Nuclear Import & Export

Question 1

What is the name of the macromolecular pore that the nuclear membrane contains? How many?

Answer 1

~500-1000 Nuclear Pore complexes (NPC)

Question 2

What does the Nuclear Pore complex contain?

Answer 2

1. ~ 30 distinct nucleoporins (Nups)
2. Membrane ring: (scaffold)
3. Core scaffold
4. cytoplasmic filaments
5. Nuclear basket

Question 3

3 distinct functions of NPC

Answer 3

Allow movement of moecules through nuclear membrane:

1. diffusion through size filtering
2. Spontaneous migration of amphiphilic molecules
3. Facilitated transport
   
   • cargo associates with an amphiphilic molecule to get it in ;)

Question 4

What are Karyopherins known as?

Answer 4

Amphiphilic Cargo transporters 
aka
Cargo receptors
aka
Importins/Exportins

Question 5

What repeats are found in Nups?

Answer 5

FG repeats

-serve as hydrophobic binding sites (filaments) for NPC

Question 6

Basic principles of facilitated transport across pore complex (4)

Answer 6

1. E dependent
2. T dependent
3. Signal dependant
4. Saturable

Question 7

What are the two families of karyopherins?

Answer 7

1. Receptor family

2. Adapters

Question 8

Overview of Receptor family Karyopherins

Answer 8

1. Contain Ran-GTP binding domain.
2. Binds with FG domains of nucleoporins (Nups)
3. Another domain binding specific cargo/multiple cargos

Question 9

Overview of Adapter Karypherins

Answer 9

1. Capable of selecting different cargos

2. Increases efficiency of transport

Question 10

NLS typically consists of which aa?

Answer 10

Basic motif: arginine + lysine rich

Question 11

NES typically consists of which aa?

Answer 11

hydrophobic motif: Leucine rich

Question 12

Why is Importin is typically considered as a heterodimer?

Answer 12

Importin usually an alpha beta pair.

Consists of an alpha (adaptor) and beta (transport receptor)

Question 13

tRNA and miRNA use what type of export mechanism?

Answer 13

Ran dependent transport

tRNA:Exp-t

Question 14

List different RNA that get exported and their respective export factors

Answer 14

tRNA:Exp-t
miRNA: Exp-5
snRNA: CRM1
mRNA: NXF1 + NXT1, Aly

rRNA: CRM1, NXF1+ NXT1

Question 15

Describe mRNA export

Answer 15

Note: not part of Karyopherin family

1. initiated by Aly linking premRNA to nuclear export
2. allows recruitment of transport factors NXF1 + NXT1
3. Remodeling by DB helicase removes ALY and forms export competent complex
4. Kapα adaptor binds mRNA
5. Kapβ transport receptor binds Kapα and mRNA
6. Protein complex and nuclear export protein and nuclear basket interact and export occurs
7. Once on the other side, a bunch of proteins associate with it and cause the complex to dissociate

Question 16

mechanisms on how cargo nucleocytoplasmic transport is regulated in:

cargo

Answer 16

1. Post translational modification
2. Post transcriptional modification
3. intermolecular/intramolecular masking
4. affinity enhancement
5. sequestration

Question 17

rRNA is ran dependent/independent transport

Answer 17

both
uses CRM1, NXF1+ NXT1

similar to snRNA and mRNA

Question 18

mechanisms on how cargo nucleocytoplasmic transport is regulated thru:

cargo

Answer 18

1. Post translational modification
2. Post transcriptional modification
3. intermolecular/intramolecular interaction

Question 19

mechanisms on how cargo nucleocytoplasmic transport is regulated thru:

transport receptors

Answer 19

1. expression
2. sequestration
   
   • via entropy barriers created by Nups
   • Ran GTP/GDP can help overcome barrier

Question 20

mechanisms on how cargo nucleocytoplasmic transport is regulated thru:

NPC

Answer 20

1. pore permeability

2. protein expression and stability

Question 21

Describe when it is necessary for BRCA2 to get imported into the nucleus and how it is able to stay localized in nucleus

Answer 21

1. BRCA-RAD51 complex is transported into nucleus for DNA repair
2. Complex is retained in nucleus bc NES on RAD51 is occluded by binding to BRCA2.
3. BRCA2 NES is occluded by BSS1 binding during stressful conditions

Question 22

Give an example of how error in nuclear transport can contribute to breast cancer

Answer 22

A mutation from Asp to His in BRCA2 prevents binding with BSS1

- which means BRCA2's NES is exposed, and the mutant is rapidly exported    - RAD51 is also exported

Since BRCA2 is unable to stay in nucleus and function in DNA repair —> genomic instability –> breast cancer

Question 23

Give an example of how error in nuclear transport can prevent TSGs from doing its job?

Answer 23

1. Normally TSG are localized in cytoplasm
2. Under stressful conditions, TSG can go into nucleus and do its job
3. Many tumors have upregulation of nuclear export proteins
   ie: CRM1 or MDM2 (exporting p53)
4. Tumor suppressors are still functional, so selective inhibitors that can prevent export can restore tumor suppressor function