Histone Modification

neurobehavioural epigenesis

from gene expression to phenotype

gene expression to cell/synapse to neural circuitry to behavioural phenotypes

chromatin

physiological template of our genome

represents a structural solution for maintaining and accessing complex genomes

Aristotle (384-322 BC)

Greek philosopher who established scientific methods for investigation and reasoning (termed natural philosophy).
Provided a theory on how embryos generate and develop.
According to him, an organism develops gradually from undifferentiated material

William Harvey (1578-1657)

English physician, provided experiments and arguments for the systemic circulation and properties of blood being pumped to the brain and body by the heart.

~1650, coined the term ‘epigenesis’ to describe development as a gradual
process of increasing complexity from initially homogeneous material in the egg (omne vivum ex ovo)

Caspar Friedrich Wolff (1733-1794)

German physiologist. During this period, the preformation theory was common (homunculus in sperm). However, he provided experiments and arguments for germ layers in the embryo, and documented the successive stages of organ formation, which supported the theory of epigenesis

Jean-Baptise Lamarck (1744-1829)

French naturalist, biologist, soldier. According to him, evolution occurs because organisms can inherit traits which have been acquired by their ancestors. For example, giraffes find themselves in a changing environment in which they can only survive by eating leaves high up on trees. So, they
stretch their necks to reach the leaves and this stretching and the desire to stretch gets passed on to later generations. As a result, a species of animal which originally had short necks evolved into a species with long necks.

Charles Robert Darwin (1809-1882)

English naturalist, geologist, biologist. According to natural selection, organisms that are better physically equipped to survive, grow to maturity, and reproduce. For example, the long necks of giraffes allowed the species to feed from the leaves which grow high on trees—rather than graze, as
short-legged, short necked animals. Therefore, there was simply an environment which included trees with leaves up high and that was a favorable food source to long-legged, long-necked animals such as the giraffe.

Trofim Denisovich Lysenko (1898-1976)

Soviet agronomist and biologist. Rejected Mendelian genetics and natural selection. His plant breeding, hybridization, and selection theories linked Lamarckism to Marxism–Leninism. Lysenkoism theory: plants are self-sacrificing—they do not die due to a lack of sunlight or moisture but so that
healthy ones may live and when they die, they deposit themselves over the growing roots to help the new generation survive. His "law of the life of species”: plants of the same "class" never compete with one another.

Nikolai Konstantinovich Koltsov (1872-1940)

Russian biologist and pioneer of modern genetics (omnis molecula ex molecula). Proposed that the shape of cells was determined by a network of tubules forming a skeleton (cytoskeleton). According to Koltsov, traits are inherited via a "giant hereditary molecule" which would be made up of "two
mirror strands that would replicate in a semi-conservative fashion using each strand as a template.” Proposed that epigenetic changes affected the expression of chromosomes. He died unexpectedly following years of government persecution...

Conrad Hal Waddington (1905-1975)

British developmental biologist, provided the foundations for systems biology,
epigenetics, and evolutionary developmental biology. In 1942, coined the term epigenetics —meaning upon, above, in addition to, or near—, to explain “the causal interactions between genes and their products, which bring the phenotype into being”, i.e., epigenesis (17 th Century concept).

What does epigenetics mean?

literally, means above, or on top of, genetics

practically, describes phenomena in which genetically identical cells or organisms express their genomes differently, causing phenotypic differences

refers to the study of heritable changes in gene expression or cellular phenotype caused by mechanisms other than changes in the underlying DNA sequence

information coded beyond the DNA sequence, such as in covalent modifications to the DNA or modifications to the chromatin structure

epigenomics

refers to the sum of genome-wide epigenetic patterns, distinguishes and defines one tissue from another, stem cells from somatic cells, and aged from young cells

genetics, in comparison to epigenetics

static, genotype, germ cell inheritance

epigenetics, in comparison to genetics

dynamic, phenotype, somatic inheritance

pathway for environmental gene programming, epigenetics

epigenator (differentiation signals, temperature variations), epigenetic initiator (DNA binding factors, non-coding RNAs), epigenetic maintainer (histone/DNA modifiers, histone variants)

epigenetic mechanisms

key determinants for organization and expression of the genome

include RNA interference, DNA methylation, histone modification

biological examples of epigenetic phenotypes

barr body, polytene chromosomes, yeast mating types, blood smear, tumor tissue, mutant plant, cloned cat, twins

epigenetically variable

almost all the cells in our body are genetically identical, yet our body generates many different cell types (time and space), organized into different tissues/organs and express different proteins`

twins and epigenetic variation

though monozygous twins share a common genotype and are genetically identical and epigenetically similar when they are young, as they age they become more dissimilar in their epigenetic patterns

genetic variation emerges

across the lifespan

while the globular core is involved in histone-histone packing and DNA-contact, 

the N-terminal tails point outwards and are available for interaction with cell signaling pathways

histone modifications

covalent attachments (chemical groups) to the amino acids of histones, usually the side chain groups

affect the affinity (binding) of histones to both DNA and to other proteins which can interact with histones

can affect nucleosome and higher order DNA structures

are predominantly (but not exclusively) added to the extended histone tail structures

histone modifications are ________ modifications

post-translational

which amino acid is modified during acetylation and ubiquitination?

