MCB 2400 Exam 3

Lysine Acetylation

ex. H4K16Ac=histone H4 lysine #16 acetylated---> associated with actively transcribed DNA
- DNA is less tightly bound: more accessible to transcription factors
- Acetylated lysine creates binding sites for specific activators

DNA Methylation

Leads to inhibition of transcription
- Removed before transcription initiation or
- Remain methylated for long-term silencing
- Attracts DNA deacetylases

DNA Methylation- Imprinting

ICR= Imprinting Control Region- it's an insulator that can't function when methylated

Mutation

An inherited change in genetic information (descendants might be cells of organisms(
- Errors during recombination (in Meiosis I and II)
- Spontaneous errors during DNA replication
- Exposure to mutagens, X rays, and cosmic rays
- Spontaneous chemical changes in DNA bases

Germ-line Mutations and Somatic Mutations

- Germ-line mutations are inherited
- Somatic mutations are not inherited
- Meiosis and sexual reproduction pass germ-line mutations to ~50% offspring
- All offspring's cells carry mutation

Spontaneous Mutations

- Mutations are random events
- Unrelated to any adaptive advantage it might confer on the organism and its environment
- A potentially favorable mutation does not arise bc the organism has a need for it
- They preexist in the population and are selected for under certain circumstances

Mutations and Evolution

Mutations are a source of variation in a population and at the heart of evolution
- Sometimes mutations work well, and sometimes not
- Ex. After a change in selective pressure, black moths dominate (context-dependent)

Classification of Mutations

Mutations cause molecular changes
- A mutation is any heritable change in genetic material
- Mutations are classified in a variety of ways

Effect of a Mutation Depends on...

What gets mutated
- Context dependent
- Third letter (also called the wobble position) has the most mutations bc the first two letters are more stable because they matter more than the third letter

Three Basic Types of Molecular Change at DNA Level

- Types of mutations at protein level: missense, nonsense, and silent mutations
-- Missense mutation- DNA changes that results in different amino acids that are encoded at a specific position in the resulting protein
-- Nonsense mutation- mutation in a sense codon that results in a change to a chain-terminating codon
-- Silent mutation- mutation in a sense codon that does not change the resulting amino acid
- If there's an insertion or deletion in the Order Reading Frame at the protein level ---> frame-shift mutations
- Frameshift mutations- reading frame is altered-->all codons downstream are affected
- Insertion or deletion of 3bp in frame- no frameshift, but missing or additional amino acid(s)

Base substitutions: Transitions and Transversions

Purines A+G= "Pure As Gold"
Pyrimidines C+T= "CuT the Pyramid"
- Purine w/ pyrimidine---> keeps sides of ladder equidistant
Number of possible transversions is double the amount of transitions but transitions is more common
- DNA repair mechanisms detect wrong DNA structure

Single Base Changes Can Result in...

Protein level changes
- Wild-type protein
- Changed codon: changed amino acid
- Changed to stop codon: truncated protein
- Changed codon: same amino acid (degeneracy of code)---> not changing anything
Mutations in protein-coding regions can change an amino acid, truncate the protein, or shift the reading frame

Expanding Nucleotide Repeats

- Large regions of simple repeat sequences present in some genes
--Ex. CAG CAG CAG CAG [...]n--(Gln)
- Can be very unstable and prone to mutation
- Can cause repeat expansion due to strand slippage
- Can cause Huntington Disease
- 6-36 repeats=normal
- 36-39 repeats=at risk
- >39 repeats=HD
- Anticipation- number of repeats correlated with age of onset and severity of disease and tend to worsen with subsequent generations (changes based on a gene)
-- Happens to people in their 50s
-- Once a person crosses a threshold, the protein doesn't fold properly---> neurons die=death

Fragile-X Syndrome

- Normal (29-31 CGG repeats), Mutation (> 200 CGG repeats)
- An excessive number of copies of CGG repeat causes loss of function of gene designated fMR1 (fragile-site mental retardation-1)
- Number of Copies of Repeat shows how chromosome structure affects DNA

Possible Effects of Mutation on Gene Function

- Neutral
- Loss of function/Gain of function= cancer
- Conditional mutation
- Lethal mutation
-- Ex. Siamese cat shows characteristic pattern of pigment deposition- cats' melanin can't fold correctly at high temperatures=conditional mutation

Forward Genetics

- Phenotype---> Mutation
- Ex. Mutant fly---> Discover the gene responsible for red pigmentation (wild type eye color)

Reverse Genetics

Mutation---> Phenotype
- Ex. Create a nonsense mutation and see that it causes no pigmentation in the eye---> knock out a gene to see what happens

What causes the occurrence of mutations?

