Histone Proteins Dr Nivedita Patnaik Chromatin is made

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 Histone Proteins Dr. Nivedita Patnaik

Histone Proteins Dr. Nivedita Patnaik

ØChromatin is made of repeating units of nucleosomes, which consist of 146 base pairs

ØChromatin is made of repeating units of nucleosomes, which consist of 146 base pairs of DNA wrapped around an octamer of four core histone proteins (H 3, H 4, H 2 A and H 2 B)

Introduction • Histones are a special group of proteins found in the nuclei of

Introduction • Histones are a special group of proteins found in the nuclei of eukaryotic cells responsible for DNA folding and chromatin formation.

 • Chemically they are • highly alkaline basic proteins • Histones are positively

• Chemically they are • highly alkaline basic proteins • Histones are positively charged • abundance of positive amino-acids, arginine and lysine

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Classes of Histones • There are two main classes of Histones: • Core Histones

Classes of Histones • There are two main classes of Histones: • Core Histones • Linker Histones • Core Histones: In core histones following families are included • H 2 A • H 2 B contain more lysine • H 3 • H 4 contain more arginine • Two of each of these core histone proteins assembles to form one octameric nucleosome core particle, and 147 base pairs of DNA wrap around this core particle.

Linker Histones • Linker histone included: • H 1 • H 5 highest lysine/arginine

Linker Histones • Linker histone included: • H 1 • H 5 highest lysine/arginine ratio • The linker histone protein H 1 binds the nucleosome at the starting and ending sites of the DNA, thus locking the DNA into place and help in the formation of higher order structure. • H 5 histones are individual proteins involve in the packaging of specific region of DNA.

 • Function of the histone protein in a chromosome • The DNA is

• Function of the histone protein in a chromosome • The DNA is housed in chromosomes in the form of nucleosomes • It is basic unit of chromosome or chromatin fiber. It is DNA duplex coiled around a core of eight histone proteins • Positively charged histones are linked with negative charged phosphate groups of DNA

 • Some histone proteins function as spools for the thread-like DNA to wrap

• Some histone proteins function as spools for the thread-like DNA to wrap around • looks like beads on a string

 • The nucleosomes + H 1 histones = 30 nm spiral Solenoid •

• The nucleosomes + H 1 histones = 30 nm spiral Solenoid • It maintains the chromosomal structure

histone modifications ØHistone proteins contain a globular Cterminal domain and an N-terminal tail ØThe

histone modifications ØHistone proteins contain a globular Cterminal domain and an N-terminal tail ØThe N-terminal tails of histones can undergo a variety of posttranslational covalent modifications including methylation, acetylation, ubiquitylation, sumoylation and phosphorylation on specific residues Ø regulate key cellular processes such as transcription, replication and repair

Post-translational histone modifications A = acetylation M = methylation P = phosphorylation U =

Post-translational histone modifications A = acetylation M = methylation P = phosphorylation U = ubiquitination ØUnlike DNA methylation, histone modifications can lead to either activation or repression depending upon which residues are modified and the type of modifications present 12

Histone Acetylation & Deacetylation • Histone acetylation • • – Histone acetyl transferases (HATs)

Histone Acetylation & Deacetylation • Histone acetylation • • – Histone acetyl transferases (HATs) Adds acetyl groups to histone tails Reduces positive charge and weakens interaction of histones with DNA Facilitates transcription by making DNA more accessible to RNA polymerase II • Histone deacetylation • • – Histone deacetylases (HDACs) Removes acetyl groups from histone tails Increases interaction of DNA and histones Represses transcription

Acetylation • It is the introduction of an Acetyl functional group to the Lysine

Acetylation • It is the introduction of an Acetyl functional group to the Lysine amino acid of the histone tail. • These reactions are catalyzed by enzymes with "histone acetyltransferase" (HAT) or "histone deacetylase" (HDAC) activity.

Effects of Acetylation • • -ve charge on histone. reduces affinity of tail for

Effects of Acetylation • • -ve charge on histone. reduces affinity of tail for adjacent nucleosomes creating a transcription permissive environment increase the access of transcription factors

Methylation • It is the introduction of an Methyl functional group to Lysine or

Methylation • It is the introduction of an Methyl functional group to Lysine or Arginine of the histone tail. • These reactions are catalysed by enzymes with "histone methyltransferase” • ‘Arg’ can be methylated once or twice, and ‘Lys’ once, twice or thrice.

Histone Methylation • Histone methyl transferases (HMTs) • Histone lysine methyl transferases(HKMTs) methylate lys

Histone Methylation • Histone methyl transferases (HMTs) • Histone lysine methyl transferases(HKMTs) methylate lys (K) residues • Protein argenin methyl transferase (PRMTs) Methylate arge(R) residues • Methylation can result in activation or repression of expression • trimethylation of histone H 3 at lysine 4 (H 3 K 4) is an active mark for transcription • dimethylation of histone H 3 at lysine 9 (H 3 K 9), a signal for transcriptional silencing

Effects of methylation • Methylation does not neutralize charge but recruit silencing or regulatory

Effects of methylation • Methylation does not neutralize charge but recruit silencing or regulatory proteins that bind methylated histones. • Chromodomain containing proteins interact with methylated histone tails. • transcription repression

Histone phosphorylation (H 3) • Histones are phosphorylated during mitosis. • Histones are also

Histone phosphorylation (H 3) • Histones are phosphorylated during mitosis. • Histones are also phosphorylated by signal transduction pathways like the ERK pathway in response to external signals. It is not known how (and if) this phosphorylation contributes to gene expression.

Histone ubiquitylation Addition and removal of Ub (a LARGE moiety) to histone tails –

Histone ubiquitylation Addition and removal of Ub (a LARGE moiety) to histone tails – • Functions largely unknown in vertebrates ubiquitylation • H 2 A K 119: repression • H 2 B K 120: activation • H 3 and H 4: DNA repair (CUL 4) de-ubiquitylation – Recrutiment of other proteins in yeast • H 2 A Dub (PCAF) • H 2 B Ubp 8 (SAGA) • Functions: transcription elongation, polycomb repression

Study of histone modifications • Histone modifications are studied using the chromatin immunoprecipitation (Ch.

Study of histone modifications • Histone modifications are studied using the chromatin immunoprecipitation (Ch. IP) assay. • Ch. IP on chip is the high throughput form of the Ch. IP assay wherein the immunoprecipitated DNA, instead of being subject to the usual PCR, is hybridized to a microarray chip with printed oligonucleotides corresponding to various regions of the genome. • This helps to study the localization of a specific histone modification to various parts of the genome.

Epigenetic therapy HDACi EZH 2 i SAHA (Vorinostat ), Depsipeptide Leads to hyperacetylation Activation

Epigenetic therapy HDACi EZH 2 i SAHA (Vorinostat ), Depsipeptide Leads to hyperacetylation Activation of silenced proapoptotic genes Pro-apoptotic genes are upregulated Lead to apoptosis in cancer cells. Cutaneous T cell lymphoma Deazaneplanocin Activate apoptotic cell death in breast cancer 26

 • Primary nonsmall cell lung cancer (NSCLC) is the main cause of malignancy-related

• Primary nonsmall cell lung cancer (NSCLC) is the main cause of malignancy-related mortality in Asian and Western populations. • positive correlation between lower levels of H 3 K 9 ac, H 3 K 9 me 3 and H 4 K 16 ac and tumor recurrence. However when patients were further clustered according to histone modification patterns (i. e. acetylation dominant, methylation dominant, co-dominant and modification negative), the acetylation-dominant group exhibited better survival prognosis, but methylation dominant and modification negative status was associated with poor prognosis

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