Chromatin Structure Study Notes

Chromatin Structure Overview

Chromatin Folding Hierarchies

  • Different structural levels of chromatin organization:

    • 2 nm: DNA double helix

    • 11 nm: Nucleosome units ("beads on a string")

    • 20-30 nm: Chromatin fiber

    • 700 nm: Fiber loops (higher-order structures)

    • 1400 nm: Mitotic chromosome

Domains of Chromatin

  • Topologically Associated Domains (TADs): Functional units within chromatin that are spatially organized.

  • Compartmentalization: Variation between heterochromatin (gene-poor, transcriptionally inactive) and euchromatin (gene-rich, transcriptionally active).

  • Chromosome Territories: Distinct regions within the nucleus designated to each chromosome, which prevents intermolecular entanglements.

Structural Proteins in Chromatin

  • Cohesin: Complex involved in holding sister chromatids together.

  • Condensin: Important for chromosome condensation during cell division.

Intended Learning Outcomes

  • After studying the lecture, you should be able to:

    1. Explain the Role of Chromatin in Gene Expression: Understand why chromatin is crucial in eukaryotic gene expression.

    2. Describe Nucleosome Features: Identify the structural and functional characteristics of nucleosomes.

    3. Relate Genome Organization to Gene Expression: Understand how genome organization affects gene expression.

    4. Propose Experimental Approaches for Nucleosome Analysis: Suggest methods to analyze nucleosome distribution in the genome.

Biological Model Organisms and Their Genomes

  • Comparison of various organisms:

    • T4 bacteriophage:

    • Phenotype: 200 nm length

    • Genome size: 169,000 bp

    • Number of genes: 289

    • E. coli:

    • Phenotype: 2.0 μm length

    • Genome size: 4,600,000 bp

    • Number of genes: 4,288

    • S. pombe (fission yeast):

    • Phenotype: 10 μm length

    • Genome size: 14,100,000 bp

    • Number of genes: 4,970

    • Drosophila melanogaster (fruit fly):

    • Phenotype: 2.5 mm length

    • Genome size: 165,000,000 bp

    • Number of genes: ~13,000

    • Homo sapiens (human):

    • Phenotype: 1.75 m height

    • Genome size: 3,200,000,000 bp

    • Number of genes: ~25,000

    • Arabidopsis thaliana (plant):

    • Phenotype: 20 cm height

    • Genome size: 119,000,000 bp

    • Number of genes: ~26,000

Operational States of Genomes

Bacterial Genomes: Default "On" State

  • In bacteria, operons are often default active:

    • 5' + glucose + lactose: operon off

    • 5' - glucose - lactose: operon off

    • 5' + glucose - lactose: operon off

    • 5' - glucose + lactose: operon on

  • Main components:

    • CAP: Catabolite activator protein (apo-activator when not bound)

    • Lactose-bound lac repressor: inhibits transcription

    • Cyclic AMP-bound activator: enhances gene expression at weak promoters

    • lac operator: Sites where repressors and activators bind to regulate transcription.

Eukaryotic Genomes: Default "Off" State

  • Eukaryotic genes are typically repressed due to:

    • Hidden promoters: Not easily accessible

    • Inaccessible cis-regulatory motifs: Require activation before expressing genes.

Chromatin: Definition and Importance

  • Definition: Chromatin is the material that forms eukaryotic chromosomes, composed of DNA and protein.

    • Origin of the term: "Chromatin" comes from the Greek word "chroma" meaning color, due to the staining properties observed by Walther Flemming in 1882.

  • Function: Chromatin maintains DNA in a transcriptionally competent form for replication, segregation, and repair.

Facts About Chromatin

  • Each human cell contains ~2 meters of DNA.

  • The human body has approximately ~10 trillion (10^13) cells.

  • A single genome can produce about ~230 different cell types, which is essential for healthy development. Loss of this cellular identity can lead to diseases.

Evolution of Histone Proteins

Common Ancestry of Histone Proteins

  • Histone fold proteins share a common evolutionary ancestor found in both archaea and eukaryotes:

    • In archaea, the basic repeating unit is a histone fold tetramer that wraps about 60 bp of DNA.

    • Functions similarly to the nucleosome in eukaryotes.

    • References include Ammar et al. (2012), demonstrating chromatin as an ancient and conserved innovation.

Conservation of Histone Proteins

  • Histone proteins are highly conserved across species:

    • Example: The H4 histone sequence with distinct substitutions among species including:

    • S. pombe (fission yeast)

    • H. sapiens (human)

    • X. laevis (African clawed frog)

    • S. cerevisiae (bakers yeast)

    • M. musculus (mouse)

    • D. melanogaster (fruit fly)

Structure and Function of Histones

Histone Fold Domains

  • Histone fold domains serve as a structural framework for DNA binding and organization:

    • Dimerization occurs via a "histone handshake" to form tetramers that organize 60 bp of DNA.

    • The formation of tetrasomes as subnucleosomal particles for genomic packaging in archaea.

Organization of the Nucleosome

  • Nucleosomes are organizational units that compact DNA:

    • The histone octamer consists of H3 and H4 tetramers surrounded by H2A and H2B dimers, organizing 145-147 bp of DNA in a left-handed supercoil.

  • Notable motifs involved in DNA bending include the ancient helix-strand-helix motif.

  • Histone fold domains provide a positively charged path for bending DNA by inserting arginine residues into the DNA minor groove.

Linker Histones

  • Role of Linker Histones (H1): Regulate chromatin accessibility and consist of a folded globular domain with extended N- and C-terminal tails.

    • Binding Region: H1 binds between the entry and exit DNA of nucleosomes, influencing gene expression regulation.

Nucleosomal Organization Patterns

Nucleosomes Around Promoters and Terminators

  • Nucleosomes are strategically placed:

    • They are not randomly distributed, rather clustered around promoters and terminators.

    • Nucleosome-free regions (NFRs) exist before transcription start sites and after termination sites for accessibility.

  • Specific patterns of nucleosome positioning can be observed based on the gene structure, enhancing gene expression regulation.

Experimental Tools for Analyzing Chromatin

Probing Open Chromatin

  • Tools such as endonucleases (e.g., DNaseI) and free radicals help analyze DNA accessibility.

  • Analysis methods include:

    • Gels

    • Microarrays

    • Next-generation sequencing

Techniques Utilized

  • ATAC-seq: A modern technique for studying chromatin accessibility. Utilizes transposase to oxidatively tag regions of open chromatin for sequencing analysis.

Mid and Higher Order Chromatin Structure

Folding of DNA into Chromosomes

  • The hierarchical folding of DNA into complex structures:

    • From 2 nm DNA to 700 nm chromatin fibers to 1400 nm mitotic chromosomes.

Fractal Structure of Chromatin

  • Chromatin exhibits fractal-like properties:

    • Resembling patterns that are consistent across scales, embodying the principles of fractals.

Summary of Chromatin Structure

  • Genomes need to be efficiently managed and protected due to their extensive lengths.

  • Histones play a crucial role in compacting DNA into nucleosomes, which neutralizes the charge and regulates accessibility for transcription and replication processes.

  • Higher-order structures arise through fractal loops and domains, enabling proper gene expression regulation.