UNSW Cell Biology

This lecture introduces how information is transferred from stable stored information (DNA) converted to an intermediate (mRNA, rRNA, tRNA) of variable stability, exported from the nucleus to the cytoplasm where mRNA is then translated into Protein. This is gene expression, the products of this process are used either within the cell, exported (exocytosis) or used to replace worn out components.

We will study this topic at the level of the cellular components and organelles involved in the process: ribosomes, endoplasmic reticulum, Golgi apparatus, vesicles (transport and secretory).

Lecture Text: Lecture Word Document 662 Kb

Page Links: Introduction | Objectives | Lecture Audio | Textbooks | Difference between Prokaryotes and Eukaryotes | Membrane Evolution | Nuclear Compartment | Nuclear Envelope | Nuclear Cytoskeleton | Nuclear Lamina | Nuclear Pores | Nuclear Bodies | Chromosomes | Chromosome Territories | Nucleolus | Chromosome Features | Telomere | Centromere | Replication Origins | Histones | Nucleosomes | Hutchinson-Gilford Progeria Syndrome | Online Textbooks | Web Links | 2007 Lecture Slides | Comments | Acronyms

Postulated that an early "coating" structure lead to the infolding of the primitive plasma membrane to form the many membrane covered organelles in the cytoplasm.

These modules may also be the evolutionary precursor to the nuclear pore structures and account for the double membrane that coats the nucleus.

Model of Membrane Evolution (Image: PLOS - A Structural Analysis of Eukaryotic Membrane Evolution)

• Nuclear envelope
• Nuclear cytoskeleton
• Nucleolus
• Chromosome territories
• Interchromatin compartment
• “speckles” interchromatin granule clusters
  • Splicing speckles or SC 35 domains
  • thought to be sites of storage of mRNA splicing factors
• nuclear bodies - Cajal and PML

Cartoon of General Nuclear Structure

(Image: Nat Rev Genet. 2007 Feb;8(2):104-15.)

• Forms structural compartment
• Nuclear envelope two concentric membranes
• Breaks down each mitosis (recycled)
• Outer membrane continuous with Endoplasmic Reticulum (Endoplasmic Reticulum is covered in Lecture 5)
• Contains holes “nuclear pores”

Nuclear Envelope showing nuclear pores and underlying nuclear matrix

(EM Image Martin Goldberg)

• intermediate filaments
• large family of different filament types (covered in Cytoskeleton Lecture – intermediate filaments)
• 10 nm in diameter, forms rope-like networks
• lamin polypeptide form dimers
  • central alpha-helical regions of two polypeptide chains are wound around each other
• assembly
  • head-to-tail association of dimers form linear polymers
  • side-by-side association of polymers form filaments

Nuclear Envelope showing underlying nuclear matrix (green)

(EM Image Martin Goldberg)

• Vertebrate lamins are classified into 2 types - A and B
• Mammalian somatic cells show 2 species of each type
  • A type - lamins A and C (Mr 74 kD and 65 kD)
  • B type - B1 and B2 (586 aa protein, Mr 66,334 Da)
• lamins phosphorylation state affects nuclear envelope assembly state
  • phosphorylation nuclear envelope disassembly
  • dephosphorylation nuclear envelope assembly
• Mutations in lamins can lead to human disease
  • More? Hutchinson-Gilford Progeria Syndrome

• Lamins also link DNA to nuclear envelope
  • Outer laminar surface is connected to inner nuclear membrane
  • inner laminar surface attached to thechromatin

1999 Nobel Prize Medicine - Günter Blobel for the discovery that "proteins have intrinsic signals that govern their transport and localization in the cell"

JCB Movie: Nuclear Pore Complex movement in interphase (9.6 MB)

• Protein complex
• External diameter of about 120 nm (30 times the size of a ribosome)
• Channel diameter 25 nm
• channels between nucleus and cytoplasm (import/export)
• passive passage of small polar molecules, ions,
• active (selective/ regulated) passage of macromolecules, proteins and RNAs

Nuclear Pore cartoon (lateral cut away view)

Cajal Bodies

• also called - nucleolar accessory bodies, coiled body, gems
• 0.1 - 2.0 microns, 1-10/ nucleus
• Gems and Cajal bodies two forms of same structure
• GEMS (Gemini of coiled bodies)