1) Spontaneous mutations- naturally occurs in cells
2) Induced mutations- external; like chemically induced by mutagens or radiation

Spontaneous Mutations: DNA Replication

- DNA replication is very accurate, less than 1 error in a billion nucleotides
- Not complementary=distortion in helix
- DNA checkpoints fix mutations
- If a mutation get through the checkpoint, then the mutation stays

Spontaneous Mutations: Strand Slippage

Occurs during DNA replication and cause small insertions or deletions

Spontaneous Mutations: Deamination

- Very common in ancient DNA
- CpG islands are "hot spots" for mutation

Spontaneous Mutations: Unequal Crossing-Over

Unequal crossing-over in meiosis I produces insertions and deletions
- If I don't line up recombination correctly, then DNA can be added or deleted---> DNA problem

Induced Mutations: Mutagens

Mutagens- chemicals that can cause mutations in DNA
- Base analogs-chemicals with structures similar to nucleotides. If present, DNA polymerase can't distinguish them and will incorporate in DNA
-- Ex. 5-Bromouracil
- Can measure rate of replication

Induced Mutations: Mutagens Graph

Different Types of Mutations

Polymerase Chain Reaction (PCR)

- PCR= DNA synthesis in a test tube
- Used to make many copies of a lot of copies of a target sequence of DNA
- To study any DNA sequence, there needs to be a lot of it (ex. the ability to measure or detect it with instruments)
- Uses hot and cold temperature to replicate DNA

Primase and DNA polymerases

Made via primer and DNA synthesis respectively
- Gyrase- cuts double-stranded DNA
- Helicase- unwinds DNA
- Origin- replicates DNA

DNA Polymerases can...

Only add dNTPs to an existing free 3'-OH
- Primer is needed to start DNA synthesis
- dNTPs build DNA strand

Components of PCR

DNA synthesis done repetitively in a test tube
- Template DNA
- Taq Polymerase- a thermal stable DNA polymerase
- dNTPs
- Primers (oligos)- short sequences that define outer bounds of the target sequence
-- Used to 'prime' the reaction by providing 3'-OH for Taq
-- Used in pairs (a 'Forward' and a 'Reverse' primer)
- Buffers- contain magnesium
- dNTPs and primers provide opportunities to incorporate a label or another sequence

Three Stages of a PCR Reaction

PCR reaction is cycled through 3 temperatures over and over. For most of the cycles the number of copies of your target DNA is doubled in copy number. (exponential amplification happens 2x)
1. 95°--->Denature- Hydrogen bonds disrupted – ssDNA
2. 55-65°---> Anneal- Primers anneal to target sequence
3.72°---> Elongation- Taq polymerase synthesizes new DNA

Gel Electrophoresis

- Detects fragments
- Used to separate molecules based on their size
- Nucleic acids are 'loaded' into a gel matrix that is in a salt buffer
- An electrical charge is passed through the gel
- Nucleic acids will travel to the positive pole
- Something is added to DNA or gel that allows visualization (like EtBr and radioactive label)---> allows nucleic acids to be seen
- Sample is run alongside a known size standard or "ladder"
- Smaller fragments 'run' faster than larger fragments
-- Fake bands---> not big reactions
-- Large bands---> big reactions

Restriction Enzymes (Restriction Endonuclease)

Cleaves invading DNA
- Isolated from bacteria (viral defense)
- Cut dsDNA in a sequence specific way
-- Usually recognition site is 4 to 8bp long
- Most recognition sequences are palindromic (reads the same backwards and forwards)

