Lecture 4

Cards (40)

  • Genome Size Variation
    Genome sizes vary significantly among different life forms, from small bacterial genomes to much larger eukaryotic genomes
  • Non-Coding DNA Proportion
    As genome sizes increase, the proportion of non-coding DNA also rises
  • In simpler organisms like bacteria, the majority of DNA is coding, while in more complex organisms, such as humans, a large part of the genome consists of non-coding sequences
  • Factors Contributing to Large Genome Sizes
    • Gene Duplication
    • Large Introns
    • Transposons
    • Repetitive DNA
    • Non-repetitive DNA
  • Gene Duplication
    Families of genes and pseudogenes may share regulatory sequences, resulting in coordinated control of gene expression
  • Large Introns
    Introns can be substantial in size and often contain sequences like retrotransposons
  • Transposons
    Includes elements such as Long Interspersed Nuclear Elements (LINEs), Short Interspersed Nuclear Elements (SINEs), retroviruses, and retrotransposons
  • Repetitive DNA
    Consists of simple sequence repeats and segmental duplications
  • Non-repetitive DNA
    The part of the genome that does not include repetitive sequences
  • Gene Duplication in Eukaryotes
    • Leads to families and super-families of genes within the genome
    • Genes in families can be dispersed or clustered, and the maintenance of clusters suggests functional coordination
  • Multiple Hemoglobin Genes
    • Human hemoglobin is made of different subunits, whose genes are located on different chromosomes
    • The expression of these genes changes developmentally, enabling the transition from fetal to adult forms of hemoglobin
  • Gene Size Variation in Eukaryotes
    • Gene sizes in eukaryotes can vary significantly; for instance, the human Huntingtin gene is much larger than its counterpart in the Fugu fish
    • This size variation is often due to the presence of large introns containing retrotransposons
  • The human haploid genome is approximately 3.2 x 10^9 bp in length
  • The human genome contains around 22,000 protein-coding genes and a large number of pseudogenes
  • The largest human gene is the dystrophin gene, with over 2.4 million bp
  • Highly Repetitive Elements
    Nearly half of the human genome is made up of repetitive elements, including transposons and retrotransposons
  • Functional Implications of Repetitive Elements
    Retrotransposons play a significant role in gene expression regulation, affecting chromatin structure, gene transcription, and pre-mRNA processing
  • Euchromatin
    • Euchromatin appears as a loosely packed form of DNA, which is transcriptionally active, allowing genes to be transcribed into RNA
    • When observed under electron micrographs, euchromatin isolated from an interphase nucleus appears as a quasi-regular thread, approximately 30 nm thick
    • Upon partial unpacking in low salt conditions, the chromatin's "beads on a string" structure becomes evident, a phenomenon first observed in 1974. This refers to nucleosomes connected by linker DNA
  • Nucleosome
    Each nucleosome, resembling a bead, consists of DNA wrapped around histone proteins and contains about 200 bp of DNA
  • Nucleosome Core Particle
    • Each nucleosome core particle comprises eight histone subunits: two each of H2A, H2B, H3, and H4
    • The nucleosome core particle can be dissociated to release individual histones, and high salt conditions can cause disassociation
  • Histone Characteristics
    Histones are basic proteins rich in arginine (R) and lysine (K), carrying a positive charge
  • Histone Expression and Purification
    Histones can be expressed and purified individually, for example in bacteria such as Escherichia coli, which naturally do not have histones
  • Interactions Between Histones
    Histones H3 and H4 can interact to form a dimer, and two H3-H4 dimers interact to form a tetramer. H2A-H2B dimers can then bind to this tetramer to form the full nucleosome
  • DNA and Nucleosome Formation
    A specific sequence of DNA is used to bind the histone octamer and form the nucleosome, facilitating the study of nucleosome structure and dynamics
  • In vitro Nucleosome and Chromatin Structure Analysis
    • Through in vitro reconstitution, crystal structures of nucleosomes have been obtained, providing detailed insights into the organization of DNA and histones
    • The addition of the H1 histone, or linker histone, to the "beads on a string" form of chromatin, can lead to the formation of a more compact 30 nm fiber, a step in further DNA compaction