lysine

which amino acid is modified during methylation?

lysine and arginine

which amino acid is modified during phosphorylation?

serine and threonine

evidence for a histone code

specific modifications correlate with specific processes, specific modification patterns and sequences, interactions of specific modified residues with classes of protein domains

an epigenetic code includes

stable (not transient) through cell cycle, heritable (through cell generations), consistent (always associated with defined chromatin behaviour), predictive (i.e. upon discovery of a new modification)

immunofractionated chromatin

acetylated fraction enriched in active chromatin

chromatin immunoprecipitation assays (ChIP)

show enrichment of acetylated histones in active promoters

neutralizing mutations in histone tails affect

gene expression

acetylated histone tails enriched in

transcribed chromatin

acetylation and gene activation/transcription

makes nucleosomal DNA more accessible for TF binding- a role in potentiation towards gene being transcribed (read)

ChIP steps

cross-link protein to DNA in living cells with formaldehyde

break open cells and shear DNA

add pre-blocked protein G agarose beads

add primary antibody of interest

immunoprecipitate to enrich for fragments bound by protein of interest

reverse cross-links and treat with proteinase K

detect and quantify precipitated DNA through PCR and hybridization methods

ChIP on microarray chip

same process, but now with antibody specific to one type of histone modification

intergenic microarray used to detect the labeled DNA based on illuminessence

histone modifications correlate with

specific genomic processes

localized histone modifications

gene or region-specific

transcription- H3-K14ac; H3-K4me

silencing- H3-K9me; H4-K20me

DNA repair- H2Ag-S129ph

broad histone modifications

regional or genomic

chromatin condensation- H3-S10ph

apoptosis- H2B-S14ph

DNA repair- core histone SUMO

histone deacetylases (HDACs)

family of 11 enzymes that remove acetyl groups from lysine amino acid on histone proteins, allowing histones to wrap the DNA more tightly 

classified into 4 classes

recruited to their target promoters through a physical interaction with a sequence-specific transcription factor

part of multi-protein complexes, e.g. Sin3a, NuRD; associated with transcriptional repression

histone acetyltransferases (HATs)

enzymes that acetylate conversed lysine amino acids on histone proteins

acetylation of lysine neutralizes the positively-charged histone, reducing affinity between the negatively charged DNA, which renders DNA more accessible to transcription factors

divided into 5 families; e.g. CREB-binding protein (CBP)

recruited to their target promoters through a physical interaction with a sequence-specific transcription factor

part of multi-protein complexes, e.g. TFTC complexes; associated with transcriptional activity

many coactivators are these

acetylation coactivators

Tetrahymena HAT= homolog of yeast GCN5, a known trx-coactivator

coactivator p300/CBP

P/CAF = “p300/CBP associated factor”

hTAF250, dTAF230, yTAF130

several coactivators for nuclear receptors (SRC-1, ACTR)

present model for chromatin activation/silencing

HAT-activity becomes recruited to promoters through specific interactions with activators, leading to local acetylation of nucleosomes around promoters

HAT-activity in coactivator-complexes

recruited to promoters by activating TF-interaction

what are HDACs associated with?

repressing transcription factors

genome acetylation

global levels surprisingly high

on average, 13 of the 30 tail Lys residues in a histome octamer are acetylated

steady-state level of acetylation is maintained by opposing actions of HAT and HDAC complexes

gene specific acetylation

targeting of HAT and HDAC complexes to promoter regions that creates specific patterns of hyper- and hypoacetylation in a background of global acetylation that correlate with transcription activation and repression, respectively

factors that can affect histone modification

diet (increases: caffeine, calorie restriction, high-salt, alcohols; decreases: copper, chromium, nickel, niacin), drugs (increases: HDACs and SIRT inhibitors; decreases: HATs inhibitors, acetyl-CoA analogs), metabolism (increases: inflammation, immune activity, infections, hypoxia, smoking, exercise, glucose; decreases: estrogens, high glucose)