Restriction Enzyme Name Comes From The Bacteria They Were Isolated From

Two Types of Cuts Made By Restriction Enzymes

Staggered- or "sticky-" ends and blunt ends

Restriction Enzymes and Recombinant DNA

Cutting different sources of DNA with same 'sticky-end' restriction enzyme allows them to be recombined

Vector

An agent that can carry DNA into a cell or organism
- Many are derived from plasmids, bacteriophages, or viruses

Cloning Vector

Makes lots of the sequence
Plasmid Cloning
- Both DNA you want to 'clone' and the plasmid are cut with the same 'sticky-end' restriction enzyme
- Mix ‘cut’ sample DNA and the
‘cut’ plasmid together with DNA
ligase
- Once target sequence is in plasmid,
plasmid is introduced to bacteria
through transformation

A Typical Plasmid Vector

- MCS- Multiple Cloning Site= where your sequence goes (polylinker site)
- 3 Essential Components of A Vector"
1) MCS
2) ori
3) Selectable marker (amp^R)
4) Screenable marker (Lac+)
-- pUC19=plasmid UC19

Identify Bacteria Containing Both Plasmid and Target DNA Using Selectable (Plasmid) and Screenable Markers (Target DNA)

- Lac Z+- encodes for gene B-galactosidase and contains MCS
- If target DNA is in the plasmid, LacZ gene is disrupted and B-galactosidase is not functional
- After transformation and plating (w/ X-gal & amp^R):
-- All colonies on plate have a plasmid (selected for with amp^R)
- White colonies on plate have target sequence (LacZ disrupted- Xgal not broken-down=no blue)
-- White colonies=Lac Z disrupted---> no B-galactosidase
-- White colonies= can make something is messed up or something worked

Expression Vectors

- All the components of a standard vector plus sequences required for transcription and translation
- Vectors can be used in eukaryotic cells...with the appropriate eukaryotic regulatory sequences

Dideoxy-DNA Sequencing

- Relies on using a "terminator" nucleotide- dideoxyribonucleoside triphosphate (ddNTPs)---> terminates DNA synthesis (No 3'-OH)
- ddNTPs are used at lower molar ratios than dNTPs so full synthesis can occur but sometimes will incorporate a ddNTP and terminate
- ddNTPs has florescent probe---> used to find last base to get synthesis

Components of Dideoxy-DNA Sequencing Reaction

•Template DNA (need lots – PCR or Plasmid)
•Taq polymerase (a thermal stable DNA polymerase)
•dNTPs (deoxyribonucleoside triphosphates)
-- (A’s,T’s, C’s, and G’s - building material for new DNA)
-ddNTPs (dideoxyribonucleoside triphosphates)-- (terminator A,T,C and G’s with some label on it)
•Primer (oligos) – short sequence within your target sequence.
- Used to ‘prime’ the reaction by providing 3’-OH for Taq.
- Only one primer used in each reaction (linear amplification).
•Buffers.

Dideoxy-DNA Sequencing Process

- During DNA synthesis, sometimes ddNTP is incorporated
• At end have fragments differing by one nucleotide each ending with it’s respective ddNTP
• All four samples are run side-by-side (at the same time) and the sequence is read ‘up’ the gel across all four lanes
- Template DNA
1) Complementary DNA is added
2) Add ddNTP
3) Stop at each ddNTP

Didoxy-DNA Sequencing Present-Day

- Historically Sanger sequencing was done in four separate tubes each with a different labeled ddNTP
- Today, all four dideoxynucleotides are used at the same time in the same tube
- Each ddNTP is labeled with a different color fluorescence

Making A Genomic Library

Problem in breaking up DNA fragments
1) Multiple copies of genomic DNA are digested for a limited time
2) Different DNA molecules are cut in different places providing a set of overlapping fragments
3) Each fragment is joined to a cloning vector...
4) ...and transferred to bacteria cells...
5) ...producing a set of clones containing overlapping genomic fragments, some containing the gene of interest
- Bacteria can be spotted on a membrane and screened with a probe to identify candidate clones

Bacterial Colonies With Vectors...

