Chapter 11 Genetics

Lectures by Kathleen FitzpatrickSimon Fraser University

Copyright © 2012 Pearson Education Inc. Mark F. Sanders John L. Bowman

G E N E T I CA N I N T E G R A T E D A P P R O A C H

A N A LY S I S Chapter 11Chromosome Structure

Erin Kelleher2/25/14

http://www.mediaresource.org/instruct.htm

Copyright © 2012 Pearson Education Inc. Genetics Analysis: An Integrated Approach

11.1 Bacterial and Archaeal Chromosomes Are Simple in Organization

• Bacteria have single chromosomes that are almost always circular. However, some species have linear chromosomes, and some species have more than one chromosome

• When there are multiple chromosomes, the largest chromosome generally harbors the essential genes

• Plasmids – extrachromosomal circular DNAs that exist in more than one copy and carry non-essential genes (antibiotic resistance genes are common 2


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Bacterial and Archaeal chromosomes are compacted into the nucleoid

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How does this process occur?1) Proteins help package the DNA2) Supercoiling – DNA becomes tightly wound


Proteins Associated with Chromosomes

• Small nucleoid-associated proteins participate in the DNA bending that contributes to folding and condensation of chromosomes

• Structural maintenance of chromosome (SMC) proteins: these attach directly to the DNA, holding it in coils or V-shapes to form large nucleoprotein complexes

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Supercoiling

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11.2 Eukaryotic Genomes

• Multiple chromosomes, always linear

• Proteins are involved in organizing, condensing and segregating chromosomes

• The DNA and associated proteins of a chromosome are called chromatin - each chromosome is approximately half DNA and half protein

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Eukaryotic Chromosome Shapes


Chromatin Composition

• Each chromosome is approximately half DNA and half protein

• About half of the proteins are histone proteins, small basic proteins that tightly bind DNA

• The remaining proteins, the nonhistone proteins, are very diverse and perform a variety of tasks in the nucleus

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Histones


Nucleosome Assembly

• Histones H2A and H2B assemble into dimers; H3 and H4 also form dimers

• Two H3-H4 dimers form a tetramer, after which two H2A-H2B dimers associate with it to form the octamer

• The wrapping of DNA around the nucleosome is the first level of DNA condensation, and compacts the DNA about sevenfold

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Chromatime that is only packaged by nucleosomes resembles beads on a string


Higher Levels of Chromatin Compaction

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Highest order packaging, interphase and metaphase

interphase

metaphase


Higher-Order Chromatin Condensation

• Chromatin loops of 20 to 100 kb are anchored to the chromosome scaffold by nonhistone proteins at sites called MARs (matrix attachment regions)

• The radial loop-scaffold model suggests that the loops gather into “rosettes” and are further compressed by nonhistone proteins

• Metaphase chromatin is compacted 250-fold compared to the 300-nm fiber

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The Radial-Loop Scaffold Model

• Chromatin loops in 20,000- 1000,000 bp

• Chromatin is packaged into loops by non-histone scaffold proteins that attach to Matrix Attachment Regions (MARs)


Roles of Higher-Order Chromatin Condensation

• Chromosome compaction allows for efficient separation of chromosomes at anaphase

• The chromatin loops formed during condensation play a role in the regulation of gene expression

• Active transcription takes place in segments of loops distant from MARs; thus larger loops have more active transcription than small loops

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Heterochromatin and Euchromatin

• Chromosome condensation varies from one part of a chromosome to another

• Regions that contain actively expressed genes and are less condensed during interphase are called euchromatin

• Regions that remain condensed in interphase and contain many fewer expressed genes are called heterochromatin

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Types of Heterochromatin

• Facultative heterochromatin exhibits variable levels of condensation, related to levels of transcription of resident genes

• Constitutive heterochromatin is permanently condensed, found prominently in centromeres and telomeres, and composed primarily of repetitive DNA sequences

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Centromeres exhibit unique packaging

Normal nucleosome Centromeric nucleosome

The N-terminal tail of CENP-A allows the binding of kinetochore proteins to the centromere


Centromeric nucleosomes facilitate attachment of the kinetochore

Centromeric nucleosome


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Budding yeast centromeres have consensus sequences that recruit centromeric nucleosomes

CDE I CDE II CDE III


Budding yeast are weird

• Centromeric DNA sequences of all other eukaryotes are highly repetitive and constitutively heterochromatic

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Variable Chromatin Condensation and Chromosomal Banding

• Densely packed heterochromatin absorbs more stain and creates dark bands.

