🧠 Nucleus

Nucleus Functions

1. Genetic Material Management

   • Storage, replication, and repair of DNA

2. Gene Expression

   • Transcription: production of mRNA, tRNA, rRNA

   • RNA splicing: processing of RNA

3. Ribosome Production

   • Synthesis of ribosomal subunits

1. Storage, Replication, and Repair of Genetic Material

• DNA Structure and Organization

  1. DNA duplex

     • 2 nm in diameter

  2. DNA is complexed with histones to form nucleosomes

     • Nucleosome is 8 core histone proteins + DNA wrapped around 1.65 times = Euchromatin

     • Histone H1 is the 9th protein that stabilizes the wrap

     • Histone + DNA wrap height = 10 nm

  3. Histones fold in on each other (like an elastic band) - Make chromatin fibers

     • 30 nm diameter

  4. Chromatin coils

     • 300 nm in diameter

  5. Coils coil

     • 700 nm in diameter

  6. Fibers are tightly coiled to produce condensed chromatids

     • Total width of 2 sister chromatids: 1400 nm

• Stretched end-to-end, DNA molecules in a single human cell would measure about 2 meters (roughly 6 feet)

• DNA Damage Sources

  • UV light exposure

  • Replication errors

  • Chemical exposure

  • Cellular metabolism

  • Ionizing radiation

• DNA Repair Mechanisms

  • DNA repair machinery corrects damage

2. Expression of Genetic Material

• Transcription

  • Transcription produces mRNAs, tRNAs, rRNAs, and other types of RNA (e.g., miRNA, eRNAs, piRNAs)

  • Promoter region: Controls transcription initiation

  • RNA received 5’ cap and poly-A tail

• RNA Splicing

  • RNA splicing occurs before nuclear export of mRNAs, removes introns and joins exons

• Note

  • This is an oversimplification of the complex process of gene expression

3. Ribosome Biosynthesis

• Noncoding RNAs (e.g., tRNA and rRNA) do not encode proteins

• These RNAs are transcribed from DNA located in the nucleolus

• Extra info:

  • rRNA is transcribed in the nucleolus from ribosomal DNA (rDNA)

  • rRNA combines with ribosomal proteins (imported from cytoplasm) to form ribosomal subunits

  • These subunits are then assembled into large and small ribosomal subunits in nucleolus

  • rRNA and ribosomal proteins form functional ribosomes, which are essential for protein synthesis

  • Ribosomal subunits are exported to cytoplasm

  • In cytoplasm, subunits assemble to form functional ribosomes, which begin process of translation (protein synthesis)

Nuclear Structure

• Nuclear envelope: Double-layer membrane surrounding the nucleus

• Nuclear membrane: Composed of an inner and outer lipid bilayer

• Nuclear pores: Channels that allow material to pass between the nucleus and cytoplasm

• Nuclear lamina: Meshwork of intermediate filaments providing structural support to the nucleus

• Nuclear content:

  • Chromatin: DNA wrapped around histone proteins, organized into chromosomes

  • Nucleoplasm: Fluid inside the nucleus, containing enzymes, nucleotides, and other molecules

  • Nucleolus: Substructure in the nucleus responsible for ribosomal RNA (rRNA) synthesis and ribosome assembly

Nuclear Envelope Structure

• 2 parallel phospholipid bilayers separated by 10-50 nm space

• Outer nuclear membrane (ONM):

  • Binds ribosomes

  • Continuous with rough ER

• Inner nuclear membrane (INM):

  • Contains integral proteins

  • Connects to nuclear lamina

Importance of Nuclear Envelope

• Separates nuclear content from cytoplasm

• Separates transcription and translation processes

• Selective barrier that allows limited movement of molecules between nucleus and cytoplasm

Nuclear Lamina

• Supports nuclear envelope

• Thin meshwork of filamentous proteins:

  • Lamins (intermediate filaments) found in animal cells only

  • Plants have nuclear lamina, but not made of lamin protein (equivalent proteins in plants is unknown)

• Bound to inner membrane of nuclear envelope (NE) by integral membrane proteins

• Provides structural support for nuclear envelope

• Attachment sites for chromatin (e.g., heterochromatin)

• Nuclear lamina forms meshwork next to nucleoplasmic leaflet of inner nuclear membrane

Nuclear Pores

• 1 μm in diameter

• Perforate nuclear envelope

• Small molecules and ions passively diffuse

• Large proteins and RNA require active transport

• Gateways between cytoplasm and nucleus

• 3000-4000 pores/nucleus

• Pores are located where inner and outer membranes fuse

• Complex structure involving arrangement of different types of proteins

Nuclear Pore Complex

• Composed of nucleoporins (NUPs) – a large family of proteins

• Octagonal symmetry / basket-like structure

• Projects into cytoplasm and nucleoplasm

• The nuclear pore complex is a supramolecular complex

  • Supramolecular = Very large and complex molecule

  • 15-30 times the size of a ribosome

• Passive diffusion of molecules that are 40 kDA or less

  • Fast

  • 100 molecules/minute/pore

• Regulated movement of larger molecules

  • Slow

  • 6 molecules/minute/pore

Nuclear Import

Requires a Nuclear Localization Signal (NLS)

• NLS consists of several positively charged amino acids within the protein sequence

Nuclear Import Mechanism

1. Protein with NLS (cargo) interacts with Importin protein in cytoplasm

2. Cargo/Importin complex interacts with FG-NUPs at the NPC and enters the nucleoplasm

3. Ran-GTP interacts with Importin; cargo dissociates and stays in nucleoplasm

4. Ran-GTP/Importin complex exits nucleus through NPC

5. GTP is hydrolyzed to GDP

   • Importin released into cytoplasm finds new cargo

Ran-GTP

• OFF with GDP

• ON with GTP

• Step 3 - Conformational change after being turned on allows for Ran-GTP to let go of cargo

  • Cargo stays in nucleus

Nucleocytoplasmic Trafficking

Nuclear import/export are critical for cellular function

• Nucleotides for transcription

• Structural proteins (e.g. lamins)

• DNA packaging proteins (e.g. histones)

• Proteins for DNA replication, repair, and transcription

• Proteins for RNA processing (splicing) and export

• Proteins for ribosome synthesis and export

Nucleolus

• Largest structure inside nucleus of eukaryotic cells

• Primary function: Biosynthesis of ribosomes

  • Ribosomes consist of:

    • Small ribosomal subunits (reads RNA)

    • Large subunits (joins amino acids to form polypeptides)

  • Each subunit consists of:

    • Ribosomal RNA (rRNA) molecules

    • Ribosomal proteins

• Prokaryotes have 70S ribosome (30S in small, 50S in large)

• Eukaryotes have 80S ribosome (40S in small, 60S in large)

Ribosome Biosynthesis

• Synthesis of ribosomal rRNAs:

  • 5.85 rRNA

  • 18S rRNA

  • 28S rRNA

  • 5S rRNA

• rRNA processing

• Assembly of subunits:

  • rRNA + ribosomal proteins

• 40S and 60S subunits exported to cytoplasm

  • Assemble into 80S ribosomes