Containing genomic DNA are spotted onto nylon membrane
- Used to see bacterial clones
Then, membranes are screened for sequences of interest through hybridization of radiolabeled probe
-- Probe- DNA complementary to sequence of interest- typically labeled with radiation or fluorescence
- One way to identify molecule of interest
- Put phosphorous in replica filter

Positional Cloning

Complementary probes of the end of one clone are used to find the position of the overlapping next clone
1) Probe complementary to end of clone A used to find overlapping clone B
2) Probe complementary to end of clone B used to find overlapping clone C
3) Probe complementary to end of clone C used to find
overlapping clone D which contains the gene of
interest
- Ex. Chromosome walking
- Eventually find the gene of interest
- Can find a gene of interest using linkage analysis with other mapped genes

FISH

Fluoresence in situ Hybridization
- Using fluorescently labeled DNA probes, can identify cellular or chromosomal location of a sequence of interest
- Requires large stretches of DNA
- Heat up and denature a sample

Gene Expression

Equal to RNA expression analysis
- Gene expression in response to
environment (e.g. therapeutic
drug)
- Gene expression differences in disease
- Tissue specific gene expression
- Developmental stage

Northern Blot Analysis

- Evaluates RNA expression in various tissues
- RNA → Gel →membrane → probed with radioactively
labeled sequence complementary to gene we are investigating
- Finds gene expression
- Disadvantages:
-- Not quantitative- can only judge relative expression levels
-- Can only test one gene at a time

Microarrays

Allows the simultaneous evaluation of expression levels for hundreds or
thousands loci
- DNA attached to solid
support
-- i.e. glass slide, gene chip
- RNA is labeled
-- During cDNA synthesis
- Can have normal expression or have different expression
- Increases through info

Isolate mRNA Using oligo-dT

1) oligo-dT chains are linked to cellulose
2) RNA is isolated and passed through the column
3) mRNA is still in the column and the oligio-dT are paired
4) RNA passes through column
5) mRNA washed via buffer that break H bond between poly (A) and oligo (T) chains
6) Leaving only mRNA with poly (A) tails
- Poly (A) selection enriches our sample for mRNAs

cDNA Synthesis

Makes complementary DNA sequence for the RNA with Reverse Transcriptase
- 1 RNA: 1 cDNA-- use relative abundance of cDNA to represent the relative abundance of RNAs in the cell

1) Reverse transcriptase synthesis cDNA strand
2) After cDNA strand is completed, hydrolyze (mechanically breakdown with water) RNA
-- mRNA is copied bc it's unstable
-- mRNA should only have exons
3) Stays with DNA polyermase to make 2nd DNA strand
4) Terminal transferase adds single-stranded tails
- Double- stranded cDNA is ready for insertion of a cloning vector

Microarrays- Purpose

- Usually comparative- Ratio of fluorescence
between two samples
-- Allows us to compare loci
- Ex.
-- Tumor vs. Normal Tissue
-- Drug vs. No Treatment
-- Embryo vs. Adult
- Only semi-quantitative-- Results from microarray experiments must be validated by a more quantitative essay (like qRT-PCR)

Microarrays- Process

- Many probes on glass slide
- Cells ---> RNA ---> cDNA w/ fluorescent
- Fluorescence Differentiation- tells how much of a gene is expressed
Results-
- Each spot= a gene

Sanger Sequencing vs. Next Generation Sequencing

Sanger Sequencing- Sequence a single target sequence in isolation (~600-1000bp)
- Each sequence needs to be isolated in large
quantities either by cloning in plasmid or
amplifying with PCR
- Need to know a sequence to target with
primers
-- Slow and take a long time

Next Generation Sequencing- Sequencing millions of nucleotides per run
- Does not require laborious subcloning or
PCR
- Thousands to millions of reactions per run
- Allows for sequencing of the entire sample
-- Not laborious and can make many at once

