sign in sign up
<<
Home , DNA clamp

Advanced Cell Biology. Lecture 14

Alexey Shipunov

Minot State University

February 13, 2013

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 1 / 34 Outline

Questions and answers

DNA DNA replication problem DNA reparation

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 2 / 34 Outline

Questions and answers

DNA DNA replication Telomere problem DNA reparation

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 2 / 34 I Primers should be used for starting chain, therefore they should be nucleic acids

I is not a perfect replicase and will make errors

I To replace/remove potentially erroneous primers, they need to be distinguishable

Questions and answers Previous final question: the answer

Why cells use RNA as DNA replication primers?

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 3 / 34 Questions and answers Previous final question: the answer

Why cells use RNA as DNA replication primers?

I Primers should be used for starting nucleotide chain, therefore they should be nucleic acids

I Primase is not a perfect replicase and will make errors

I To replace/remove potentially erroneous primers, they need to be distinguishable

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 3 / 34 DNA DNA replication

DNA DNA replication

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 4 / 34 DNA DNA replication

How are added in DNA replication

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 5 / 34 DNA DNA replication Sliding clamp (DNA clamp)

I Keeps DNA attached to the template

I Form a ring around different DNA

I Most are trimers of PCNA proteins

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 6 / 34 DNA DNA replication Human DNA clamp protein (trimer of PCNA)

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 7 / 34 DNA DNA replication

DNA sliding clamp movie

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 8 / 34 DNA DNA replication DNA replication complex

1. DNA: old helix, leading strand, lagging strand 2. DNA polymerases 3. DNA 4. SSBP 5. DNA clamp 6. Primase 7. DNA ligase

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 9 / 34 DNA DNA replication DNA replication machine (simplified)

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 10 / 34 DNA DNA replication DNA replication machine (more realistic)

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 11 / 34 DNA DNA replication

DNA replication sewing machine movie

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 12 / 34 DNA DNA replication

DNA replication in general: movie I

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 13 / 34 DNA Telomere problem

DNA Telomere problem

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 14 / 34 DNA Telomere problem

I Lagging strand cannot reach the end of DNA molecule

I Every replication cycle lost parts of from its ends

I To prevent a loss on meaningful DNA fragments, telomerase extends chromosome with new telomere sequences

I In humans, telomeres are several thousands of TTAGGG sequences

I Telomeres are also recognizable ends of

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 15 / 34 DNA Telomere problem Telomerase and telomere

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 16 / 34 DNA Telomere problem

Telomere movie

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 17 / 34 DNA Telomere problem Telomere theory of aging

I In humans, telomerase is only active in germ cells, stem cells and certain white blood cells

I Olovnikov (1971) postulated that lost of DNA ends will eventually stop division of cells and may stimulate senescence of cells

I There is a strong support of this hypothesis with some cell types (e.g., blood vessels wall cells); however, mice with knocked-out telomerase gene do not show significantly less lifespan

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 18 / 34 DNA DNA reparation

DNA DNA reparation

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 19 / 34 DNA DNA reparation Mutation theory of ageing and/or cancer

I Accumulation of mutations will result in a constant loss of functions

I Cells will either degrade or start to go out of control (cancer)

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 20 / 34 DNA DNA reparation DNA mismatch repair

−7 I Normally, error rate of DNA polymerase + proofreading is ≈10 −9 I DNA mismatch repair proteins decrease it to ≈10

I They react on DNA conformation deviations; recognize newly synthesized strand by nicks, and remove wrong fragments which are later replaced with DNA polymerase and ligase

I Some cancers are results of mutations in DNA mismatch protein genes

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 21 / 34 DNA DNA reparation DNA mismatched repair system

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 22 / 34 DNA DNA reparation Depurination and deamination

I Depurination is detaching A and G from its sugar; it results in deletion*

I Deamination is C to U conversion; it results in substitution*

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 23 / 34 DNA DNA reparation Depurination and deamination

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 24 / 34 DNA DNA reparation Pyrimidine dimers

I UV radiation often results in forming of pyrimidine (T or C) dimers

I Dimers will stop DNA replication and transcription

I The final result is often a melanoma type of skin cancer

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 25 / 34 DNA DNA reparation Pyrimidine dimers

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 26 / 34 DNA DNA reparation Skin melanoma

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 27 / 34 DNA DNA reparation Basic way of DNA repair

1. Damage recognition 2. Excision 3. Resynthesis 4. Ligation

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 28 / 34 DNA DNA reparation DNA repair flow

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 29 / 34 DNA DNA reparation Nonhomologous end-joining

I The quick-and-dirty mechanism of reparation when both strands are broken

I The reparation will result in deletion

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 30 / 34 DNA DNA reparation Nonhomologous end-joining

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 31 / 34 What is the difference between deletion and substitution?

DNA DNA reparation Final question (1 point)

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 32 / 34 DNA DNA reparation Final question (1 point)

What is the difference between deletion and substitution?

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 32 / 34 DNA DNA reparation Summary

I Ends of chromosomes are constantly shortening and extending with new telomeres

I DNA replication system is a multienzyme complex

I Ends of chromosomes are constantly shortening and extending with new telomeres

I DNA suffers from multiple damaging events; multiple reparation systems are trying to lower mutation risks

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 33 / 34 DNA DNA reparation For Further Reading

A. Shipunov. Advanced Cell Biology [Electronic resource]. 2011—onwards. Mode of access: http://ashipunov.info/shipunov/school/biol_250

B. Alberts et al. Essential Cell Biology. 3rd edition. Garland Science, 2009. Chapter 6.

Shipunov (MSU) Advanced Cell Biology. Lecture 14 February 13, 2013 34 / 34