Mitochondrial Replication

Question 1

triple-strand specific RNAse

Answer 1

needs a displaced strand in the RNA-DNA complex
- 3’ end of RNA primer is cut with triple-strand specific RNAse H

Question 2

SAR11 clade

Answer 2

• alphaproteobacteria, Rickesettsiales
• able to handle oxygen
• these were the ancient symbiont ancestor to eukaryotic mitochondria
• this group of organisms represent 1 in 3 of cells in the ocean’s surface

Question 3

how is mitochondrial DNA passed?

Answer 3

• mitochondria are usually transmitted by the female
• we all have maternal mitochondria only

Question 4

D-loop

Answer 4

• does not code for anything, free to mutate and used to study human evolution
• molecular clock
• old light strand is displaced and remains single-stranded

Question 5

molecular clock region

Answer 5

• D-loop
• mutations in the D-loop
• mutations do not affect gene product and is not selected against

Question 6

what is the rate in which mitochondria have mutations?

Answer 6

• mitochondria are 15 more likely to have mutations than the nuclear genome
• this is because there are no histones and are more exposed to radiation and it exists as single strand DNA so it is subject to more oxidation, metabolic reaction that alters the bases

Question 7

how many origins are there in human mitochondria and what direction does the replication fork go?

Answer 7

• unidirectional
• each strand of mitochondria has its own origin that fire at different times
• 2 unidirectional replicons

Question 8

what is the placement of mit-RNA genes?

Answer 8

• position 57
• the arrested 75 D-loop primer is used when the mtDNA is severely depleted - positions 151 and 191

Question 9

light strand promoter

Answer 9

• initiate transcription of mRNA
• initiate RNA primers for DNA replication

Question 10

which strand encodes (serves as template) the most genes?

Answer 10

• heavy strand
• outside of the circle

Question 11

what are the three primers used in mitochondrial replication?

Answer 11

• 2 for stress conditions and one for normal replication
• stress conditions: origins 151 and 191
• normal conditions: origins 57

Question 12

what is the significance of the D-loop?

Answer 12

• a primer
• does not encode for anything
• free to mutate with few constraints

Question 13

how many origins of replications are present in the mitochondrial genome?

Answer 13

2 - one on the heavy strand and one on the light strand

Question 14

what kind of replication priming occurs within the D-loop?

Answer 14

under stress conditions

Question 15

what enzyme cuts the RNA primer in order to generate the end to be extended by DNA polymerase?

Answer 15

triple strand specific RNAseH

Question 16

what are the composition of the D-loop of human mitochondria?

Answer 16

• RNA portion of the primer is 600 bases
• DNA portion of the primer 1,400 bases
• the primer for replication of the H strand is comprised of an RNA-DNA composite molecule
• single D-loop with an opening of 7S

Question 17

tetrahymena mitochondrial DNA

Answer 17

• has 6 D-loops at a time (many origins), and plant chloroplasts have 2

Question 18

what is the origin of replication of mtDNA?

Answer 18

• illustrates the principle that an origin may be used to initiate a single strand only in the case of D loops
• another example is PhiX174 rolling circle replication
• an origin can be a sequence of DNA that initiates DNA synthesis using one strand as a template

Question 19

what is mit RNA polymerase single subunit related to?

Answer 19

• phage T7, T3, and SP6 polymerase
• the protein-encoding RNA have a short polyA tail added at the 3’ end

Question 20

which enzyme processes the short RNA primer for synthesis of the H-strand of mitDNA?

Answer 20

triple strand-specific RNase H

Question 21

What is thought to be the ancestral organism group that gave rise to mitochondria?

Answer 21

Oxygen dwelling bacteria could deal with oxygen through oxidation and reduction
SAR11

Question 22

where did many mitochondrial genes migrate to?

Answer 22

• nucleus

Question 23

where are the H and L strand origins located?

Answer 23

• origin on the H strand to make L
• origin on L strand to make H

Question 24

how does mitochondrial DNA replicate?

Answer 24

• RNA primer within the D-loop is synthesized the new H strand
• 3’ end of RNA primer is cut with triple-strand specific RNase H
• old H strand is displaced and new H strand primed
• RNA primer is extended with mtDNA polymerase to make a 1,400 base primer
• L strand synthesis is initiated using origin on H strand
• synthesis of L strand is initiated when its origin is exposed by the movement of the first replication fork
• then the old H strand is displaced with the L strand is partially replicated and completes
• the second strand is made with the old L strand and a new H strand is made
• gaps in new strands sealed