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DNA in Nucleus

Why DNA is not “naked” in the cell

DNA is packaged and associated with proteins for:

  • regulation and modification
  • protection
  • saving space

DNA binds to DNA-binding proteins; together, DNA + proteins are called chromatin.

DNA-binding proteins: histones and chromatin organization

Histone proteins and the nucleosome

Histone core (octamer):

  • \(2\times H2A\), \(2\times H2B\), \(2\times H3\), \(2\times H4\) form the core that DNA coils around

Properties :

  • histones are rich in basic amino acids (positively charged)
  • DNA is negatively charged (phosphate backbone)

Key numbers :

  • each core has ~150 bp wrapped around it (about 2 full circles)
  • this structure is a nucleosome

Histone H1 (linker histone)

  • histone H1 binds outside the core and stabilizes the complex
  • links one nucleosome core complex to another
  • overall appearance: “beads on a string”

Higher-order chromatin packing

10 nm chromatin fiber (“beads on a string”)

  • 10× packaging

30 nm chromatin fiber (solenoid)

  • the 10 nm fiber forms a solenoid state around a center axis
  • ~6 histones per turn
  • core is histone H1
  • 30 nm chromatin fibers
  • 40× packaging

Radial looping

  • 30 nm chromatin fibers loop around a “nuclear flag pole” (a DNA-binding protein)
  • 20–50× packaging

Non-histone proteins

  • non-histone proteins also participate in chromatin structure (example: ROC)

Euchromatin vs heterochromatin

  • heterochromatin (darker): transcriptionally inactive
  • euchromatin (lighter): transcriptionally active

Histone tail modifications (chromatin regulation)

Acetylation

  • enzyme: histone acetyltransferase (HAT)
  • effect: opens chromatin

Mechanism :

  • uses acetyl-CoA → releases CoA
  • acetylizes histone tail, replacing \(+\) charge with an acetyl group
  • reduces ionic attraction between histones and DNA

Deacetylation

  • enzyme: histone deacetyltransferase (HDAC)
  • effect: closes chromatin

Mechanism :

  • uses water → releases acetate
  • restores \(+\) charge on histone tail

Chromosome structure landmarks

Centromeres

  • constricted regions of chromosomes linking sister chromatids
  • linkage via cohesin complex
  • kinetochore: where spindle fiber attaches

Chromosome arms

  • p arm: small arm
  • q arm: long arm

Centromere position categories

  • metacentric: \(p = q\)
  • submetacentric: \(p < q\)
  • acrocentric: \(p \approx 0\)

Chromosome banding notation

  • defined by staining pattern of chromosomes
  • format: location #/X/Y, p/q/none, position two digits.subband

Telomeres

Telomeric repeat

  • telomeres contain repeats: \(5'\text{-TTAGGG-}3'\)

FISH staining

  • FISH: fluorescence in situ hybridization

T-loop / D-loop structure

  • not all double-stranded at the very end
  • a single-stranded region loops back into double-stranded DNA (t-loop)
  • creates a displacement (d-) loop
  • provides binding sites for proteins / shelterin complex

Functions of telomeres

  • protection from nucleases
  • protection from chromosome fusion
  • protection from DNA repair

Telomere shortening

  • 50–200 nt lost
  • due to replication problems and environmental problems when DNA opens up

Hayflick limit and cellular senescence

  • Hayflick limit: ~40–60 cell cycle limits
  • then cells enter senescence (permanent growth arrest)

Telomerase

Components :

  • telomerase reverse transcriptase (hTERT)
  • telomerase RNA (hTR)

Function:

  • adds telomeric repeats to chromosome ends