called regulatory elements
        • transcription factors activate or repress transcription, and different regulatory proteins are expressed in different cells
        • many human genes have more than 20 regulatory elements
        • one example of a transcription factor is the steroid hormone receptor-ligand complex

  4. RNA Translation
    • synthesis of proteins from mRNA
    • takes place on ribosomes in the cytoplasm
    • a codon, which is a triplet of three mRNA bases, encodes for one amino acid
    • each codon is recognized by a tRNA molecule, which adds the correct amino acid to the growing peptide chain
    • most amino acids are coded by more than one codon
    • the start codon is AUG; there are three stop codons
    • most proteins are modified by some combination of proteolytic cleavage, glycosylation, phosphorylation, and sulfation
    • many of the modifications occur in the Golgi apparatus

Cell-to-Cell Interactions

  • multi-celled organisms have evolved an elaborate system of cell-to-cell communication
  • extracellular information that cells process is provided in the form of various humoral or contact-mediated signals
  • these signals (ligands) exert their effects by binding with specific transmembrane receptor proteins
  • this is then transduced into a cytoplasmic signal, which initiates a second messenger cascade within the cell

  1. Chemical Signaling
    • various strategies have evolved to transport information to cells at some distance

    1. Endocrine Signaling
      • long-range signaling requires the transport of signals via the bloodstream
      • specific messenger molecules (hormones) are secreted by specialized cells which are usually organized into glands
      • endocrine signals reach every cell in the body, but only those cells that are able to bind the signal and translate its message will respond

    2. Paracrine Signaling
      • paracrine signals act over short distances
      • typical examples are cytokines, eicosanoids, and biologically active amines such as serotonin or histamine
      • do not normally enter the circulation in sufficient concentration to affect distant cells
      • various control mechanisms normally keep paracrine signals confined to their immediate region (proteinases, soluble receptors)
      • spillover of paracrine signaling molecules can have profound systemic effects (tumor necrosis factor, IL-1)

    3. Autocrine Signaling
      • autocrine signals act on the secreting cell itself, provided that the cell can receive its own signals
      • this provides feedback regulation of cell function

    4. Synaptic Signaling
      • nerve cells transmit information by electrical excitation across long distances to well defined target cells
      • target cells are contacted through synaptic signaling by the release of specific neurotransmitters

  2. Receptors
    • whether the route of signaling follows an endocrine, paracrine, autocrine, or synaptic fashion, the ligand will bind to a protein receptor that translates the signal into information that leads to a specific reaction of the target cell
    • water-soluble ligands (proteins) bind to cell surface receptors
    • lipid-soluble ligands can cross biologic membranes and bind to receptors in the cytoplasm or nucleus

    1. Intracellular Receptors
      • thyroid and steroid hormones pass directly through the plasma membrane and bind to receptor proteins inside the cell
      • resulting complex is then transported into the nucleus
      • hormone-receptor complex then binds to certain regions of the DNA, regulating transcription of various genes

    2. Cell Surface Receptors
      • binding of ligands to cell surface receptors initiates a cascade of intracellular second messenger systems that regulate cell function directly or change gene expression

      1. Tyrosine Kinase Receptors
        • translate specific signals into tyrosine kinase activity
        • controls cellular events by phosphorylation of tyrosine residues located in certain intracellular proteins
        • this causes a conformational change sufficient to initiate a specific intracellular response
        • receptor family includes epidermal growth factor (EGF), platelet-derived growth factor, transforming growth factor-α, insulin receptor
        • erb B oncogene encodes for a mutation of the receptor for EGF

      2. G Protein-linked Receptors
        • G proteins function as mediators between G protein-linked receptors and membrane-associated enzymes
        • G proteins can function as stimulators or inhibitors of signal transduction
        • binding of the ligand to its receptor changes the configuration of the receptor, which enables binding of the G protein to the receptor
        • GDP of the G protein is then replaced by GTP
        • G protein then dissociates from its receptor
        • G protein can now bind to and activate specific membrane-bound enzymes
        • oncogenes such as the ras proteins are involved in G protein-coupled signal transduction

        G-Protein Linked Receptors
    3. Intracellular Processing of Signals
      • following receptor-ligand interactions, events occurring in cell membranes must be translated into messages that can be interpreted within the cell
      • this is organized through stimulation of second messenger systems

      1. cAMP
        • ubiquitous intracellular messenger in all animal cells
        • produced by adenylate cyclase using ATP
        • activity of adenylate cyclase is regulated by G protein-coupled cell surface receptors
        • 2nd messenger function of cAMP is mediated through activation of cAMP-dependent protein kinases
        • these enzymes in turn activate other enzymes by phosphorylation

Cell Death

  • to maintain tissue homeostasis, cell proliferation must be balanced against cell death
  • cell death is responsible for removing senescent cells and cells with genetic damage beyond repair

  1. Apoptosis
    • programmed cell death is activated by two pathways, the extrinsic pathway, and the intrinsic pathway
    • in the extrinsic pathway, cell surface death receptors bind to proapoptotic ligands (TNF)
    • the intrinsic pathway is activated when intracellular sensors (p53) detect proapoptotic stimuli such as irreparable genetic damage
    • both activated pathways result in activation of caspases (cysteine aspartase proteases) that cleave proteins after aspartic acid residues
    • activated caspases set off a terminal series of events by cleaving key cellular proteins – nuclear and cytoskeletal structural proteins, DNA repair proteins
    • process results in formation of vesicles called apoptotic bodies, which are then eliminated by phagocytic cells
    • dysregulation of apoptosis can lead to cancer or autoimmune diseases

  2. Autophagy
    • characterized by massive vacuolization of the cytoplasm without chromatin condensation
    • resulting vesicles are called autophagosomes
    • autophagosomes fuse with lysosomes, leading to degradation of engulfed cytoplasmic material and organelles
    • plays an important role in protecting against infection, neurodegeneration, and tumor development
    • autophagy is controlled by at least 11 highly conserved genes
    • process is less well understood than apoptosis







References

  1. Simmons and Steed, pgs 3 - 12
  2. O’leary, 4th ed., pgs 1 – 43
  3. Sabiston, 19th ed., pgs 24 - 39
  4. Schwartz, 10th ed., pgs 443 - 453