DNA Replication and Repair Flashcards
What is the cell cycle?
What are the stages to mitosis and what is produced at the end of the process?
- Produces two diploid (46 chromosomes) daughter cells identical to parent cell
- Occurs in all somatic cells (non-gametes i.e., body cells)
- Chromosome duplicate using DNA replication to form two sister chromatids. This is still referred to as one chromosome because the chromatids are connected via a centromere
Phases of mitosis:
o Prophase
Involves the condensation of the chromosomes and breakdown of nucleolus and nuclear envelope
**o Metaphase **
The duplicated chromosomes line up along the middle of the cell
**o Anaphase **
Pulls the separate chromatids to polar ends of the cell
**o Telophase **
Involves the formation of new nuclear envelopes followed by the splitting of the cell in cytokinesis to form two diploid daughter cells identical to parent cell
Tell me the stages to meiosis
- Produces four haploid (23 chromosomes) cells- called gametes
- Starts with a germ cell
The phases are:
**o Prophase I **
Crossing over happens between homologous chromosomes resulting in what is known as a recombinant chromosome
Homologous means that the two chromosomes are the same size and code for the same gene
These crossing over events are what accounts for a large amount of genetic diversity between individuals
**o Metaphase I **
Line up in homologous pairs in the middle of the cell
**o Anaphase I
o Telophase I
o Prophase II
o Metaphase II
o Anaphase II
o Telophase II **
* The two cells after the first division contain 2 different sets of chromosomes
* The second division results in the separation in the sister chromatids producing 4 haploid cells each with a different set of 23 recombinant chromosomes i.e., gametes
In what stage of the cell cycle does DNA replication occur?
Synthesis stage (S-phase) of interphase before it enters cell division
How is DNA replication described and why is this term used?
- DNA replication is** semi-conservative**- each new strand contains half of the original DNA- where complimentary base pairing works
- DNA replication is a complex process that involves several enzymes and other proteins. It occurs in 3 main stages, what are they?
o** Initiation
o Elongation
o Termination **
What are the stages of initiation of DNA replication?
Initiation
- During initiation, the enzyme helicase unwinds and opens (“unzips”) the DNA helix:
o Breaks H bonds between the bases
o Exposes the DNA to enzymes and proteins involved in the replication process - DNA unwinds at the origin of replication- this is also where the formation of the new replicated DNA strands begins
- There are multiple origins of replication on the eukaryotic chromosome allowing replication to occur simultaneously across the genome
- Prokaryotic chromosomes are circular and only have one origin of replication
- As helicase unwinds the DNA helix additional proteins are needed for stability
- Topoisomerase is an enzyme that prevents the DNA double helix from overwinding and getting tangled
* Single-stranded binding proteins (SSB) bind to the separated strands to prevent them from annealing back together
What are the stages of elongation in DNA replication?
- Once DNA strand are unwound DNA primase (a RNA polymerase) builds primers using RNA nucleotides
- DNA polymerase uses these primers as a starting point then adds nucleotides complementary to parent strands at the 3’ end
- DNA polymerase can only synthesise DNA in the 5’ to 3’ direction, therefore
o One new strand is made continuously- the leading strand
o The other strand (the lagging strand) is made in small 5’ to 3’ sections known as **Okazaki fragments **
What are the stages to Termination in DNA replication?
- As synthesis proceeds, an enzyme with endonuclease activity removes the RNA primers
- The gaps between the Okazaki fragments are sealed with DNA nucleotides by an enzyme called DNA ligase
- Once replication reaches the ends of the linear chromosomes (telomeres) a new set of enzymes needs to be employed
Overview of DNA replication (bacteria diagram)
Whats the name for the end of DNA strands and what the end replication problem with this?
**Telomeres- the end replication problem **
- The end of the linear chromosomes is known as telomeres, which have repetitive sequences that do not code for a particular gene
- DNA polymerase is unable to replicate the 3’ end of eukaryotic chromosomes without a primer
- This means that during cell division the end remain uncopied- over time the end of the chromosomes gets progressively shorter as cells continue to divide until the cell reaches its Hayflick limit- a cellular age limit
o Cell age correlates with organism aging
o When they get too short they get flagged for apoptosis or cell death, and they are unable to divide anymore - The enzyme **telomerase **recognises the sequence at the end of the telomeres
o Has an inbuilt RNA template which allows extension of the parent strand in the 5’-3’direction
o Produces more repeats which will allow binding of primer
o DNA polymerase can then replicate the original sequence found at the end of the telomere and no information is lost
Tell me about telomere replication
-
Telomerase is not active in adult somatic cells
o over time the shortening of telomeres this contributes to aging - Is only active in cells that need to constantly divide- e.g., germ cells, lymphocytes, and some adult stem cells
- Can also be active in some cancer cells
DNA replication is highly accurate, but mistakes such as the wrong base added can occur
If left uncorrected these errors can have serious consequences- mutations, cancer
What are the different types of DNA repair?
Proofreading
Mismatch repair
Nucleotide excision repair
Tell me about DNA repair by proofreading?
- DNA polymerase can **“proofread” **as it synthesises new DNA
o Also, has 3’ exonuclease action which allows removal of the incorrect nucleotide
o Can then add on the correct nucleotide
Tell me about DNA repair via mismatch repair
If errors not correcting during replication:
* After synthesis of new DNA strand, specific repair enzymes recognise the mis-paired nucleotides, excise it from the DNA and replace it with the correct base
- Mechanism in eukaryotes is poorly understood
- In E.coli the adenine bases are methylates in parent DNA, but they are not in the daughter strands:
o This allows enzymes to know which strand/ base is incorrect
The human MutS enzymes involved in mismatch repair are; MLH1, MLH3, PMS1 and PMS2
Tell me about DNA repair via nucleotide excision repair
- Similar to mis-match repair but employed mostly for damaged bases
- Such as when UV exposure causes thymine dimers
o Two adjacent thymine nucleotides on the same strand that are covalently bonded together
o If they are not removed, this will lead to a mutation- this is common in individuals sensitive to sunlight and predisposed to skin cancers in early life - Enzymes cut out the segment of DNA containing the abnormal bases
- The gap is filled with complementary bases by DNA polymerase with help from DNA ligase
the enzyme involved in this is excision nuclease
