Nucleus

Question 1

Heterochromatin Vs. Euchromatin

Answer 1

Heterochromatin is transcriptionally inactive, is more densely packed, marked by specific histone modifications that restrict gene expression, and can be spread and limited by boundaries or insulators. Euchromatin is less condensed and generally associated with active gene expression.

Question 2

Histone Modifications and the Histone Code””

Answer 2

Histone code refers to post-translational modifications via specific proteins on histone tails that regulate gene expression and chromatin structure.

Methylation: linked to heterochromatin formation and gene slicing.
Acetylation: associated with active transcription.

Reader complexes interpret modifications, and writer and eraser complexes propagate or remove these marks.

Question 3

Methods to determine Chromatin position

Answer 3

Fluorescence in situ hybridization (FISH) uses fluorescent probes to detect specific DNA sequences and shows spatial organization in the nucleus.

Chromosome conformation capture (3C) maps physical interactions between chromatin regions, revealing their 3D organization.

Question 4

Gene Activity and Nuclear Position, example

Answer 4

Genes’ localization within the nucleus can change depending on their activity. Inactive genes may be positioned at the nuclear periphery, while active ones are often found in transcriptionally active domains.

For example, tagging a gene with a fluorescent marker and imaging its position during activation/inactivation could reveal positional changes relative to the nuclear envelope.

Question 5

Position Effect Variegation (PEV) vs. Insulators

Answer 5

PEV: gene silencing due to proximity to heterochromatin.

Insulators: act as barriers, preventing the spread of heterochromatin into active regions.

Question 6

Losing selectivity in the Nuclear Pore

Answer 6

FG- repeat Nups with inert proteins, disrupting the barrier function while retaining the pore structure.

Question 7

Different types of Nucleoporins

Answer 7

1. Structural nucleoporins: Provide a scaffold for the NPC (e.g., FG-Nups with phenylalanine-glycine repeats, critical for selective gating).
2. Channel nucleoporins: Line the central channel, mediating interactions with transport receptors.
3. Anchoring nucleoporins: Attach the NPC to the nuclear envelope and maintain its stability.

Question 8

Protein nuclear import

Answer 8

1. Key components:
   Cargo proteins: Contain a Nuclear Localization Signal (NLS).

Importin receptors: Recognize NLS and transport cargo.

Ran-GTP/Ran-GDP gradient: Maintains directionality.

2. Process:
3. Cargo protein with a nuclear localization signal (NLS) binds to an importin.
4. The complex moves to the nuclear pore and interacts with FG-repeat nucleoporins.
5. Importin-cargo complex translocates through the pore.
6. Inside the nucleus, RanGTP binds to importin, releasing the cargo.
7. Importin and RanGTP return to the cytoplasm, where RanGTP is hydrolyzed to RanGDP for recycling

Question 9

Protein Nuclear Export

Answer 9

1. Key components:
   Cargo proteins: Contain a Nuclear Export Signal (NES).

Exportin receptors: Recognize NES.

Ran-GTP: Powers export.

2. Process:
3. Cargo with a nuclear export signal (NES) binds to exportin and RanGTP in the nucleus.
4. The complex moves through the nuclear pore.
5. RanGTP hydrolyzes to RanGDP in the cytoplasm, releasing the cargo.
6. Exportin and RanGDP return to the nucleus.

Question 10

Regulation of Nuclear Import

Answer 10

Post-translational modifications:
Phosphorylation of cargo proteins may expose or mask their NLS.

Sequestration: NLS-bearing proteins can be retained in the cytoplasm by anchoring proteins.

Signal transduction: Activation of pathways (e.g., during stress) can recruit specific import machinery.

Ran cycle regulation: Alteration of the Ran gradient can affect transport directionality.

Question 11

Levels of chromatin compaction and supporting experiments

Answer 11

Nucleosome Formation: DNA wraps around histone octamers to form nucleosomes, the basic unit of chromatin. This provides a packaging ratio of about 6. X-ray crystallography has revealed the structure of nucleosome core particles​

30 nm Fiber: Nucleosomes are compacted further into a 30 nm fiber, often modeled as a zigzag or solenoid structure. Electron microscopy (EM) evidence supports this model by imaging reconstituted chromatin fibers​.

Chromosome Territories: Fluorescence in situ hybridization (FISH) experiments demonstrate that chromosomes occupy distinct territories within the nucleus during interphase.