Next- Generation Sequencing

- Simultaneously sequence million of nucleotides at time
- See sample (DNA, RNA, etc)
1) Fragment sample
-- Randomly sheer sample (required length depends on which platform you are using)
2) Ligate adapters
-- Adapters provide a known sequence that you can use to isolate or target with primers
3) Isolate fragments and amplify
-- Amplification is typically some form of PCR and is done to increase signal (from step 4)
4) Synthesize DNA with primers targeting adapter sequence
-- Incorporation of nucleotides detected by detecting chemical change or using some sort of fluorescent signal

Next Generation and 3rd Generation Sequencing Makes $1,000 Genome Possible

- Computational speed, power and storage becomes limiting factors

Next Generation Sequencing- Composition

- Millions of short sequencing reads are assembled into contigs based on sequence similarity
- Short sequencing reads are made into continuous segments based on sequence similarity

RNA-Seq

- After cDNA synthesis, use Next Gen. Sequencer to sequence all of RNA pool--> finds abnormal genes
- Once everything sequenced, align to reference genome and evaluate differential expression levels between samples
- Coverage- range that sequences need to stay between

ChIP-seq (Chromatin Immunoprecipitation)

- Where DNA interacts with proteins, crosslink the DNA to the proteins
- Shear DNA into fragments
- Precipitate protein of interest with antibody for that protein (DNA associated will precipitate too)
- Reverse crosslink and isolate associated DNA
- Sequence recovered pool of associated DNA

Eukaryotic Genes Are Very Repetitive

Human genome
~58% repetitive
~1.5% exonic= proteins
- 2000- 90% of genome was done
- April 2022- Complete genome
- Map-based vs. Shot gun sequencing
-- Today- more shot gun sequencing

Contig Assembly

- Overlap of Restriction Enzyme digest pattern, DNA marker or other known sequence can be used to determine clone order to assemble a Contig
-- Contig= contiguous sequence

Whole Genome Sequencing: Map-Based Approach

- Use detailed genetic and physical maps to identify the least number of clones to cover the genome (minimum tilting path)
- Then sequence and assemble the small number of clones

Whole Genome Sequencing: Whole Genome Shotgun Sequencing

- Shear entire genome into small pieces and clone
- Sequence millions of clones.(e.g. 3.2 billion bp/700bp ≈ 4.5 million sequencing reactions)
- Assemble all those sequences
- Blow up genome---> re-sequence it

Human Genome History

- Human Genome 1st draft completed in 2000
- Completed by both NIH & DOE as well as Celera (In UK Sanger Institute)
- NIH (Francis Collins) used Map-based sequencing
- Celera (Craig Venter) used Shotgun sequencing
- Helps drive the development of sequencing technology

Gene Function

- Roughly 2% of human genome is protein coding
- Only about half has any known function
- Functional genomics- characterizes what genome sequences do, what their function is
- Difficult to identify entire genome

Most Genomic Variation Is In Single Nucleotide Polymorphisms (SNP)

- GWAS: Genome Wide Association Study- uses pedigree and linkage using SNP haplotypes to identify disease loci
~ 4 million nucleotides differentiate humans
-- Differences in genomes are used as markers
- Haplotype is inheritable

Most Genomic Variation Is In Single Nucleotide Polymorphisms (SNP)- Pedigree and Linkage vs. Manhattan Plot

- Uses pedigree and linkage using SNP haplotypes to identify
- Manhattan plot shows genomic regions associated with a trait
- Autism and height associated with loci---> complex traits---> can lead to therapies to help

Other Genome Projects

The Cancer Genome Atlas
- Characterize 20,000 primary cancers with matched normal samples
- 33 cancer types
- 2.5 petabytes of genomic, epigenomic, transcriptomic and proteomic data
- 80% of genome is functional
- 20% is protein coding

ENCODE Project (Encyclopedia of DNA Elements)
- Effort to identify and characterize regulatory sequences
- Trying to figure out letters

Synteny

Co-linearity btwn related genomes
- Genomes are descended from a common ancestor
- Genomes break apart and fuse with different pieces
-- Part of speciation
-- Break locations are reused over time

Homologous Sequences Are Evolutionarily Related

- Genes that are evolutionarily related are homologous (similar to each other bc they're related by descent)
- Paralogs – Homologous genes in the same organism arising by duplication of a single gene in the evolutionary past
- Orthologs – Homologous genes found in different
species that evolved from the same gene in a common ancestor

