Module 02 - Section 05

Question 1

What does Homologous Recombination (HR) refers to?

Answer 1

Recombination between two DNA molecules of similar sequence (i.e. homologous chromosomes or sequences with small allelic difference) occurring in all cells

Question 2

When does Homologous recombination take place&

Answer 2

During Meiosis and mitosis in eukaryotes, and repair of DSBs in all organisms

Question 3

In what phase of meiosis are each pair of homologous chromosomes replicated to create four chromosome copies (2 copies, or chromatids per chromosome)?

Answer 3

S Phase

Question 4

In what phase of meiosis do homologous sister chromatid pairs align?

Answer 4

First meiotic prophase (prophase I)

Question 5

In one round of meiosis, the four sets of chromosomes are segregated through two cell divisions (meiosis I and meiosis II) to create how many gametes?

Answer 5

4

Question 6

Are the gametes haploid or diploid?

Answer 6

Haploid

Question 7

In meiosis, (a) what segregates at the first cell division? (b) What about the second cell division?

Answer 7

(a) Homologous chromosomes segregate in the first cell division
(b) Sister chromatids segregate in the second cell division

Question 8

What provides the physical link between sister chromatids?

Answer 8

Proteins called cohesins

Question 9

During meiosis I, because the chromosomes to be segregated are not related by a recent replication event, what molecular device ensure that only homologous chromosomes are linked and segregated?

Answer 9

Homologous recombination; recombinational crossovers provide the accurate alignment of homologous chromosomes at the metaphase plate during meiosis

Question 10

Name a similarity and a difference between recombinational DNA repair during replication and meiotic recombination

Answer 10

Similar: many of the same enzymes are involved in both and both begin with DSBs
Difference: DSBs are programmed events in meiosis I

Question 11

What are the roles of crossing over in meiosis?

Answer 11

(1) Create the physical link essential for proper chromosomal segregation
(2) Sister chromatids are no longer identical - generating genetic diversity

Question 12

What is a chiasmata?

Answer 12

Actual crossover sites between chromatids of homologous chromosomes

Question 13

Describe the initiation of meiotic recombination in yeast

Answer 13

As cell enters meiosis, DSBs are intentionally introduced at multiple locations along one chromatid of each chromatid pair - These breaks are not random, and yet are not entirely predictable

Question 14

Which protein catalyzes the formation of DSBs

Answer 14

Spo11

Question 15

Describe the mechanism of Spo11

Answer 15

Uses an active-site Tyr residue as a nucleophile in a transesterification reaction (Process of exchanging the organic group of an ester with the organic group of an alcohol)

• Each Subunit cleaves 1 strand of DNA with the phosphodiester bond replaced by a 5’-phosphotyriosyl linkage
• reaction halts

Question 16

Which key proteins process the broken ens of DSBs? (5)

Answer 16

• Mre11-Rad50-Xrs2 complex
• Sae2
• Sgs1
• Exo1 or DNA2

Question 17

What is the role of Mre11-Rad50-Xrs2 complex in processing DSBs?

Answer 17

• Removes Spo11 by cleaving the DNA several base pairs on the 3’ side of Spo11 (Spo11 cleaves on the 5’ side)

Question 18

What is the role of Sae2 in processing DSBs?

Answer 18

After Mre11-Rad50-Xrs2 complex has cleaved off Spo11, it degrades the DNA a bit more.

Question 19

What is the role of Sgs1, nucleases Exo1 or Dna2 in processing DSBs?

Answer 19

extended degradation of the 5’ end to create long 3’ single-stranded extensions

Question 20

Describe the mechanism of meiotic recombination

Answer 20

(1) RPA binds ssDNA regions
(2) Dmc1 (recombinases) and Rad51 are loaded onto the 3’ extensions (aided by mediator proteins)
(3) Sequential cycles of binding, sampling and release of dsDNA performed until homology is detected
(4) Stable DNA joint is formed by intertwining of the ssDNA (same recombination of section 4)

Question 21

What is gene conversion?

Answer 21

Non-crossover exchange of genetic material

Question 22

Which resolution of Holliday intermediates lead to the exchange of genetic material?

Answer 22

Both crossover and non-crossover

Question 23

How to determine if crossover or non-crossover resolution of Holliday intermediates

Answer 23

(1) Non-crossover; both junction are cleaved the same way (XX or YY)
(2) Crossover; junctions are cleaved using different sites (XY or YX)

Question 24

Describe the catalysis of Holliday intermediate resolution

Answer 24

(1) RuvA; Binds specifically to Holliday junction, recognizes the structure of the DNA junction and keepts it in a box-like state
(2) RuvB; surrounds 2 of the 4 arms of Holliday intermediate which propels DNA outward through hole in its donut shape
(3) RuvC: Holliday intermediate resolvase replaces one of the RuvA and cleaves strands in opposing amrs of the Holliday intermediate - nicks 2 strands with same polarity
(4) Nicks are sealed into a different arrangement by DNA ligase