Altering the genetic message: Mutation Cancer. Chapter 17 Altering the genetic message: Mutation Cancer.

= DNA Damage + DNA Repair. MUTATIONS = change in the genetic message = change in the nucleotide sequence of a gene = DNA Damage + DNA Repair. 1. Types of mutations 2. Causes of DNA damage 3. DNA (damage) repair

Types of mutations. A. Gene mutations a. point mutations or base-pair substitution b. frame-shift mutations B. Chromosomal mutations (duplication, inversion, deletion, translocation)

Causes of DNA damage. (tautomeric shift, deamination, 1. Spontaneous (tautomeric shift, deamination, depurination, looping-out) 2. Oxidation 3. Chemical - base-pair analogs - base-modifying agents - intercalating agents - agents that cause ‘bulky’ lesions 4. Physical: - UV light - X-ray

Causes of DNA damage. - tautomeric shift - deamination - depurination 1. Spontaneous - tautomeric shift - deamination - depurination - strand slippage/looping-out

Tautomeric shift of bases Common Form Uncommon Form

Tautomeric shift leads to altered base pairing.

Depurination of DNA Apurinic sites

De-amination of cytosine or methyl-cytosine.

Strand slippage during DNA replication 5’TACGGAC 3’ 3’ATGCCTGACTTTGC 5’ Newly synthesized DNA Template DNA 5’TACGGACTG 3’ 3’ATGCCTGACTTTGC 5’ T Newly synthesized DNA loops out, … 5’TACGGACTGAAACG 3’ 3’ATGCCTGACTTTGC 5’ T … resulting in the addition of one nucleotide in the new strand.

Strand slippage during DNA replication 5’TACGGAC 3’ 3’ATGCCTGACTTTGC 5’ Newly synthesized DNA Template DNA 5’TACGGACTG 3’ 3’ATGCCTGCTTTGC 5’ Template DNA loops out, … A … resulting in the omission of one nucleotide in the new strand. 5’TACGGACGAAACG 3’ 3’ATGCCTGCTTTGC 5’ A

Causes of DNA Damage 2. Oxidative damage (Respiration, Mixed function oxidases, Inflammation)

Causes of DNA damage. 3. Physical

UV-Light (254 – 260 nm) Pyrimidine dimers Gross distortion DNA molecule

Malignant Melanoma

Causes of DNA damage. 3. Physical There are two main ways radiation can damage DNA inside living cells. Radiation can strike the DNA molecule directly, ionizing and damaging it. Alternately, radiation can ionize water molecules, producing free radicals that react with and damage DNA molecules.

Causes of DNA damage. 4. Chemical - base-pair analogs - base-modifying agents - alkylating agents - base deaminating - intercalating agents - agents that cause ‘bulky’ additions

Causes of DNA damage 4. Chemical: Base Analogs Miss-pairing

Alkylating agents add alkyl groups  Cause altered base pairing E.g., Ethyl-methane-sulfonate (mustard gas), Nitroso-guanidine.

Base deamination E.g., nitrous acid, bisulfite

Intercalating agents Actinomycin Ethidium Bromide Acriding Orange Tetracycline Proflavine Intercalating agents

Bulky additions grossly distort the DNA

Aflatoxins … ‘bulky’ additions, gross distortions of the DNA

Benzo (a) pyrene

Error-prone DNA repair Cellular Responses to DNA damage Error-free DNA repair Apoptosis Error-prone DNA repair Mutation

DNA Repair: 1. Direct correction of DNA repair a. Proofreading of DNA polymerase b. Repair of alkylating damage 2. Repair involving excission of base pairs a. General excission repair system (UvABC) b. Repair by glycosylases and AP endonucleases c. Mismatch-repair system d. SOS-repair system

Proofreading Capacity of DNA Polymerase

Repair by Alkyl-transferases

General Excision repair system

Xeroderma pigmentosa: Deficiency of the general excision repair system.

Glycosylase/ AP endonuclease Repair system

Repair by DNA glycosylase and AP endonucleases.

Mismatch Repair System

Hereditary Non-Polyposus Colon-rectal Cancer (HNPCC)

Error-prone repair by end-joining.

Altering the genetic message: Cancer. Chapter 17 Altering the genetic message: Cancer.

Cancer Independent of growth signals Tissue invasion metastasis Insensitive to growth inhibitors Sustained angiogenesis Cancer Changed energy metabolism Limitless replicative potential Avoid apoptosis

Normal cells Cancer cells loose contact inhibition, grow on top of each other, and become rounded.

Normal Cancer

Normal (contact inhibition) Cancer

Rous sarcoma virus

J. Michael Bishop (1936 - ) and Harold E J. Michael Bishop (1936 - ) and Harold E. Varmus (1939 - ) won the Noble Prize in 1989 for proving that viruses contain a cancer-causing gene derived from the genome of the organism they infect. Specifically, they showed that chicken Rous Sarcoma Virus (RSV) carried an oncogene called v-src and this gene was an intronless version of a normal chicken gene called c-src Nobel Prize 1989

Chicken Rous Sarcoma Virus (RSV) carried an oncogene called v-src and this gene was an intronless version of a normal chicken gene called c-src.

Active tumor suppressor gene(s) Inactive tumor suppressor gene(s) DNA Damage + DNA Repair Active tumor suppressor gene(s) Proto-oncogene(s) MUTATIONS Oncogene(s) Inactive tumor suppressor gene(s) Gain-of-function Dominant phenotype Loss-of-function CANCER Recessive phenotype

Retinoblastoma. S-phase EF2 Rb Cdk2-cyclin E DNA mRNA DNA polymerase P + DNA mRNA DNA polymerase S-phase

CANCER: A multi-hit process of mutations accumulating in proto-oncogenes and tumor suppressor genes.

Familial Adenomatous Polyposis Coli

Colonoscopy Results

JB Weitzmann and Nosh Yaniv Nature 1999, 400 p401

What causes cancer? 1. Environmental carcinogens - chemical (e.g., cigarette smoke) - physical (e.g., UV radiation) 2. Host carcinogens (e.g., inflammation) 3. Viruses:

TUMOR VIRUSES Papiloma virus Hepatitis B virus Human Herpes virus 8 (Kaposi) Human Herpes virus 4 (Epstein Bar) Human T lymphotropic virus

This woman has hepatitis B and is suffering from liver cancer This woman has hepatitis B and is suffering from liver cancer. She was a Cambodian refugee and died 4 months after she arrived in a refugee camp (average life expectancy after diagnosis of liver cancer is 6 months)

Kaposi syndrome: Human Herpes Virus 8)

Cutaneous B cell lymphoma

HTLV Leukemia

The End.