Genes A1 and A2 are paralogs
Genes B1 and B2 are paralogs
Genes A1 and B1 are orthologs
Genes A2 and B2 are orthologs

Transgenic Organisms

- Injects DNA into pronuclei can result in DNA integrating into genome of organism by nonhomologous
- CRISPR- Cas9- can be used to target specific locations in genome to introduce changes

DNA Fingerprinting- Short Tandem Repeats (STRs)

- Use variations in the genome to identify a person
- AKA microsatellites- known places or variation between individuals

DNA Fingerprinting- Combined DNA Index System (CODIS)

- FBI system of STR Loci to identify people and solve crimes
- Originally used 13 loci (1990)
- Added 7 more loci (2017)

Cancer

A group of disorders character by cell proliferation
- Cells do not respond to normal controls of cell division
- Cells divide rapidly and constantly= tumor growth---> cells become a mass
-- Crowd normal cells and rob nutrients
- Benign tumor- cells remain localized
- Malignant tumor- cells invade other tissue (more dangerous)
- Advanced tumors shed cells that travel to other parts of the body= metastasis

Cell Cycle

- Molecular decision made at Step 3
-- DNA mechanisms need to fix nucleotides

Types of Cancer Causing Genes: Tumor Suppressor Genes

- Tumor suppressor genes- mutated recessive-acting inhibitory genes that cause cancer
-- Both copies of genes are copied
-- Like brakes in a car---> bad but can work if there's only one

Types of Cancer Causing Genes: Oncogenes

- Mutated dominant-acting stimulatory genes that cause cancer
-- Proto-oncogene- oncogene prior to mutation
-- Like gas in a car
-- Very quick and excessive
-- Contributes to cell growth

Types of Cancer Causing Genes: DNA Repair Genes

Mutations in DNA-repair genes can increase
the likelihood of acquiring mutations in these genes

Knudson's Multistep Model of Cancer (1971)

- Explains retinoblastoma
- Unilateral Sporadic
-- No family history
-- Very rare
-- Mutations are less common
- Bilateral Familial
-- Cancer in both eyes
-- Very common
-- Mutations are more common
- Cancer is multistep- if one or more required mutations are inherited, less mutations would be required to produce cancer

Clonal Evolution of a Tumor

Most tumors arise from somatic mutations that accumulate in a person's life span
1) 1st mutation cell predisposed to proliferate faster
2) 2nd mutation causes cell to divide rapidly
3) 3rd mutation cell undergoes structural changes
4) 4th mutation causes cell to divide uncontrollably and invade other tissue
- Rate of clonal evolution depends on the frequency where new mutations arise

Other Genes Invoved

- Telomerase- activation of telomerase allows cells to divide indefinitely
-- Hits other genes
- Apoptosis genes- genes involved in signaling ‘programmed cell death’ are often mutated
-- Other genes are mutated
- Vascularization genes- genes that stimulate angiogenesis are often overexpressed
-- Tumor cells reactivate vascularization cells
- Cell adhesion genes- genes involved in intercellular interactions and attachment
-- Don't stick together well bc of leaky blood vessels---> causes problems due to metathesis
- miRNAs- microRNAs are often mis-regulated

Cell Cycle Genes are Mutated In Cancer (Ex. Retinoblasma---> G1/S Checkpoint)

- Kinase- an enzyme that adds a phosphate to a protein
- Cyclin-dependent kinases (CDKs)- kinases that control key events of the cell cycle
-- CDKs are functional only when associated with a cyclin (another protein)
---Cyclin determines proteins
-- Each cyclin appears at specific points in the cell cycle
-- When bound to a CDK, which cyclin is present will determine which protein the CDKs phosphorylate

Retinoblastoma Protein Helps Control the Progression Through G1/S Checkpoint

1) RB binds E2F and keeps it inactive
-- Sequesters E2F---> prepare cell for S phase
-- Cyclin-D--CDK and Cyclin-E--CDK starts to become more abundant
2) Increasing conc. Of cyclin-D-CDK
and cyclin-E-CDK phosphorylate RB
-- E2F available
3) Once phosphorylated, RB is inactive and releases E2F (a TF)
-- Go to S phase
4) E2F binds DNA and stimulates
transcription of DNA replication genes
- RB= tumor suppressor