PML Bodies

• promyelocytic leukaemia nuclear bodies
• also called PODs, ND10 or Kremer bodies
• Function unknown
• regulation of diverse cellular functions?
• viral infection, cellular transformation, innate immunity, growth control, apoptosis
• dynamic hubs sensing stress and DNA damage

(Images: http://redbone.umassmed.edu/)

• not “visible” at interphase, condense for mitosis (1,000 fold)
  • condensation allows chromosomes to move along mitotic spindle without breaking or tangling
• eukaryotes have separate chromosomes
  • Human 24 pairs, 22 autosomes, 2 sex chromosomes
• diploid 2 copies of each chromosome (inherited one male/one female)
  • except male sex chromosomes X from mother Y from father
• DNA and protein
• packing of DNA
• DNA structure
• encodes genome (humans 30,000 genes, draft sequence published in 2001)
• DNA genes encode RNA and proteins
• DNA can also encodes nothing

Human Idiogram

Karyotype from a female human lymphocyte (46, XX)

Chromosomes were hybridized with a probe for Alu sequences (green) and counterstained with TOPRO-3 (red). Alu sequences were used as a marker for chromosomes and chromosome bands rich in genes.

PLoS - Three-Dimensional Maps of All Chromosomes in Human Male Fibroblast

• Space within the nucleus occupied by individual chromosomes
• Several different models as to how these territories interact

Click to see movie of chromosome territories.

(Images: Nat Rev Genet. 2007 Feb;8(2):104-15.)

• Appearance
  • Fibrillar center, dense fibrillar component, and granular component
  • Changes during cell cycle
  • each mitosis - nucleolus breaks up as chromosomes condense
  • after mitosis - nucleolus reforms from coalesce of tips of 10 chromosomes
• Nucleolus size depends on cell metabolic activity
• Sites of ribosomal (rRNA) gene transcription, processing, and ribosome assembly
• All cells contain multiple copies of rRNA genes

Cartoon of nucleus structure showing Nucleolus

(Image: Conference on The Dynamic Nucleus: Questions and Implications, London 2002)

• at ends of all chromosomes (not bacterial DNA circular)
• roles in chromosome replication and maintenance
• replication
  • for replicating the ends of linear chromosomes
• maintenance
  • proposed to provide each cell with a replication counting mechanism that helps prevent unlimited proliferation
• each cell division shortens telomere 50–100 nucleotides
• DNA 100s to 1,000s repeats of a simple-sequence containing clusters of G residues (humans AGGGTT)
• Telomerase enzyme maintains length

• directs movement of each chromosome into daughter cells every time a cell divides
• centromere embedded in heterochromatin
• satellite DNA sequences (AT-rich) repeated many thousands of times
• proteins assemble on this to form Kinetochore
  • attachment site for spindle microtubules

• DNA replication initiates at multiple origins (ori)
• in both prokaryotic and eukaryotic DNA
• multiple origins in eukaryotes (human genome about 30,000 origins)
• each origin produces two replication forks (moving in opposite directions)

A single DNA Replication Fork

• only in eukaryotes
• small postitively charged proteins (binds negatively charged DNA)
• not sequence specific binding (as in transcription factors)
• 4 core histones (H2A, H2B, H3, and H4)
• 2 linker histones (H1/H5)

Histone Protein Organization with DNA

• formed by DNA wrapped around histones
• unit particle of chromatin (nucleosomal histones) (discovered 1974)
• EM unfolded DNA has "beads on a string" appearance
• second order folding forms 300 nm fibre
• condensed DNA for mitosis 700 nm fibre

DNA increasing Orders of Packing

• In more than 80% of cases the gene defect responsible for HGPS is a single spontaneous mutation in codon 608 of the LMNA gene, which encodes both lamin A and lamin C
• progerin is a mutant form of the nuclear architectural protein lamin A

(Images: PLoS - The Cell Nucleus and Aging)

Seee also: Scaffidi P, Misteli T. Lamin A-dependent misregulation of adult stem cells associated with accelerated ageing. Nat Cell Biol. 2008 Mar 2; [Epub ahead of print]

HGPS patients cell nucleus has aberrant morphology (bottom, right) rather than the uniform shape typically found in healthy individuals (top, right).

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Notice also that in some slides I have added annotations in brackets with my initials (MH - )

Links: Current Course Outline 2008