epigenetics and developmental genetics

week 9 stibs

epigenetics

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

• covalent modifications to DNA and histones form a secondary code which dictates gene expression

epigenetic trait

stable, mitotically and meiotically heritable phenotype that results from changes in gene expression without alterations in DNA sequence

epigenome = epigenetic state of a cell

epigenators

signals that are received by cell that stimulate response via an intracellular pathway

epigenetic initiators

• action of initiators defines location at which epigenetic changes in chromatin will take place

• they can switch specific genes on/off and recruit enzymes that add/remove epigenetic tags

epigenetic maintainers

• epigenetic modifications are maintained by epigenetic maintainers

• conserve and sustain epigenetic changes in present and future generations

• operate anywhere in the genome

• depend on initiators to specify loci at which chromatin modifications will take place

• ensure that epigenetic modifications are transmitted to daughter cells by mitosis

DNA methylation

• addition of methyl groups to selected cytosine bases

• blocks transcription factor access

  • localised silencing of gene expression

• promotes chromatin condensation

  • regional silencing of gene expression

• DNA methylation patterns are maintained during DNA replication

• example of epigenetic change

  • stable alterations to gene expression

  • transmitted from one gen to the next

  • important during differentiation

imprinting related congenital disorders

• rare congenital diseases that affect growth, development and metabolism caused by changes in gene regulation, gene dosage (or in genomic sequences-rare)

examples of imprinting-related congenital diseases

• Beckwith-Wiedemann Syndrome

  • overly large infants, macroglossia (tongue larger than other structures), internal abnormalities, characteristic facial features

  • IgF2 gene activation on maternal genome

Rett Syndrome

• multiple neurological defects, loss of motor skills and speech

• mutations in MeCP2 methyl-binding protein

• activation of genes that are normally repressed, loss of heterochromatin

X inactivation of DNA methylation

• one X chromosome becomes increasingly methylated to control X chromosome gene dose

• random in placental mammals

• forms heterochromatin

• formation of Barr Body

histone acetylation

• addition of acetyl groups to histones via lysine residues

• linked to active genes

• evidence that chromatin becomes looser, promoting transcription

• (histones are positively charged so bind to negative DNA), acetylation of lysine and arginine neutralises this positive charge

microRNAs

• act as regulators of gene expression

• they can also function as tumour suppressors and oncogenes (play a role in cancer)

developmental genetics

• study of how genes control growth and development of an organism throughout its life cycle

hierarchy of cell fate

• specification/commitment

  • cell type not yet determined, can be switched to another fate

• determination

  • cell fate can no longer be transformed

• differentiation

  • changes in biochem, structure, function

development and gene expression

• variable gene activity hypothesis

  • activation/inactivation of different genes at diff times in diff cells

• spatial and temporal interactions to modify fates of cells

methodologies in developmental genetics

• model organisms

• nuclear transfer

• cell ablation

• fate mapping

• mutant screens

• fluorescence microscopy

differential gene expression

• detecting mRNA

  • in situ hybridisation

  • microarray analysis

  • RNA sequencing

• detecting protein expression

  • immunostaning

  • proteomics

• transgenic

  • genetically modified organisms

major mechanisms of cell fate determination

1. asymmetric localisation of determinants

2. induction between non-equivalent cells

3. lateral inhibition between equivalent cells

mechanisms to communicate positional info

• signalling morphogen gradients

• cell-cell contact

Caenorhabditis elegans

• nematode worm: well understood genetics, completed genome sequence

• developmental lineage from fertilised egg to adult has been mapped

• 959 somatic cells

1. asymmetric localisation of determinants

mRNAs and proteins move to poles along microtubules

2. induction: non-equivalent cells

• P2 signals EMS so E different from MS

• PS signals ABp so Abp is different from ABa (Notch-Delta)

Wnt in humans

• 19 different Wnt genes identified

  • specification of dorsal/ventral axis

  • formation of brain, muscle, gonads

  • homozygosity of WNT3 causes tetra-amelia (absence of all 4 limbs)

