Lecture 3 Unit 4 Strobel - Homologous Recombination

Question 1

When do double strand breaks occur?

Answer 1

when both strands of a DNA double helix are severed

Question 2

What are the 2 severe consequences to the genome when double strand breaks occur?

Answer 2

• major chromosomal rearrangements
• deletion of larger chromosome segments

Question 3

What are some things that cause double strand breaks?

Answer 3

• DNA replication blockage/collisions
• ionizing radiation
• chemical agents

Question 4

Name the 2 strategies for repairing double strand breaks.

Answer 4

Homology-directed repair and non-homologous end-joining

Question 5

What does Homology-directed Repair do?

Answer 5

repairs the broken DNA ends using homologous DNA sequences as a template
* this is high fidelity
* this needs a sister chromatid

Question 6

What does Non-homologous end-joining do?

Answer 6

joins the ends of a double-strand break back together WITHOUT USING A TEMPLATE
* fidelity is variable
* can be used at any time, but may result in imprecise double-strand break repair

Question 7

What occurs during the 1st step of non-homologous end-joining?

Answer 7

Step 1: end-binding proteins assemble onto the DNA ends at the double strand break

Question 8

What occurs during step 2 of non-homologous end-joining?

Answer 8

Step 2: a protein crossbridge is assembled between the end-binding proteins
* the protein crossbridge keeps the DNA ends from drifting apart

Question 9

What occurs during step 3 of non-homologous end-joining?

Answer 9

Step 3: the DNA ends are processed in preparation for repair
* for successful repair, the DNA must have 3’ OH and 5’ phosphate ends

Question 10

What occurs during step 4 of non-homologous end-joining?

Answer 10

Step 4: the processed DNA ends are ligated together (the ends come together)

Question 11

What occurs during the 1st step in the general mechanism of Homologous Recombination Repair?

Answer 11

Step 1: End processing: the 5’ ends at the double-strand break are digested in the 5’ to 3’ direction
* this yields 3’ overhang that are complementary to one strand

Question 12

What occurs during Step 2 in the general mechanism of Homologous Recombination Repair?

Answer 12

Step 2: Strand exchange; the 3’ overhangs base pair with the complementary region in the DNA that will be used as a repair template
* the overhangs will be used as primers

Question 13

What occurs during Step 3 in the general mechanism of Homologous Recombination Repair?

Answer 13

Step 3: DNA synthesis; the 3’ overhangs are extended by DNA polymerase and once DNA synthesis is complete, they are ligated to seal the double strand break
* this results in a complex DNA structure called a double-Holliday junction

Question 14

What occurs during Step 4 in the general mechanism of Homologous Recombination Repair?

Answer 14

Step 4: Resolution: the two DNA duplexes are disentangled so that the double strand break is repaired

Question 15

What does homologous recombination enable the exchange of?

Answer 15

it enables the exchange of genetic material between chromosomes
* this is called sister chromatid exchange

Question 16

Why does sister chromatid exchange NOT result in new alleles during mitosis?

Answer 16

because the sister chromatids are genetically identical

Question 17

When does gene conversion occur?

Answer 17

when one allele is converted to the allele on the homologous chromosome

Question 18

Name the model that was proposed to explain the phenomenon of gene conversion.

Answer 18

the Holliday Model
* proposed by Robin Holliday

Question 19

What is the first step proposed in the Holliday Model?

Answer 19

Nicks are introduced at identical sites in one strand of two homologous chromatids

Question 20

What is the second step proposed in the Holliday Model?

Answer 20

the strands invade the helix of the opposite chromatid by base pairing with the complementary strand

Question 21

What is the third step proposed in the Holliday Model?

Answer 21

the invading strands are linked to the opposite chromatid to yield a Holliday Junction

Question 22

What is the fourth step proposed in the Holliday Model?

Answer 22

The Holliday junctions can migrate laterally, so that strands from each homologous chromatid are swapped

Question 23

How does homologous recombination lead to gene conversion?

Answer 23

1. DNA mismatch repair of heteroduplexes
2. DNA gap repair

Question 24

What does Branch Migration of the Holliday Model cause?

Answer 24

• it causes a heteroduplex to form
• heteroduplexes contain mismatches and will be repaired by the cell