DNA Repair DNA Repair

Uracil-DNA Glycosylase

DNA is continually assaulted by damaging agents (oxygen free radicals, ultraviolet light, toxic chemicals).

Fortunately, the cell has multiple mechanisms to identify and correct mutations.

Yeasts have 50 different enzymes involved in DNA repair.

Damage occurs in the form of deletions and mutations (changes in the sequence of DNA bases that make up the genetic code).

The DNA repair process may be a major factor in aging, health, and longevity.

An animal's ability to repair certain types of DNA damage is directly related to the life span of its species

Genetic defects in DNA repair are associated with genetic or familial susceptibility to cancer.

Figure from the Duke University Medical Center and the Graduate Program in Molecular Cancer Biologyhttp://mcb.mc.duke.edu/

Cell nuclei and chromosomes from a human brain tumor

Mitochondrial DNA is more susceptible to damage than nuclear DNA. Furthermore, mitochondrial DNA damage increases exponentially with age.

Several diseases that appear late in life, including late-onset diabetes, have been traced to defects in mitochondria.

Causes of Mutations

DNA replicationSpontaneous depurination and deaminationUV lightOxidationChemically induced

DNA Mismatch Repair (E. coli)How does the cell know which strand

to repair?Deoxyadenosine methylase (dam)Old DNA methylatedNew DNA unmethylatedMismatch repair system cuts

unmethylated DNAIn eukaryotes the recognition may be

nicks.

DNA Repair and Cancer

Do humans have a mismatch repair system?

hMSH, human Mut S HomologhMLH, human Mut L HomologWhat phenotype would be expected

from a mutation in hMSH or hMLH?Hereditary nonpolyposis colon

cancer (HNPCC), Lynch syndrome II

Depurination and Deamination

Loose A or G

Loose NH2 groups from C (forms U)

Base-Excision Repair Pathway All organisms contain a specific repair

pathway which removes uracil from DNA.

DNA Damage From UV Light

Single-strand breaks - mostly sealed by DNA ligase so don't contribute to lethality

Double-strand breaks - often lethal because can't be resealed by ligase so degraded by nucleases

Alteration of bases - this type of oxidative damage is usually lethal because forms a replication barrier at that site

Ionizing radiation causes three types of damage to DNAIonizing radiation causes three types of damage to DNA

uvrABCD Repair System

What would be the phenotype in humans from an accumulation of thymine dimers in somatic cells?

What would be the phenotype in humans from mutations in uvrABCD genes?

Xeroderma Pigmentosum (XP)

Disorders Associated with Defective DNA Repair

Xeroderma pigmentosum (XP)Skin photosensitivity; Early onset skin cancer

Cockayne SyndromeDwarfism; Precociously senile appearance Sensitivity to sunlight

Bloom Syndrome

Dwarfism, low-birth-weight type;

Life-threatening infections Predisposition to neoplasia

Helicase: Bloom syndrome (genomic instability in somatic cells, leads to telangiectatic facial erythema, photosensitivity, dwarfism and other abnormalities)

BRCA1 DNA repair protein Breast and Ovarian cancer, early onset

MSH2 DNA repair protein Hereditary non-polyposis colon cancer

Mutations in DNA repair genes lead to an increase in the frequency of other mutations.

Ataxia-Telangiectasia

Approximately 10% of children diagnosed with AT will develop a malignancy in childhood or early adulthood.

The gene defect in AT patients allows the formation of a much higher level of chromosome translocations.

Transcription occurs in nucleus, translation occurs in cytoplasm:

RNA must move across nuclear membrane.

Transcription in eukaryotes Transcription in eukaryotes

Regulation involves multiple enzymes & proteins -30 'TATAAAA' (Hogness box) promoter -50 ~ -500 enhancer sequences control rate

Post-transcriptional processingPost-transcriptional processing

poly-A 'tail' (5'-AAAAAA-etc-AAAAAA-3') added to 3' end (Fig.12.10a) 7mG 'cap' (7-methyl guanosine, 7mG) added to 5' end splicing of heterogeneous nuclear RNA (hnRNA) (Fig.12.10b) up to 90% of transcript is removed exons are retained ("expressed") introns are removed ("intervening") 10 ~ 20 exons / 'gene'