Silence of the Genes

Genetics

• The study of inheritance

Gregor Mendel(1823-1884)

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Mendelian Ratios

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Mutation

ATGCGAGCGAGTATGCGATCGAGT Genotype

Phenotype

Epigenetics

• Heritable changes in gene expression that do not involve changes in DNA sequence.

Epi-mutation

ATGCGAACGAGTATGCGATCGAGT Genotype

Phenotype

DNA Modifications

Histone Modifications

Proteins

Nucleosome

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DNA

histones

DNAMethylation

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histones

CH3

HistoneMethylation

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histones

CH3

Examples of Epigenetics

X-Inactivation

inactivation

male female

X Y X Xunequal expression

equal expressionX Y X X

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Barr Body

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Calico Cat

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X Xo O

X Xo O X Xo O

orangeallele

blackallele

black sector orange sector

Other Examples of Epigenetics

Imprinting

• Imprinted genes are expressed differently depending on whether they are inherited through the maternal or paternal parent.

Horses and Donkeys

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male female

female

Gynogenotes

• Embryos containing two female genomes

– do not develop normally– fail due to underdeveloped extraembryonic

placental tissue

Androgenotes

• Embryos containing two paternal genomes

– result in abnormal (often overgrown) embryo– display overdeveloped extraembryonic

placental tissue

Maternal vs. Paternal Imprinting

• Male genome wants to promote growth.

• Female genome wants to inhibit growth.

Resetting Methylation Patterns

Primordial Germ Cells

male

female

met

hyla

tion

developmental time

Battle for Maintaining DNA Methylation

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Battle to Maintain Methylationm

ethy

lati

on

developmental time

Fertilization

Female Strategy

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Genes that promote growth

Fertilization

Male Strategy

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Antisense

Genes that inhibit growth

Fertilization

Resetting Imprints is Important for Proper Development

Dolly

• First mammal to be cloned from an adult cell.

• Developed illness common in older sheep (arthritis)

• Probably due to abnormal imprinting

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Gene Silencing

HeterochromatinDensely staining condensed chromosomal regions; believed to be transcriptionally inert.

EuchromatinA chromosomal region that stains normally; thought to contain the normally functioning genes.

Heterochromatin

Paul Fransz

B. McClintock

IV

Heterochromatin

• Centromeric regions

• Telomeric regions

Position Effect Variegation

W+

W+

W+

W+

heterochromatin

spreading

suppressed

enhanced

Post-Transcriptional Gene Silencing

Anti-Sense

Sense Also Works?

Guo and Kemphues, (1995)

What is Causing Silencing?

?

dsRNAPost Transcriptional

?

Fire et al. (1998)

RNAi

?

Fire et al. (1998)

RNAi

AAA

AAA

Dicer

RISC

dsRNA

AAA

AAA

Dicer

RISC

dsRNA

RNAi

RNA Silencing

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cosuppression

quelling

RNA Silencing

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Post TranscriptionalGene Silencing

(PTGS)

RNA Silencing

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RNA Silencing

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RNA Interference(RNAi)

RNA Interference

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DNA Methylation

Mette et al. (2000)

RNAi and Transcriptional Silencing

Transcriptional Silencing

Mette et al. (2000)

Micro RNAs

Grishok et al. (2001)

dicer

RNAi Genes in S. pombe?

RNAi Genes in pombe?

• Yes!!

• Mutation of these genes results in loss of centromeric silencing.

• Loss of RNAi reveals centromeric transcripts.

• dsRNA from centromere targets transcriptional silencing.

Cen

Cen

RNAi

Small RNAs

Cen

RNAi

SilencingMachinery Small RNAs

Cen

RNAi

SilencingMachinery

Small RNAs

Cen

RNAi

• Is important for initiation and maintenance of heterochromatin at the centromere.

• Could be involved in other silencing phenomena

Where else may RNAi be functioning to silence genes?

X Chromosome Inactivation(RNAi?)

Xist

Tsix

Heard et al (2001)

Xist

Tsix

Heard et al (2001)

RNAi

SilencingMachinery

X Chromosome Inactivation(RNAi?)

Xist

Tsix

Heard et al (2001)

RNAi

SilencingMachinery

X Chromosome Inactivation(RNAi?)

Imprinting(RNAi?)

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Antisense

RNAi

SilencingMachinery

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Imprinting(RNAi?)

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Antisense

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Imprinting(RNAi?)

RNAi

SilencingMachinery

HeterochromatinDensely staining condensed chromosomal regions; are not necessarily transcriptionally inert. RNAi is important for its initiation and maintenance.

EuchromatinChromosomal regions that do not densely stain; thought to contain functioning genes that maybe transcriptionally active.