• Loosely packed euchromatin absords less stain and creates light bands

• Giemsa is the most commonly used stain so we call these “G-Bands”


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Banding and Chromosome StructurePrior to DNA sequencing, banding patterns allowed scientists to reliably identify chromosomes and define regions within those chromosomes


Karyotyping• Reliable identification of

chromosomes also allows for us to identify aneuploid individuals – those with more or less than two copies of each chromosome

• Most common human aneuploidies• Down syndrome – trisomy 21• Trisomy 18 and 13• XXX, XO, XXY


Fluorescent In-Situ Hybridization (FISH), chromosome painting


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Chromosome painting – an alternative strategy to identification by G-band


Structural Rearrangements in Human Cancers


Nucleosome Distribution and Synthesis During Replication• Nucleosomes interfere with DNA replication via DNA

polymerase.

• Therefore, nucleosomes must be removed from the DNA to allow for replication, then quickly replaced after DNA synthesis is complete

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Nucleosomes and Gene Expression

• Nucleosomes also interfere with the recruitment of RNA polymerase to initiate transcription.

• chromatin remodeling – nucleosomes are displaced to expose promoter and other regulatory sequences

• Chromatin remodeling relies on chemical modifications to histones in nucleosomes are epigenetic marks or epigenetic modifications

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Histone Modifications

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Acetyl groups

- histones are modified at their N-terminus

- methyl groups generally confer more compact chromatin that reduces gene expression

- acetyl groups generally confer more open chromatin that increases gene expression


Epigenetics and Histone Modification

• Epigenetic – a heritable change in gene expression that is NOT caused by a change in DNA sequence

• Histone modifications are epigenetically heritable because they can be transmitted through cell division and across generations

• The retention of some old histones during DNA replication provides a mechanism for maintaining the modifications and passing them to daughter cells

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Important Features of Epigenetic Modification

1. Alters chromatin structure

2. Transmissible during cell division

3. Reversible

4. Directly associated with gene transcription

5. Does not alter DNA sequence

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Drosophila as a model for chromatin modification

• Position effect variegation – genes that are adjacent to heterochomatic regions exhibit variable expression due to variable spreading of heterochromatin

• To identify proteins involved in chromatin packaging, scientists look for genes that modify position effect variegation

• Mutations of these genes enhance or suppress the variegating phenotype, E(var) and Su(var)


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Su(var) and E(var) Mutations

• Several dozen E(var) and Su(var) mutations have been identified in Drosophila

• Genetic analysis supports the hypothesis that chromatin is dynamic and associated with gene expression

• Genes have been identified that encode proteins that make epigenetic marks on histone proteins (adding methyl, acetyl, and phosphoryl groups)

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Su(var) genes, HP-1 and HMT

• Methylated histone locations (e.g., H3K9me) are methylated by HMTs

• These act as sites of HP-1 binding, and help condense chromatin structure and silence gene expression

• Mutations in either gene lead to failure to remodel chromatin to a condensed state, and thus suppress variegation

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Wild-type Cell

Su(var) Mutant Cell


Chromatin remodeling and X-chromosome inactivation

Random X-inactivation is a mechanism of dosage compensation, which equalized the expression of sex-linked genes between males and females


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XIST- X inactivation specific transcript

- XIST is expressed only from the X-chromosome that was randomly chosen for inactivation

- XIST coats the X and recruits HMTs

- Methylated histones recruit HP1 and forms a silenced X-chromsome (Barr body)


Eukaryotic Interphase Chromosome Territories

Chromosome territory: a 3D space in the nucleus that is occupied by individual chromosomes

Interchromosome domain: channels for movement of proteins, enzymes, and RNA molecules


Dynamic Chromosomes

• Chromosomes do not occupy the same territory in each nucleus, but once confined to a territory, a chromosome does not leave until the M phase is initiated

• However, chromosomes are active within their territories and move, twist, and turn during transcription and DNA replication

• Chromosomes appear to be anchored in their territories by their centromeres

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Centromeric clustering in yeast

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Chapter 11 Genetics