Loss of Heterozygosity

- People heterozygous for a tumor-suppressor gene are predisposed to cancer
- Loss of heterozygosity often leads to cancer in a person heterozygous for a tumor-suppressor gene

Cell Cycle Progression Is Regulated By External Factors

- Hormones and growth factors are unable to pass through the cell membrane + Typically bind to cell surface receptors that transmit the message into the cell = Signal transduction pathway
- Ex. Ras signal transduction pathway
- Inactive Ras binds guanosine diphosphate (GDP)
- Active Ras binds guanosine triphosphate (GTP)

Ex. Ras Signal Transduction Pathway

Mutates ~800x
- Binding of growth factor causes conformation change and phosphorylation
- Adaptor molecules bind to receptor and link to Ras
- Ras binds GTP and is activated

Ex. Ras Signal Transduction Pathway- Cascade of Reactions

Cascade of reactions---> activated MAP kinase moves to nucleus and activates transcription factors that stimulate transcription of cell cycle genes

Clonal Evolution of Colorectal Cancer

- Loss of normal tumor- suppressor gene APC
1) Polyp (Small growth) forms on colon wall
2) A benign, precancerous tumor grows
- Activation of oncogene RAS
3) An adenoma (benign tumor) grows
- Loss of tumor-suppressor gene p53
4) A carcinoma (malignant tumor) develops
- Other changes; loss of anti-metastasis gene
5) The cancer metastasizes (spreads to other tissue through bloodstream)

Many Cancers Have Recurring Chromosomal Abnormalities

Burkitt's lymphoma
- Has recurring reciprocal translocation btwn q-arms of chromosomes 8 &14

Many Cancers have Recurring Chromosomal Abnormalities

- Philadelphia Chromosome- BCR-ABL1 fusion protein
-- Chronic myelogenous leukemia
- Spectral Karyotyping Fluorescence in situ hybridization (SKY FISH)

Many Cancers Have Chromosomal Abnormalities- Karyotype of HCC38 Cells

Derived from breast cancer
- Hypertriploid
- 37 structural abnormalities involving all chromosomes except 6 and 16
- And more...

Cancer Types

- Occurs in many tissue types
- Public health issue
- Model to study cell cycle control

Cancers Are Influenced by Environmental Factors

Hong Kong vs. Salt Lake City, Utah

Viruses Are Associated With Some Cancers

Virus----> Cancer
- Human papilloma viruses (HPVs)---> Cervical, penile, and vulvar cancers
- Hepatitis B Virus---> Liver cancer
- Human T-cell leukemia virus 1 (HTLV-1)---> Adult T-cell leukemia
- Human T-cell leukemia 2 (HTLV-2)---> Hairy-cell leukemia
- Epistein-Barr Virus---> Burkitt lymphoma, nasopharyngeal cancer, Hodgkin lymphoma
- Human Herpes Virus---> Kaposi sarcoma
- Merkel Cell Polyomavirus---> Merkel Cell Carcinoma

Mendel's Rules of Inheritance Dealt W/ Qualitative Traits

- Looks at discrete traits
- Genotypic ration- 1:2:1
- Phenotypic- 3:1

Simple Single-Gene Inheritance

Discontinuous Characteristic vs. Continuous Characteristic

- Discontinuous characteristic- allows predictions about genotypes and genetic cross
- Continuous characteristic- are polygenic and influenced by the environment= multifactorial

Quantitative Characteristics

Environmental factors often influence quantitative characteristics
- Can be difficult to see which phenotype to use
- If a trait is polygenic:
-- Many genotypes are possible
-- Several genotypes might produce the same phenotypes
-- Often environmental factors influence phenotype
- Quantitative genetics- genetic analysis of complex characteristics
- Quantitative trait loci (QTL)- Chromosomal regions contain genes that influence a quantitative trait
-- Region produces variation
- Knowing variation can determine loci, genes, etc.