DNA damage and repair

Why is the stability of DNA essential for cell survival?

It encodes the genetic instructions for the development and functioning of all known living organisms.

Which biological macromolecule is unique in its ability to undergo repair?

DNA.

What are the two primary consequences of DNA damage in proliferating cells?

Cell death or mutation.

The accumulation of mutations in proliferating cells can lead to the development of _.

Cancer

What is a primary consequence of DNA damage in non-dividing cells regarding gene expression?

The blocking of transcription.

In non-dividing cells, a decline in tissue repair caused by DNA damage leads to _.

Aging

How is 'endogenous DNA damage' defined?

Damage occurring spontaneously within the cell.

Identify three sources of endogenous DNA damage.

Hydrolysis, oxygen species, and by-products of metabolism.

How is 'exogenous DNA damage' defined?

Damage caused by reactions with molecules from outside the cell.

Identify three external factors that serve as exogenous sources of DNA damage.

UV light, X-rays, and carcinogens (or chemotherapeutics).

Why is DNA damage more critical than damage to proteins or lipids?

DNA contains genetic information and cannot be replaced, unlike proteins or lipids.

Which chemical bond is susceptible to hydrolysis during depurination?

The N-glycosidic bond connecting the purine to the deoxyribose sugar.

What is the result of a depurination event in the DNA helix?

An abasic site.

Approximately how many depurination events occur per human genome per day?

18,000.00

What are the potential mutagenic outcomes of an abasic site?

Deletion mutations, substitution mutations, or strand breakage.

The removal of amino acids from DNA bases by hydrolysis is known as _.

Deamination

What base is created when cytosine undergoes deamination?

Uracil.

Why are transition mutations more likely to occur than transversions?

Substituting a double ring structure for another double ring is more likely than substituting it for a single ring.

What type of protein is generated as a result of a frameshift mutation?

Missense proteins.

Which specific DNA lesion is induced by exposure to UV light?

Pyrimidine dimers.

How do pyrimidine dimers physically affect the DNA helix?

They cause distortion and prevent the DNA from being flexible.

Besides dimers, what other toxic lesions can UV light induce?

Interstrand DNA crosslinks and DNA-protein crosslinks.

Why are DNA crosslinks considered highly toxic to the cell?

They block both replication and transcription.

Which enzyme scans DNA and flips bases out of the helix to identify damage in Base Excision Repair (BER)?

Uracil glycosylase.

In BER, what is the role of uracil glycosylase once a damaged base is identified?

It cleaves the bond between the base and the sugar to leave an abasic site.

Which enzyme cuts the phosphodiester backbone at the 5' side of an abasic site during BER?

AP endonuclease.

What is the function of phosphodiesterase in the BER pathway?

It removes the leftover sugar-phosphate residue after the backbone is cut.

Which enzyme adds the correct new nucleotide to the gap during Base Excision Repair?

DNA polymerase.

In BER, which enzyme is responsible for sealing the final nick in the sugar-phosphate backbone?

DNA ligase.

Which enzyme identifies helix distortions in the Nucleotide Excision Repair (NER) pathway?

Excision nuclease.

How does excision nuclease initiate the repair process in NER?

It makes two cuts in the sugar-phosphate backbone, one on either side of the damage.

What is the role of DNA helicase in Nucleotide Excision Repair?

It unwinds the DNA to peel away the damaged single-stranded segment.

What does DNA polymerase use as a template to fill the gap during NER?

The healthy opposite strand.

When does a cell employ Translesion Synthesis (TLS)?

As a last resort to bypass major DNA lesions that stall replication.

What occurs when a replicative polymerase encounters a bulky lesion?

It stops because it cannot fit the damaged bases into its active site.

Which chemical 'flags' signal the replicative polymerase to release the DNA during a stall?

Covalent modifications to the sliding clamp.

How do the active sites of translesion DNA polymerases differ from replicative polymerases?

They are much larger to accommodate distorted or damaged bases.

What is the primary genetic risk associated with translesion synthesis?

It lacks precision in template recognition and substrate base choice.

Why are translesion polymerases unable to fix mistakes made during synthesis?

They lack proof-reading activity.

Which types of mutations are most frequently caused by translesion synthesis?

Base substitutions and single nucleotide deletions.

What allows high-fidelity replicative DNA polymerase to reload after translesion synthesis?

The removal of covalent modifications from the sliding clamp.

Which protein complex binds to DNA ends and recruits DNA-PKcs in Non-Homologous End Joining (NHEJ)?

Ku70/8.

What is the role of Artemis in the NHEJ pathway?

It trims nucleotides at messy or mismatched DNA ends to make them ligatable.

Which complex catalyses the formation of new phosphodiester bonds to close a break in NHEJ?

The XRCC4-DNA ligase IV complex.

What is the major disadvantage of Non-Homologous End Joining?

Genetic information is often lost due to the loss of nucleotides surrounding the break site.

How does the MRN complex initiate Homologous Recombination (HR)?

It trims the 5' ends of broken DNA to create single-stranded DNA tails.

What is the function of Replication Protein A (RPA) in Homologous Recombination?

It coats single-stranded DNA tails to prevent them from tangling or degrading.

Which proteins facilitate 'strand invasion' during Homologous Recombination?

Rad51, Rad52, Rad54, and tumour suppressors BRCA1 and BRCA2.

In HR, what term describes the point where broken DNA is physically linked to its template sister chromatid?

Holiday junction.

Why is Homologous Recombination considered more accurate than NHEJ?

It uses the sister chromatid as a guide to restore missing information without loss.

Identify the three points in the cell cycle where DNA damage is detected.

G1, entry to S-phase, and entry into mitosis.

Which kinases associate with the site of DNA damage to block the cell cycle?

ATM/ATR.

How does p53 contribute to cell cycle arrest after DNA damage?

It is stabilised and activates the expression of p21.

What is the specific function of p21 in the cell cycle checkpoint?

It renders the G1/S-CDK complex inactive to prevent cycle progression.

What is the cellular fate if DNA damage cannot be successfully repaired?

Apoptosis.

What is the inheritance pattern of Xeroderma pigmentosum?

Autosomal recessive.

Patients with Xeroderma pigmentosum have a 2000-fold increased risk of which condition?

Skin cancer.

Which tumour suppressor genes are specifically associated with inherited breast cancers?

BRCA1 and BRCA2.

What allows cancer cells to grow very fast despite DNA damage?

They are able to replicate in the presence of damage.

Define the concept of 'synthetic lethality'.

A combination of two specific genetic defects leads to cell death, even though either defect alone does not.