Noncoding sequencesLevels of packingGene regulationChemical Modifications

Organization of the Eukaryotic Chromatin Structure

Human Genome20,000-

30,000 genesmost of the

DNA does NOT encode for a protein or RNA (is not a gene)

http://upload.wikimedia.org/wikipedia/commons/6/64/Components_of_the_Human_Genome.jpg

Coding & Noncoding Sequences

http://www.hhmi.org/biointeractive/dna/DNAi_coding_sequences.html

Classes of Noncoding Sequences

IntronsRegulatory

sequencesHighly repetitive

sequences:Tandemly repetitive

DNA (tandem repeats)

Interspersed repetitive DNA

Telomeres

Noncoding DNA

Tandemly Repetitive DNA (e.g. VNTRs)

Interspersed Repetitive DNA

(e.g. transposon)

Repetitive DNA

Noncoding DNA

Introns Regulatory sequences

Telomere

Tandemly Repetitive DNAShort identical DNA sequences repeated

in seriesMany types, often classified by length of

the repeatVariable Number Tandem Repeats

(VNTRs)Number of repeat variesSequence can be repeated different

number of times

Tandemly Repetitive DNA ApplicationUseful for DNA Fingerprinting and

studying evolutionOSC Lab: VNTR at the D1S80 locus

Length of repeat: 16 bpRepeated sequence: GTGGTCCTCCTTCCTG Number of repeats: 14-41Total length: 200-700 bp

Example: Fragile X SyndromeFirst triplet repeat identified was the one

in Fragile X syndromeRepeat sequence: CGG triplet on X

chromosomeNormal fragile X allele: repeated 30x in

the 5’ untranslated region of the first exon

Fragile X syndrome: repeat is over 100xThe most common single known cause

of intellectual disability and autism

Example: Huntington’s Disease

Neurodegenerative disorder that affects muscle coordination and leads to cognitive decline and psychiatric problems

Repeat sequence: CAG Individuals affected with

HD typically have at least 36 repeats or greater.

Repeats

Disease

< 27 -27 – 35 -36 – 39 + / -> 39 +

Repeat Size

Median Age at Onset * (years)

39 66 (72-59)40 59 (61-56)41 54 (56-52)42 49 (50-48)43 44 (45-42)44 42 (43-40)45 37 (39-36)46 36 (37-35)47 33 (35-31)48 32 (34-30)49 28 (32-25)50 27 (30-24)

*Age by which 50% of individuals will be affected

Specific numbers of CAG repeat correlates to disease. Walker FO (2007). "Huntington's disease". Lancet 369 (9557): 218–28. Normal HD gene CAG repeats range from 10 - 27 repeats. A few normal individuals have intermediate HD gene CAG repeats of 27-35 repeats. A current review of 1,049 persons (the majority of whom were symptomatic) has provided a determination of the likelihood of an age-of-onset for a given CAG repeat size for repeats between 39 50 repeats (Brinkman et al., 1997; Am. J. Hum. Genet. 60:1202-1210).

Comparing Prokaryotic and Eukaryotic DNA

Prokaryote EukaryoteDNA Circular LinearLocation of DNA

Not in nucleusNucleoid region

Contained in nucleus

DNA bound with proteins?

No. Naked. Yes

Chromosome Structure2 arms, divided at

centromere:p arm – petit armq arm – long arm

Centromeres:region where sister

chromatids are connectedmade up of repetitive

sequences

Activity: DNA PackingMaterials:

ThreadScissorspill covers

Instructions:Cut out 20 m of threadPut the thread in the pill cover

DNA Packing AnimationTry to ignore the strange music

http://www.youtube.com/watch?v=N5zFOScowqo

Levels of DNA packing1st level: nucleosome2nd level: solenoid /

chromatin3rd level: looped domain4th level: metaphase

chromosome

http://www.nature.com/nri/journal/v3/n11/images/nri1225-i2.jpg

HistonesProteins around which DNA coils to form

chromatinContain positively charged R-groups which

bind to the negatively charged DNA (phosphate groups)DNA backbone has a negative chargehistones have a positive charge

5 typesCore: H2A, H2B, H3, H4Linker: H1

First level of packing: NucleosomeNucleosome = DNA +

core histonesDNA wrapped twice

around an octamer of core histones consisting of:2 of each core histone:

H2A, H2B, H3, H4Note: H1 is not part of

the nucleosome, but is attached to the DNA near the nucleosome

10 nm in diameterhttp://biology.kenyon.edu/courses/biol114/Chap01/chrom1.gif

http://bio.research.ucsc.edu/people/boeger/images/research%20interest%20images/nucleosome%20bigger.jpg http://bioweb.wku.edu/courses/Biol566/Images/NucleosomeF09-30A.JPG

Beads-on-a-stringA nucleosome looks

like a bead (histone) with string wrapped around it (DNA) Beads-on-string

Consists of histone octamer and DNA

http://219.221.200.61/ywwy/zbsw(E)/pic/ech9-19.jpg

http://www.nature.com/nri/journal/v3/n11/images/nri1225-i2.jpg

SUPERCOILING BEGINS!!!

