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MolCell Mega Class > PRIVETTE-VINNEDGE 2 > Flashcards

PRIVETTE-VINNEDGE 2 Flashcards

1
Q

What are the three phases of preparation for cell division in the cell cycle?

A

Interphase (G1, S, G2)

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2
Q

What are the growth phases of the cell cycle?

A

G1 and G2

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3
Q

What is the name of the DNA synthesis phase of the cell cycle?

A

S phase

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4
Q

What happens during the M phase of the cell cycle?

A

Mitosis (cell division)

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5
Q

How are transitions between the phases of the cell cycle regulated?

A

By cell cycle checkpoints

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6
Q

Where does DNA replication begin?

A

At distinct loci called Origins of Replication

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7
Q

What type of organisms have origins of replication with a consensus DNA sequence?

A

Prokaryotic organisms and yeasts

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8
Q

How do eukaryotic organisms identify origins of replication?

A

Based on contextual clues

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9
Q

What are three contextual cues that eukaryotic organisms use to identify origins of replication?

A

Local DNA topology; Chromatin modifications; DNA composition

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10
Q

What type of DNA sequence is usually found at origins of replication?

A

AT-rich sequence

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11
Q

What is the term for loosely packed chromatin that is open for transcription?

A

Euchromatin

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12
Q

What is the term for densely packed chromatin?

A

Heterochromatin

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13
Q

What type of chromatin is generally thought to be replicated first?

A

Euchromatin (early replication)

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14
Q

What type of chromatin is thought to be replicated later?

A

Heterochromatin (late replication)

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15
Q

What proteins bind to DNA origins of replication?

A

Origin recognition complex (ORC) proteins

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16
Q

What happens to many potential origins of replication in G1?

A

They are identified and primed, but only some “fire” and are used in S phase

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17
Q

How long does the S phase typically last?

A

8-10 hours

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18
Q

What proteins are involved in the E2F mediated transcription that occurs during the G1-S phase transition?

A

Cyclin-E and -A

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19
Q

What do Cyclin-E and -A activate?

A

CDK2

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20
Q

What does the CyclinE-CDK2 complex do to trigger the S phase?

A

It phosphorylates (inactivates) Rb

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21
Q

What roles do Cyclin E, Cyclin A, and CDK2 play in the S phase?

A

They have essential roles in initiating DNA replication

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22
Q

What is DNA origin licensing?

A

The process of identifying and preparing origins of replication for DNA synthesis

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23
Q

How many origins of replication are there per Mb of DNA?

A

25-40

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24
Q

What proteins bind to ORC during DNA origin licensing?

A

Cdc6 and Cdt1

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25
Q

What proteins are recruited after Cdc6 and Cdt1 bind?

A

MCM2-7 proteins

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26
Q

What is the result of inactivating licensing?

A

The pre-initiation complex

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27
Q

What phosphorylates Cdc6?

A

S phase CyclinA/E-CDK2 complexes

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28
Q

What protein binds to and inhibits Cdt1?

A

Geminin

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29
Q

What is the main purpose of licensing?

A

To prevent re-replication, ensuring DNA is only replicated once per cell cycle

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30
Q

What does DDK stand for?

A

Dbf4-dependent kinase

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31
Q

What does the DDK complex consist of?

A

Cdc7 and Dbf4

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32
Q

What does the DDK complex phosphorylate?

A

The Mcm complex

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33
Q

What does the DDK complex promote the loading of?

A

Cdc45 and Sld3

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34
Q

After cyclinA/E-CDK2 initiates licensing inactivation, what activates the MCM helicases?

A

GINS complex and DNA polymerase epsilon (pol ε)

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35
Q

What is the composition of the GINS complex?

A

A tetramer of Sld5, Psf1, Psf2, and Psf3

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36
Q

What is another name for the CMG complex?

A

Replicative helicase

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37
Q

What is the composition of the CMG complex?

A

Cdc45, MCM2-7, GINS

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38
Q

What is recruited to complete the replication progression complex (RPC)?

A

DNA polymerases

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39
Q

What does the formation of the replication fork allow?

A

DNA synthesis to begin

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40
Q

What makes up the replisome?

A

RPC + polymerases + PCNA + Replication Factor C

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41
Q

What is another name for Replication Factor C?

A

Clamp loader

42
Q

What does PCNA stand for?

A

Proliferating Cell Nuclear Antigen

43
Q

Describe the shape of PCNA.

A

Ring-shaped

44
Q

What does PCNA encircle?

A

DNA

45
Q

What is the function of PCNA?

A

Acts as a “sliding clamp”

46
Q

What DNA polymerase does PCNA act as scaffolding for?

A

DNA polymerase delta (pol δ)

47
Q

What is PCNA used as a marker of?

A

S phase

48
Q

In what direction are both strands of DNA copied during replication?

A

5’ to 3’

49
Q

What are the names of the two strands of DNA being copied during replication?

A

Leading strand and lagging strand

50
Q

What proteins run ahead of the helicases during replication?

A

Topoisomerases

51
Q

What do topoisomerases do to DNA?

A

Nick DNA to relieve tension from supercoiling and help unwind DNA

52
Q

What is the result of the uncoupling of polymerase and helicase during replication?

A

Exposed single-stranded DNA (ssDNA)

53
Q

What proteins coat the ssDNA when the polymerase and helicase uncouple?

A

RPA, then Rad51

54
Q

What do RPA and Rad51 initiate after coating the ssDNA?

