Eukaryotic DNA

• Eukaryotic DNA replication is similar to replication in prokaryotes, but more complex
• Eukaryotic DNA replication shares many features with replication in bacteria

  • Double-stranded DNA unwound at replication origins
  • Replication forks are formed
  • Bidirectional synthesis creates a leading and lagging strands
  • Eukaryotic polymerases require 4 deoxyribonucleoside triphosphates, a template, and a primer
• Eukaryotic DNA replication is more complex due to

  • More DNA
  • Linear chromosomes
  • DNA complex with proteins
• Eukaryotic chromosomes contain multiple origins of replication to allow the genome to be replicated in a matter of minutes to a few hours
• Yeast genes contain 250-400 origins and are called autonomously replicating sequences (ARSs)
• These ARSs contain and 11-base-pair consensus sequence flanked by other short sequences involved in efficient initiation
• Eukaryotic origin also control timing of DNA replication
• The pre replication complex (pre-Rc) assembles at replication origins
• In early G1 phase of the cell cycle, replication origins are recognized by a 6-protein complex, the origin recognition complex (ORC) which tags the origin as the site of initiation
• 3 DNA polymerases are involved in replication of nuclear DNA. 1 involves mitochondrial DNA replication. Others are involved in repair processes.
• Alpha polymerase - RNA-DNA primer, initiation of DNA synthesis
• Delta polymerase - lagging strand synthesis, DNA repair, proofreading
• Epsilon polymerase - leading strand synthesis, proofreading
• Pol alpha, delta, and epsilon are the major forms of the enzyme involved in initiation and elongation
• Pol alpha possesses low processivity, a term that reflects the length of DNA that is synthesized by an enzyme before it dissociates from the template
• Pol alpha function in the synthesis of the RNA primers during initiation on the leading and lagging strands
• Once the primer is in place, polymerase switching occurs, Pol alpha is replaced by pols delta and epsilon for elongation
• The end of linear chromosomes are problematic during replication
• Telomeres at the ends of linear chromosomes consist of long stretches of short repeating sequences and preserve the integrity and stability of chromosomes
• Lagging strand synthesis at the end of the chromosome is a problem because once the RINA primer is removed, there is no free 3’-hydroxyl group from which to elongate
• Telomerase directs synthesis of the telomere repeat sequence to fill the gap
• Telomerase is a ribonucleoprotein with an RNA that serves as the template for the synthesis of its DNA complement - reverse transcription
• In most eukaryotic somatic cells, telomerase isn’t active
• With each successive cell division, telomeres shorten and erode, causing further cell division to stop
• Malignant cells maintain telomerase activity and are immortalized