  • abnormal Wnt signalling associated with formation of tumours

• Wnt gradients work with TGF-beta signals to set up organiser

Notch-Delta signalling pathway

• P2 to ABp: short range system

• direct cell-cell contact

• developmental fate of adjacent cells

3. lateral inhibition: signalling between equivalent cells

• main role of Notch-Delta: specifying fate of equivalent cells in population

• interaction involves neighbouring cells

Notch-Delta in humans

• 4 different Notch receptors

• important in:

  • neuronal function and development

  • angiogenesis

  • haematopoiesis

  • T cell lineage committment

  • formation of heart

• dysregulated in many cancers, implicated in many diseases

inductive signalling and lateral inhibition work together

• vulval development in C. elegans : 3 rounds of cell-cell interactions that transmit/receive signals from other cells

human homologues of pain rule genes

• cleidocranial dysplasia (CCD) mutation in human CBFA/RUNX2 gene (Runt homologue)

• key transcription factor associated with osteoblast differentiation

• results in skeletal defects: open fontanel, no clavicles

segment polarity genes

• expression of segment polarity genes controlled by pair-rule gene products

• active in single band of cells extending around embryo surface (divides embryo into 14 parts)

• includes cell signalling gene products as well as TFs

holoprosencephaly

• Shh pathway involved in:

  • patterning of CNS

  • induction of floor plate

• defects in Shh: incomplete cleavage of prosencephalon (deformed head)

homeotic genes

• they specify parts of adult body

• activated by gap and pair rule gene products: determines which adult structures will be formed by each body segment

• homeotic mutants: structure formed by one segment transformed into that formed by another

Hox genes (drosophilia)

• 2 clusters of homeotic genes on chromosome 3

• homeobox encodes homeodomain TFs

Hox genes functions

• spatial and temporal colinearity

• highly conserved

• in genomes of all animals including vertebrates

• fundamental role in shaping body and appendages (limb malformations caused by HOXD genes)

mutation in HOXD13

synpolydactyly (SPD) : characterised by extra fingers and toes, bone abnormalities in hands and feet

morphogenesis: how tissues acquire positional information and develop in specific places

pattern formation laid down during embryogenesis:

• definition of cells of region

• establishment of signalling centers (provide positional information)

• differentiation of cells in a region in response to additional cues

• must be arranged in temporal-spatial pattern

TGF-β superfamily

• large group of structurally related
  genes –homo- or heterodimers
  • TGF-β
  • Bone morphogenetic protein (BMP)
  • Activin
  • Vg1
  • Nodal-related proteins
  • Trigger gene expression eg. activin
  activates expression of Xbra,
  goosecoid

BMP (bone morphogenetic protein)

• growth factor

• mutation in cartilage derived morphogenetic protein 1: skeletal abnormalities (brachydactyly)

• expressed throughout Xenopus blastula

• antagonists: chordin, noggin, follistatin

clock and wavefront model of somitogenesis (somites developing in embryo)

• oscillations of network of genes/gene products (FGF, Wnt, Notch)

• cells oscillate regularly between permissive and non-permissive states

• clock of signalling generated by Notch feedback loop that comes into contact with cells in permissive state (directed by wavefront: FGF, Wnt)

• undergoes epithelial mesenchymal transition

• pinch off from msot posterior pre somite

• Shh and BMP signalling also involved

fibroblast growth factors (FGF)

• FGF binds to FGF receptors (heparan sulphate proteoglycan interacts with 2 or more FGF molecules)

• induces dimerisation

• mitogens: pluripotent GFs

  • involved in limb development, neural induction, angiogenesis, keratinocyte organisation

FGF mutations

• FGF (23 genes) mutation: Vit D resistant rickets

• FGFR (4 genes) : differs in ligand affinities and tissue distribution

  • mutation lead to 12 different conditions

  • craniosynostosis syndromes

  • short-limb skeletal dysplasias

gastrulation

• organiser in humans (primitive streak)

• signals surrounding tissues

• induction: neural plate formation

• lateral inhibition

• establishes ecto, meso and endoderm