B2.060 DNA Replication Flashcards

1
Q

what phase of the cell cycle does DNA replication take place during?

A

S

synthesis

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2
Q

what does semi conservative mean?

A

parental strands are the template for synthesis of new DNA

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3
Q

what direction is DNA synthesized in?

A

nucleotides added to the 3’ end of growing DNA polymer

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4
Q

what are the steps of DNA synthesis

A
  1. Identify origins of replication
  2. Unwind dsDNA
  3. Initiate (primer synthesis)
  4. Elongate (fix over-winding)
  5. ligate newly synthesizes DNAs on lagging strand
  6. terminate replication
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5
Q

what is the function of the origin recognition complex?

A

ORC binds to origin DNA at late M/early G1 phase

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6
Q

what is the function of cdc6 +cdt1 in replication?

A

recruit/load six subunit MCM helicase complex onto origin during late M/ early G1 phase

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7
Q

what happens when cdc6 and cdt1 are release?

A

exit nucleus and are degraded until after mitosis

this makes sure only one replication event occurs per cell cycle (licensing replication)

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8
Q

what are the components of the pre-replicative complex?

A

ORC
cdt1
cdc5
MCM

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9
Q

what occurs in the transition from pre-replicative to replicative complex?

A

CDK phosphorylated MCM, activating helicase
MCM unwinds DNA
ORC leaves
ssDNA coated by replication protein A (RPA)

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10
Q

how are primers synthesized?

A
  1. DNA primare makes a 10-15 nt RNA

2. DNA pol alpha adds 30 nt onto RNA

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11
Q

how many primers are at the beginning of the leading strand?

A

one

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12
Q

how many primers are required on the lagging stand?

A

many for ongoing synthesis

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13
Q

what enzyme synthesizes DNA?

A

DNA pol d/e

PCNA (proliferating cell nuclear antigen) clamos the DNA pol d/e onto the DNA

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14
Q

what are okazaki fragments?

A

new DNA on lagging strand
1000 bp long
each has RNA primer
must be joined together

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15
Q

what is the over winding problem?

A

strand separation at the replication fork causes positive supercoiling of the double helix downstream

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16
Q

what enzyme helps w the over winding problem and how?

A
topisomerase 1 (DNA gyrase in bacteria)
introduces negative supercoils ahead of fork by creating a single stranded nick which allows DNA to spin
17
Q

describe the process of okazaki fragment maturation

A
  1. DNA pol d/e initiates at primer, elongates DNA to next primer
  2. DNA pol d/e displaces primer and endonuclease cuts off “flap”
  3. DNA pol d/e fills in DNA, leaves single nick
  4. DNA ligase seals the nick w AT{
18
Q

what is a nick?

A

absence of a phosphodiester bond in a DNA backbone

19
Q

what is telomerase and what is its function?

A

reverse transcriptase enzyme

helps lengthen telomeres that tend to shorten w successive replication events

20
Q

what is the Hayflick limit?

A

approx. 50 cell divisions before senescence

due to shortening of chromosomes

21
Q

which types of cells express telomerase?

A
stem cells
cancer cells (immortality)
22
Q

what are some mechanisms as using DNA replication as a drug target?

A

block/reduce synthesis of nucleotides
inhibit DNA polymerase
cross link DNA strands; no separation
incorporation of DNA analogues into growing chains

23
Q

what types of diseases can be targeted at DNA replication?

A

viral/microbial infections

neoplastic diseases

24
Q

how do drugs like acyclovir /foscarnet select for viruses and not destroy all cells?

A
  1. viral enzyme needed to phosphorylate/activate drug

2. drug blocks viral DNA polymerase specifically

25
what are the 3 checkpoints of cell cycle control?
G1-S: integrity of DNA -R: restriction point (late G1) G2-M: DNA duplication M: attachment of kinetochores to spindles
26
what 2 key proteins are involved in cell cycle control?
p53- genome integrity/replication | RB-conditions are "pro-division"
27
describe the cyclin-CDK-RB process
cyclin proteins are expressed at specific points in the cell cycle CDKs are activated when bound to cyclins phosphorylation of RB allows cell cycle to proceed past checkpoints
28
what happens to p53 under normal conditions?
p53 degrades during cell cycle | allows for progression past chekpoints
29
what happens to p53 during cell stress/DNA damage?
p53 stabilizes, halts cell cycle cells recover>transverse cell cycle damage too severe>apoptosis
30
what are the 3 classes of regulators?
cyclins CDKs CDK inhibitors
31
how are CDK inhibitors controlled?
signaling events indicating cell is not ready to divide | p53 increases translation of CDK inhibitors too
32
what phosphorylates cdc6 and cdt1?
CDKs