ANHB3323 (4) - Nucleus

What happens inside the Nucleus (4 things)

1. transcription (DNA to mRNA)

2. processing/export of mRNA

3. DNA replication

4. DNA integrity/repair

Birth of Daughter Nuclei

Each cell division:

• nuclear envelope breaks down and is reformed

Nuclear factors have to be re-imported via nuclear pores

Nuclear organisation has to be re-created

• Each cell is established by the formation of a new nucleus

Nuclear envelope

Double Membrane:

1. Outer = continuous with Rough ER

2. Inner = associates with nuclear lamina

has pores for import/export, molecules sit in the space between the two sheets of membrane

Nuclear import/export

All molecules pass through nuclear pores

Passive diffusion:

- for molecules under 20-30kDa

Large proteins:

- have export/import signals

- need cargo molecules which act as carriers

Nuclear lamina

Composed of lamina proteins:

• gives structural stability to nucleus

Mutations in Lamin Genes

Cause/lead to:

• progeria (premature ageing)

• other muscle diseases

Lamin Mutations

1. Nucleus cannot withstand mechanical stress

2. Alters nuclear organisation = gene expression changes

What is found inside the Nucleus?

Chromatin = DNA + packaging proteins

What is Chromatin connected to?

the Nuclear Lamina

Chromosome Territories

defined location of each chromosome:

- some at the periphery

- some in the centre

What influences Chromosome Territories?

Cell type + shape

Size of chromosome (larger chromosome are usually at the periphery)

Fluorescent in Situ Hybridisation (FISH)

Used to localise DNA sequences

How:

• short fragments ('probe') of DNA complement sequence of interest

• probe is labelled with fluorescent dye

• target DNA is deanatured, allowing probe to anneal

• It is possible to FISH on multiple chromosomes are the same time

Nucleolus (general info)

Forms around ribosomal DNA repeat

• Densest part of cell

Nucleolus function

Site of:

• ribosome production

• subnuclear sequestration of regulatory molecules

Paraspeckles (general info)

Stress-induced subnuclear bodies

Built around long noncoding RNA (NEAT1)

Paraspeckles function

Regulate gene expression by:

- Sequestration of paraspeckle proteins

Paraspeckle substructure

Has distinct zones:

- Core

- Shell

- Patch

Molecular movement

Despite crowding, molecules move rapidly throughout the nucleus

• inactive genes in the middle

• Actively transcribed genes are found at the edges or outside the territory

• genes cluster together to be transcribed

How to determine how fast molecules move in living cells

Photodynamics, via fluorescent protein fusion:

- bleach protein

- image recovery of fluorescence (how long it takes to recover)