http://www.bio.miami.edu/dana/104/nucleosome.jpg

Second level of packing: Solenoids / chromatin

Solenoid: H1 histones aggregate causing 6 nucleosomes to coil together

30 nm in diameterstack on top of each other

forming the chromatin fiber

http://www.nature.com/nri/journal/v3/n11/images/nri1225-i2.jpg

Third level of packing: looped domainsChromatin fibers forms

loops which attach to nonhistone proteins

Nonhistone proteins form a scaffold

300 nm in diameter

http://bioweb.wku.edu/courses/Biol566/Images/ChromatinF09-35.JPG

http://www.nature.com/nri/journal/v3/n11/images/nri1225-i2.jpg

Fourth level of packing: Metaphase ChromosomeLooped domains coil and fold, further

compacting chromatinResult in the characteristic metaphase

chromosome seen in karyotypesSuch packing occurs during mitosis

DNA Packing Video

http://www.hhmi.org/biointeractive/dna/DNAi_packaging_vo2.html

http://www.dnai.org/c/index.htmlhttp://www.youtube.com/watch?v=bwVjYxcDQ5I

TutorialDiscovery of chromosome packaging

http://www.dnaftb.org/dnaftb/29/concept/index.html

Gene Regulation TopicsReview cell cycleMetaphase chromosome in mitosisInterphase chromatin in G1, S, G2

DNA replicationtranscription

Gene Regulation DefinedThe ability to control whether a gene is

actively being transcribed or notThis regulation can occur at the:

Structural level (e.g. supercoiling of DNA)Molecular level (e.g. negative feedback to

suppress enzymatic activity)

http://www.biologycorner.com/resources/cellcycle2.jpg

Cell Cycle

SupercoilingFor a gene to be expressed

(transcribed), the chromosome at that region must be uncoiled

Different levels of supercoiling affects transcription

Allows control over which genes are expressed

Mitosis: PMAT

http://upload.wikimedia.org/wikipedia/commons/4/4b/Chromatin_Structures.png

DNA is “inactive” because it is in the most condensed form (metaphase chromosome)

thus information is inaccessible

Interphase ChromatinMuch less condensed than during mitosisScaffold may be less defined (e.g. attach

to inside of nuclear envelope)But order still exists:

Each chromosome occupies a restricted area with the nucleus

Nucleosome structure stays intact throughout cell cycle

Most still maintain the solenoid and looped domain structures

Interphase Chromatin Different levels of

packing on various regions of the same chromosome

Euchromatin: loosely packed region on chromatin, active transcription

Heterochromatin: densely packed region on chromatin, inactivated

Eu = true; Hetero = different

Chromatin Structure during Transcription

http://upload.wikimedia.org/wikipedia/commons/4/4b/Chromatin_Structures.png

http://upload.wikimedia.org/wikipedia/commons/4/4b/Chromatin_Structures.png

MaCS only

The following sections are for the MaCS grade 12 biology class only

EpigeneticsStudy of heritable

changes in gene expression caused by mechanisms other than the DNA sequence

Modifications often affect both chromatin structure and gene regulation

Modifications include:DNA methylationHistone acetylation

http://cerch.org/wp-content/uploads/2011/02/DNA-methylation-image.jpg

Methylation

Attachment of methyl groups (-CH3) to DNA bases, usually at cytosine

Occurs after DNA is replicated (S-phase) In general, methylation occurs on genes

that are not expressedMethylation suppresses gene expressionRemoving methylation turns genes “on”

Histone Acetylation

Attachment of acetyl group (-COCH3) to histones

Acetylation changes histone shape reducing its grip on the DNA

Results in more loosely packed DNA, available for transcriptionAcetylation activates gene expressionDeacetylation suppresses gene expression

Summary of Modifications affecting chromatin structure and gene regulation

http://cnx.org/content/m26565/latest/graphics35.jpg

Summary of Modifications affecting chromatin structure and gene regulation

http://cnx.org/content/m26565/latest/graphics35.jpg

Genomic ImprintingImprinting: suppression of genes so that only

1 allele is expressed Established in germline cellsMaintained in all somatic cells derived from the

parent cellMost genes are not imprinted: both alleles

are expressedImprinted genes:

Expressions regulated by epigenetic factorsInactivated gene can be either maternal or

paternalGenes are expressed from the non-imprinted

allele

http://www.fluorous.com/images/epigenetics.jpg

Epigenetics Videos PBS NOVA Now “Epigenetics” (13:03)

http://www.youtube.com/watch?v=7WEHoCA1hpo

BeginBeforeBirth “What makes us who we are?” (4:11)http://www.youtube.com/watch?v=9AfBsTAQ8zs

BBC Horizon “The Ghost in Your Genes” (49:06)

http://www.youtube.com/watch?v=Q6cQSR3mPm8