DNA Packaging

The Nucleus and DNA Packaging

Prokaryotic Cells

• Prokaryotic Cells – no nucleus

• Prokaryotes divide by fission.

• Most prokaryotes have one circular chromosome.

• As DNA replicates, each of the two resulting DNA molecules attaches to the plasma membrane.

• As the cell grows, new plasma membrane is added between the attachment points, and the DNA molecules are moved apart.

• Cytokinesis separates the one cell into two, each with a complete chromosome.

Eukaryotic Cells

• Eukaryotic cells divide by mitosis or meiosis.

• Eukaryotes usually have many chromosomes.

• Eukaryotes have a nucleus, which must replicate and, with few exceptions, divide during cell division.

Structure of Nucleus

• Usually the largest organelle in the cell.

• Contains most of the cell’s genetic material (DNA).

• Replication of DNA and the first steps in decoding it for protein production take place in the nucleus.

Nuclear Pores

• Transport of molecules across the nuclear envelope.

• From nucleus:

  • RNA

  • ribosomal proteins

• To nucleus

  • proteins (DNA polymerase and lamins)

  • carbohydrates

  • signalling molecules and lipids

• 8 Large protein granules surround each pore

DNA Packaging Problem

• Each nucleus ~ 5 - 10 μm in diameter

• Nucleolus can take up to 25% of its volume

• 2 m DNA in each nucleus

• Roughly 1/266,667 times ‘shorter’

• The question is: How is this possible?

Chromosomes

• In the nucleus, the DNA molecule is packaged into thread-like structures called chromosomes.

• Chromosomes are not visible if the cell is not dividing.

• Each chromosome has a constriction point called the centromere, which divides the chromosome into two sections, or “arms”.

• The short arm is labelled the “p arm”. The long arm is labelled the “q arm”.

• The location of the centromere on each chromosome gives the chromosome its characteristic shape and can be used to help describe the location of specific genes.

DNA Storage

• Humans have 46 chromosomes (22 pairs plus two non-paired sex chromosomes).

• One chromosome can have as few as 50 million base pairs or as many as 250 million base pairs.

• There’s a copy of our entire DNA sequence in almost every cell in our body.

• Over 99% of our DNA sequence is the same as other humans’.

DNA Compaction

• Proteins called histones compact chromosomal DNA into the microscopic space of the eukaryotic nucleus.

• The resulting DNA-protein complex is called chromatin.

• Proteins are added to DNA to make it more compact.

• This coiling requires work, and energy is needed to perform work.

• Within the nucleus, histones provide the energy (mainly in the form of electrostatic interactions) to fold DNA.

Histones

• Histones are a family of small, positively charged proteins termed H1, H2A, H2B, H3, and H4.

• DNA is negatively charged, due to the phosphate groups in its phosphate-sugar backbone.

Nucleosomes

• The basic repeating structural and functional unit of chromatin is the nucleosome, which contains nine histone proteins and about 166 base pairs of DNA.

Further DNA Packaging

• Packaging DNA into nucleosomes shortens the fiber length about sevenfold.

• A piece of DNA 1 meter long will become a "string-of-beads" chromatin fiber just 14 cm long.

• Chromatin is further coiled into an even shorter, thicker fiber termed the "30-nm fiber“, because it is approximately 30 nm in diameter.

• The 30-nanometer fiber may be highly irregular and not quite the uniform structure depicted in textbooks.

Chromatin Structure

• Chromatin is the name given to the mixture of DNA, Histones, and other proteins that make up Chromosomes.

• It is divided between heterochromatin (condensed) and euchromatin (extended) forms.

• The functions of chromatin are:

  • to package DNA into a smaller volume to fit in the cell,

  • to strengthen the DNA to allow mitosis and meiosis, and

  • to control gene expression and DNA replication.

Transcription & Replication

• Both require the two strands of DNA to separate temporarily to allow polymerases access to the DNA template.

• Nucleosomes and folding of chromatin into 30-nm fibers pose barriers to enzymes.

• Cells must have means of opening up chromatin fibers and/or removing histones transiently.

• There are two major mechanisms by which chromatin is made more accessible:

  • Histones can be enzymatically modified.

  • Histones can be displaced by chromatin remodeling complexes.

• These processes are reversible.

Cell Division

• Nuclear envelope breaks down, and microtubules attach to chromosomes.

• Thick, coiled chromosomes are lined up in the center of the cell on the metaphase plate.

• Spindle fibers are attached to the chromosomes.

Levels of DNA Packaging

1. At the simplest level, chromatin is a double-stranded helical structure of DNA. (2 nm)

2. DNA is complexed with histones to form nucleosomes. (11 nm)

3. Each nucleosome consists of eight histone proteins around which the DNA wraps 1.65 times.

4. A chromatosome consists of a nucleosome plus the H1 histone.

5. The nucleosomes fold up to produce a 30-nm fiber…

6. …that forms loops averaging 300 nm in length.

7. The 300-nm fibers are compressed and folded to produce a 250-nm-wide fiber. (700 nm)

8. Tight coiling of the 250-nm fiber produces the chromatid of a chromosome. (1400 nm)