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M1W2 lectures > DNA replication > Flashcards

DNA replication Flashcards

1
Q

what are the 2 prinicpal functions of DNA?

A
  • template for DNA replication
  • template for RNA replication
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2
Q

where does DNA replication take place?

A
  • replication forks
  • allows both strands to be synthesized at the same time
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3
Q

what is the leading strand in DNA replication?

A

the DNA strand that is continuously synthesized in the direction of the fork (from 5’ to 3’)

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4
Q

what is the lagging strand in DNA replication?

A

the DNA strand that is synthesized discontinuously against the replication fork (from 3’ to 5’)

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5
Q

list the steps of DNA replication

A
  1. helicases unwind the parental double helix
  2. single-strand binding proteins stabilize the unwoumd parental DNA
  3. the leading strand is synthesized continuously in the 5’ to 3’ direction by DNA polymerase
  4. the lagging strand is synthesized discontinuously. primase synthesizes a short RNA primer which is extended by DNA polymerase to form an Okazaki fragment
  5. after the RNA primer is replaced by DNA (by another DNA polymerase) DNA ligase joins the okazaki fragment to the growing strand
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6
Q

what is DNA polymerase?

A

the enzyme that synthesizes DNA and is only able to synthesize DNA in the 5’ to 3’ direction
- requires a primer sequence which is synthesized by the enzyme primase

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7
Q

describe what occurs during leading strand synthesis

A
  • primase enzyme synthesizes short RNA primer using the DNA template
  • DNA polymerase can then add nucleotides and synthesize DNA in the 5’ to 3’ direction at the free 3 OH group
  • DNA polymerase then moves along the ‘old’ strand in the 5’ to 3’ direction, creating a new 3’ to 5’ strand
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8
Q

describe what occurs during lagging strand synthesis

A
  • lagging strand synthesis is discontinuous
  • primase synthesizes numerous primers which are used by DNA polymerase for DNA synthesis
  • the DNA fragments are called Okazaki fragments
  • exonuclease enzyme removes the RNA primers and DNA polymerase fills the gaps
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9
Q

describe the synthesis of both DNA strands

A
  • both strands synthesized at the same time
  • synthesis occurs 5’ to 3’
  • the leading strand is synthesized in the forward direction and the lagging strand is syntehsized in the reverse direction

end replication problem: ends of DNA are not replicated which directly leads to telomere shortening/loss

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10
Q

what are the 2 types of DNA damaging agents?

A
  • exogenous
  • endogenous
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11
Q

list some of the endogenous sources of DNA damage

A
  • errors in replication (errors introduced by DNA polymerase is scarce)
  • reactive oxygen species - ROS (byproduct of metabolism)
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12
Q

list some of the exogenous sources of DNA damage

A
  • UV (200-300nm) radiation from the sun
  • other radiation (X-ray and gamma rays)
  • mutagenic chemicals (e.g. DNA intercalating agents - can get inside double helix and cause damage)
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13
Q

how do healthy cells respond to DNA damage?

A
  • repairing DNA using the cells DNA repair mechanism
  • kill the cell via apoptosis
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14
Q

list the types of mutations that can arise if DNA damage is not repaired

A

point mutations:
- silent mutations
- missense mutations
- nonsense mutations

frameshift mutations

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15
Q

what is a point mutation?

A

substitution of one base pair for another (mismatched bases)

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16
Q

what happens when a silent mutation arises?

A

no effect on the protein
- silent mutations code for the same amino acid

17
Q

what happens when a missense mutation arises?

A

the protein function is altered or becomes non-functional
- amino acid is changed

18
Q

what happens when a nonsense mutation arises?

A

stop codon arises
- codon is changed to a stop codon

19
Q

what happens when a frameshift mutation arises?

A

shifts the reading frame in all codons following the mutation site
- occurs when one or more DNA base pairs are inserted into the gene
- changes the ‘frame’ of protein translation

20
Q

what are the effects of endogenous DNA damage?

A
  • glycosyl bond linking DNA bases with deoxyribose is labile (fragile) and causes spontaneous loss of purines or pyrimidines
  • creates and apurinic/apyrimidinic (AP) site
  • base pair deletions occur and can cause breaks when DNA replicates
21
Q

what are 3 causes of point mutations?

A

DNA replication of:
- mismatched bases
- deamination
- oxidation by reactive oxygen species (ROS)

22
Q

define mismatched base pairing and list its effect(s)

A
  • misincorporation of a nucleotide during replication (e.g. T matched to G instead of A)
  • can cause point mutations when DNA replicates
23
Q

define deamination and list its effect(s)

A
  • amino groups of nucleic acid bases are unstable (e.g. loss of amino group from C results in U)
  • can cause point mutations when DNA replicates
24
Q

define oxidation by reactive oxygen species (ROS) and list its effect(s)

A
  • reactive oxygen species (ROS) result from oxidative metabolism and ionizing radiation
  • produces hyperactive oxygen (e.g. radicals) which can modify nucleotides
  • can cause point mutations when DNA replicates - common one is G-T mutation
25
Q

list the effects of exogenous DNA damage

A
  • methylation/alkalation
  • interstrand cross-links
26
Q

define methylation/alkalation and list its effect(s)

A
  • many environmental chemicals can modify DNA bases by addition of a methyl group (methylation) or alkyl group (alkalation)
  • can cause point mutations when DNA replicates
27
Q

define interstrand crosslinks and list its effect(s)

A
  • alkylating agents (cancer drugs) can cross-link both strands by attaching abses on both strands
  • causes a block in replication, can also cause breaks
  • cross-links can also be generated by UV and ionizing radiation
28
Q

what kind of DNA damage does UV light cause?

A
  • photodamage
  • most frequent photoproducts are pyrimidine dimers (can be between T-T, C-C, or T-C)
  • causes mispairing during replication (TT pairs with CC instead of AA)
  • this causes a distortion of the double helix which inhibits replication and causes the cell to die
  • it can also induce interstrand cross-links
  • UV light is absorbed by the nucleic acid bases and the resulting influx of energy can induce chemical changes
29
Q

what kind of DNA damage does ionizing radiation cause?

A
  • single strand and double strand breaks
  • breakage of chromosomes can cause translocations, duplications (amplification), inversions, deletions
  • can also cause interstrand crosslinks which blocks DNA replication
30
Q

what is the cellular DNA damage repair mechanism?

A
  • DNA Damage Response (DDR)
  • DDR can ‘sense’ DNA lesions and activate the G1 DNA damage checkpoint via activating p53 protein
31
Q

list the steps involved in most repair pathways

A
  1. recognition
    detection of the damage
  2. incision
    cutting into the damaged DNA
  3. excision
    removal of the damaged part (exonucleases)
  4. polymerization
    gap filled by DNA polymerase
  5. religation
    joining of the gaps by DNA ligase

failure to repair DNA damage can result in gene mutations which can lead to cancer

M1W2 lectures (4 decks)

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