DNA Repair and Oncogene- Induced Replication Stress

Question 1

What is neoplasia?

Answer 1

Tissue composed of cells with the ability to grow beyond their normal confines

Question 2

What is genomic instability linked to?

Answer 2

DNA damage

Question 3

What is gammaH2AX?

Answer 3

A phosphorylated form of histones which is generated when DNA damage occurs

Question 4

What is ATM?

Answer 4

A DNA damage response kinase which signals in the presence of DNA breaks

Question 5

What is Chk2?

Answer 5

A DNA damage response kinase which signals in response to DNA damage

Question 6

What is p53?

Answer 6

A tumour suppressor which activates a DNA damage induced checkpoint, promoting apoptosis and senescence

Question 7

What is increased DNA damage correlated with?

Answer 7

More 53BP1 foci which is correlated with increased p53 mutations

Question 8

What are the stages of cancer progression?

Answer 8

Normal -> Hyperplastic -> Dysplastic -> Neoplastic -> Metastatic

Question 9

What is hyperplasia?

Answer 9

The tissue appears normal but there are too many cells

Question 10

What is dysplasia?

Answer 10

The cells appear abnormal and the relative number of different cell types are abnormal
Precancerous state

Question 11

What two factors may be the cause of DNA damage in precancerous cells?

Answer 11

Telomere shortening and defects in the DNA repair proteins

Expression of oncogenes

Question 12

In precancerous cells, what is the general trend?

Answer 12

The cells responding to DNA damage increase

Apoptosis increases

Question 13

In cancerous cells, what is the general trend?

Answer 13

Less cells respond to DNA damage than precancerous cells but more than normal cells
Apoptosis decreases

Question 14

What is the less aggressive nature of precancerous lesions a result of?

Answer 14

Tumourigenesis barrier imposed by the DNA damage checkpoint (p53)

Question 15

What is responsible for the continuous levels of DNA damage in precancerous cells?

Answer 15

Oncogenes deregulate DNA replication -> leads to collapsed replication forks and DNA breaks which initiate DDR

Question 16

What is the process of replication stress?

Answer 16

DNA replication -> Replication fork stalling due to anything which block replication e.g. bulky DNA lesions -> Replication fork collapse -> one-ended break

Question 17

How does transcription-replication collisions occur?

Answer 17

In a codirectional encounter - upregulation of transcription makes the replication fork catch up with the RNA polymerase and causes a collision
Head on encounters - The replication fork and the RNA polymerase hit each other head on

Question 18

In terms of replication, what do oncogenes cause?

Answer 18

Shortened G1 phase

Replication origins to fire in intragenic regions

Question 19

What is intragenic origin firing related to?

Answer 19

Increased transcription-replication collisions and formation of DNA breaks

Question 20

Why can R-loops be harmful?

Answer 20

They block DNA replication

Question 21

What prevents R loops?

Answer 21

RNA binding proteins

Question 22

What is SFPQ?

Answer 22

A splicing factor

Question 23

What happens if the splicing factor is removed?

Answer 23

Increase in single stranded DNA and DNA-RNA hybrid
Increased DNA replication
Increased DNA breaks due to collapse of replication forks
Increase DNA damage signalling
- Increased phosphorylation of H2AX
- Increased phosphorylation of Chk1

Question 24

What does SFPQ interact with?

Answer 24

DHX9 and RNA Pol II

Question 25

What is DHX9?

Answer 25

A DNA helicase (unwinds DNA)

Question 26

What does DHX9 loss cause?

Answer 26

DNA replication rescue

Question 27

In terms of R-loops, what does DHX9 KO cause?

Answer 27

No effect on R loops

Question 28

In terms of R-loops, what does DHX9 + SFPQ KO cause?

Answer 28

Reduction in R loops

Question 29

In terms of R-loops, what does SPFQ KO cause?

Answer 29

Gain in R loops

Question 30

If RNA pol II is phosphorylated on serine 2, what will it do?

Answer 30

Terminate RNA pol II

Question 31

If RNA pol II is phosphorylated on serine 5, what will it do?

Answer 31

Initiate RNA pol II

Question 32

If RNA pol II is phosphorylated on serine 2 and serine 5, what will it do?

Answer 32

Elongate RNA pol II

Question 33

How are R loops formed?

Answer 33

without splicing factors, DHX9 binds to the phosphorylated serine 2 of RNA pol I and doesn't leave in the beginning like it is supposed to

Question 34

Where does DHX9 bind to RNA pol II in normal cells?

Answer 34

Serine 5

Question 35

What do R loops cause RNA pol II to do?

Answer 35

Be stuck on the DNA

Question 36

What is the pull down of splicing factors dependent on?

Answer 36

DHX9 and RNA

Question 37

How does DXH9 cause splicing factors to bid to RNA pol II?

Answer 37

DHX9 binds RNA secondary structures and unwinds the nascent transcript -> RNA binding proteins bind nascent RNA to form ribonuclear protein

Question 38

What happens if there is no DXH9?

Answer 38

RNA secondary structures persist, preventing the formation of RNA-DNA hybrid

Question 39

What happens if there is no RNA binding proteins?

Answer 39

Nascent RNA can pair with DNA complement to form RNA-DNA hybrid -> RNA pol II is trapped -> increased transcription-replication conflicts and genomic instability