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DNA damage and repair

High Yield Notes
15 pages
Transcript

Content Reviewers:

Rishi Desai, MD, MPH

Our DNA is like a library - found in the nucleus of our cells - with thousands of books.

Some of these books - called genes - are extremely important, because they carry the recipes for every single protein found in the cell.

Now, on a molecular level, DNA is made up of two strands of nucleotides, so each gene is just a segment of this nucleotide sequence.

Nucleotides of DNA are made out of a sugar - deoxyribose, a phosphate, and one of the four nucleobases - adenine, cytosine, guanine, and thymine - or, A, C, G, T for short.

The nucleotides on one strand pair up using hydrogen bonds with nucleotides on the opposing strand, to create the double-stranded DNA: specifically, A bonds with T, and C bonds with G, so they’re called complementary bases.

Now, the goal of DNA is to store information and pass it onto their daughter cells, and to use this information to create proteins.

To do this, there are two critical processes - DNA replication and gene expression.

DNA replication occurs during the cell cycle - more specifically, during the S phase of interphase.

So, the cell cycle is made up of interphase - when the cell prepares for division - and mitosis - or the actual splitting of the cell in two daughter cell.

Interphase has 3 subphases - G1, S and G2, and during the S subphase, the cell replicates its DNA, so that the two daughter cells get the exact same DNA during mitosis.

If we zoom onto the double- stranded DNA, we can see that during DNA replication, the two DNA strands are separated by an enzyme called DNA helicase.

Then another enzyme, DNA polymerase, uses each of the single strands as a template and adds complementary nucleotides to it.

Gene expression, on the other hand, is the process of decoding the information stored in the DNA in order for the cell to make proteins, and it includes transcription and translation.

Transcription is where RNA polymerase copies the nucleotide sequence of the gene and creates a messenger RNA molecule, or mRNA that has the same sequence, with one tweak: it has uracil nucleotides - or U - instead of thymine.

Now during translation, cell organelles called ribosomes “read” the mRNA molecule in 3 nucleotide “words”, called codons - with each 3 nucleotide sequence coding for an amino acid that will eventually become part of the protein.

So, for the cell to keep functioning, the DNA strands need to remain intact, or at least mostly intact, in order to pass on or express unaltered genetic information.

Unfortunately, the cell is exposed all the time to both endogenous, and exogenous or environmental factors that can damage the DNA.

Luckily, if DNA gets damaged, the cell can enter a special phase outside the cell cycle - the G0 phase - where DNA repair mechanism try to fix the damage.

If the DNA damage starts to pile up - a cell will typically go down one of three paths.

First, the cell might go into senescence - which is when the cell stops dividing.

Second, the cell might undergo apoptosis, which is programmed cell death.

Third, the cell might begin to undergo uncontrolled cell division and develop into a tumor.

None of these paths are ideal, so it’s essential for cells to fix reversible DNA damage and prevent too many DNA mutations from occurring.

Broadly speaking, there might be single strand damage - or double strand damage, and the cell has mechanisms to address both situations.

Single strand damage can happen because of endogenous causes - like errors in DNA replication - or exogenous factors - like harmful chemical or physical agents.

Single strand damage is fixed by three repair mechanisms: mismatch repair, base excision repair, and nucleotide excision repair.

During replication, DNA polymerase can sometimes put in the wrong nucleotide - like pairing adenine up with a cytosine instead of a thymine.

This is called a mismatch, and it happens about 60,000 times per replication - so 1 out of 100,000 nucleotides.

Now the first way to fix a mismatch is right after it happens - because DNA polymerase is quite a resourceful enzyme, and it can look over its shoulder to check for errors and see if it put the right nucleotide in.

Kinda like checking an essay for typos before sending it in.

If DNA polymerase finds a mismatch, it goes back and acts as an exonuclease - meaning, it removes the wrong nucleotide from the newly synthesized DNA strand and replaces it with the correct nucleotide.

This is called proofreading - and while it still leaves some mismatches behind, it reduces the error rate to about 600 times per replication - so 1 out of 10 million nucleotides.

Next is mismatch repair - which relies on special proteins, called MSH proteins - and fixes the remaining errors after replication.

When MSH proteins see a mismatch in a newly synthesized strand, they recruit an enzyme - called endonuclease - that acts like a pair of scissors and severs the nucleotide bonds from the DNA strand.

Then another enzyme - called exonuclease - removes the damaged segment of DNA, leaving a gap in the daughter DNA strand.

Then, DNA polymerase can come in and fill this gap with new nucleotides - kinda like when you have to go back and rewrite a paragraph of your essay if your supervisor tells you there's an error in there.

And finally, once DNA polymerase is done matching correct nucleotides, another enzyme, DNA ligase, seals the bonds and the damage is successfully repaired.

But even mismatch repair leaves a tiny number of errors, which is six errors per cell division - so 1 out of 1 billion nucleotides.

Of course, nothing compared to the original 60,000.

Now, base excision repair comes in when the cell's DNA suffers damage from exposure to harmful chemicals or physical factors.