A

Fork reversal

55
Q

How are stalled forks restarted?

A

Homologous recombination DNA repair or branch migration

56
Q

What two proteins coordinate to control DNA replication and prevent re-replication?

A

Cdt1 and Geminin

57
Q

Through what mechanism do Cdt1 and Geminin prevent re-replication?

A

DNA origin licensing

58
Q

What can happen if Cdt1 accumulates incorrectly or geminin is lost?

A

DNA replication can occur multiple times without mitosis

59
Q

What does FUCCI stand for?

A

Fluorescent Ubiquitination-based Cell Cycle Indicator

60
Q

What does FUCCI allow for?

A

Live cell imaging of the cell cycle

61
Q

What color is the Cdt1 protein fragment in FUCCI?

A

Kusabira-Orange 2 (mKO2)

62
Q

What phase of the cell cycle does the Cdt1 protein fragment represent in FUCCI?

A

G1 phase

63
Q

What color is the geminin protein fragment in FUCCI?

A

Azami-Green 1 (mAG1)

64
Q

What phases of the cell cycle does the geminin protein fragment represent in FUCCI?

A

S-G2-M phases

65
Q

Why were eGFP and RFP not suitable for FUCCI?

A

They were too stable, resulting in constant expression across the cell cycle.

66
Q

Who developed FUCCI?

A

Asako Sakaue-Sawano

67
Q

When and where was the development of FUCCI published?

A

In Cell in 2008

68
Q

Why were Cdt1 and geminin protein fragments used in FUCCI?

A

Because they are only present during specific phases of the cell cycle and are rapidly degraded, providing accurate cell cycle phase information.

69
Q

Why did the protein fragments in FUCCI have to be used together?

A

To distinguish between G1 and S/G2/M phases, as each fragment represents a different stage of the cell cycle.

70
Q

In flow cytometry, what is used to label newly synthesized DNA?

A

Nucleotide analogs of thymidine, such as BrdU and EdU

71
Q

What is the difference between BrdU and EdU detection?

A

BrdU uses antibodies for recognition, while EdU uses Click chemistry

72
Q

What fluorescent dye binds to all DNA in flow cytometry?

A

7-AAD

73
Q

What are CldU and IdU?

A

Thymidine analogs used to label newly synthesized DNA in DNA fiber assays

74
Q

How are DNA fibers spread for analysis in DNA fiber assays?

A

On tilted slides

75
Q

What method is used to label CldU and IdU in DNA fiber assays?

A

Immunofluorescence

76
Q

What can DNA fiber assays be combined with to test their impact on DNA replication?

A

Various DNA damaging agents or gene expression variants

77
Q

What can be rearranged in DNA fiber assays to test different hypotheses?

A

Order of pulses and treatments

78
Q

What are telomeres?

A

Repetitive DNA sequences at the end of chromosomes

79
Q

What happens to telomeres with each cell division?

A

They shorten due to incomplete replication of the ends of DNA (end replication problem)

80
Q

Why are telomeres difficult to replicate during S phase?

A

Repetitive sequences confuse the polymerases, they form secondary structures (t-loop), and they are heterochromatic.

81
Q

What is the t-loop?

A

A secondary structure formed by invasion of the 3’ end of the telomere into the duplex sequence with TRF1/2 proteins

82
Q

What does the t-loop protect DNA from?

A

Degradation and being treated as a DNA double-strand break

83
Q

True or False: You only want to replicate DNA once per cycle.

A

True, doing more replication will cause polyploidy (extra replicates of the chromosome(s)), or cancer

84
Q

What is the Hayflick limit?

A

The number of times a cell can divide before it stops due to telomere shortening

85
Q

What is the vertebrate telomere sequence?

A

5’-TTAGGG-3’

86
Q

What is the full name of TERT?

A

Telomerase reverse transcriptase

87
Q

What does TERT do?

A

Creates ssDNA from an RNA template

88
Q

What is another name for the telomerase RNA component?

A

TR/TERC

89
Q

True or False: PCNA can be an indicator of S phase in immunofluorescence.

A

True

90
Q

Who won the 2009 Nobel Prize for the discovery of telomerase?

A

Carol Greider, Elizabeth Blackburn, and Jack Szostak

91
Q

What does telomerase add to the chromosome?

A

DNA to the 3’ end

92
Q

What DNA polymerase converts most of the ssDNA to dsDNA after telomerase extension?

A

DNA polymerase alpha/primase

93
Q

What does TRAP stand for?

A

Telomere Repeat Amplification Protocol

94
Q

What does TRAP involve?

A

In vitro creation of ssDNA by telomerase and PCR-based detection of telomeres by filling in the ssDNA

95
Q

What is the typical appearance of TRAP reaction products?

A

A ladder of bands in 6bp increments (telomere repeat length)

96
Q

What can loss of telomere capping and function cause?

A

Substantial genome instability

97
Q

What happens to telomeres to protect the DNA when capping and function are lost?

A

They fuse

98
Q

What does persistent telomere shortening result in?

A

Hayflick limit and senescence/apoptosis

99
Q

In what percentage of tumors is telomerase upregulation found?

A

85-95%

100
Q

Can hTERT overexpression alone immortalize normal somatic cells?

A

Yes, but it does not transform them

101
Q

Give an example of a telomerase inhibitor that may be an anti-cancer drug.

A

Imetelstat

102
Q

What are cohesins?

A

Protein complexes that promote sister chromatid cohesion after DNA replication

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