  • Chromatin Packaging and Organization
    • DNA compaction increases from the "beads on a string" structure (11 nm fiber) to a 30 nm fiber, and further to higher-order structures like extended chromatin (300 nm), compacted chromatin (700 nm), and ultimately metaphase chromosomes (1400 nm)
    • During mitosis, DNA is highly compacted into chromosomes, which are much shorter than the extended length of DNA
    • Sperm DNA is even more densely packed than typical chromatin, primarily using protamines instead of histones
  • Chromosome Territory
    • Chromosomes occupy distinct domains within the interphase nucleus, allowing for regulated access to DNA for transcription and replication
    • The presence of nucleosomes and histones poses questions about how transcription can occur without obstruction and whether histones interfere with DNA replication
  • Histone 'Tails'
    Extend from the nucleosome and can undergo post-translational modifications
  • Histone Modifications and the 'Histone Code'
    • Acetylation of lysine residues is common and generally associated with transcriptional activation by loosening chromatin structure
    • A term describing the hypothesis that the pattern of histone modifications acts like a code in regulating gene expression
  • Chromatin Remodeling
    1. Histone-modifying complexes are recruited to chromatin by gene activator proteins
    2. Nucleosomes can be removed, repositioned, replaced, or covalently modified, affecting DNA accessibility for transcription machinery
  • Gene Silencing
    • Methylation can lead to chromatin compaction or relaxation, affecting gene expression
    • Involve histone modifications that create a more compact chromatin state, hindering transcription factor access
  • Complexity of the Histone Code
    • Histones can be modified in various ways, not only by acetylation or methylation but also by phosphorylation, ubiquitination, and other chemical modifications
    • Each modification can influence different outcomes in DNA-related processes, such as transcription, replication, and repair
  • Maintenance of Histone Modifications During Transcription
    • Histone modifications are part of epigenetic regulation, which can be maintained across cellular generations
    • The maintenance of histone marks during and after transcription is complex, as RNA polymerase II must navigate through nucleosomes
  • Transcription Through Nucleosomes
    1. As RNA polymerase II transcribes DNA, it encounters nucleosomes which may be temporarily removed or repositioned
    2. Post-translational histone marks are generally maintained or re-established after transcription to preserve the regulatory state
  • Origins of Replication
    • Controlled to ensure precise DNA replication. In yeast, these origins are known as Autonomous Replication Sequences (Ars)
  • Histone Management During Replication
    • During replication, parental histones are distributed to new strands, and new histones are incorporated
    • Histone modifications and DNA methylation are copied to maintain gene expression patterns
  • Replication Process
    1. Initiation involves the formation of a pre-replicative complex (pre-RC) at origins, which includes ORC (Origin Recognition Complex), Cdc6, Cdt1, and Mcm helicases
    2. In S phase, the pre-RC is activated, allowing the recruitment of DNA polymerases and other replication factors
    3. As DNA unwinds, parental histones are partially retained while new histones are added. NAP-1 and CAF-1 chaperone new histones to ensure proper nucleosome assembly
  • Histone Management During Replication
    • H3-H4 tetramers are likely retained and reassembled onto new DNA, while H2A-H2B dimers are displaced and replaced
    • New histones are synthesized, acetylated to maintain an open chromatin structure, and then incorporated into nucleosomes
    • After replication, new histones undergo modifications to replicate the epigenetic state of the parental chromatin, ensuring that daughter cells maintain the same gene expression patterns
  • Epigenetic Regulation
    • DNA Methylation and Histone Modification interplay to regulate gene expression, typically silencing genes through chromatin condensation
    • Both histone modifications and DNA methylation are crucial for passing on epigenetic information to daughter cells, influencing traits and susceptibility to conditions
  • Studies indicate environmental conditions (like diet and stress) experienced by parents can influence gene expression patterns in offspring through epigenetic mechanisms