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Cell and Developmental Biology > Protein Targeting > Flashcards

Protein Targeting Flashcards

1
Q

What are target sequences?

A
  • a.a sequences that act as address labels
  • directs proteins to correct destination in a cell
  • e.g. KDEL sequences
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2
Q

What is the KDEL sequence?

A

Lys-Asp-Glu-Leu

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3
Q

What does the nuclear pore consist of?

A 
Scaffold Nucleoporins
Channel nucleoporin
Disordered region
- line the pore, portions extend into the pore space
Nuclear Basket
- extends into the nuclear space
Cytosolic fibrils
- extend to the cytosol
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4
Q

What are nucleoporins?

A

proteins that make up the nuclear pore

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5
Q

What are importins?

A

Importins are a type of karyopherin that transports protein molecules into the nucleus by binding to specific recognition sequences (nuclear localisation sequences)

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6
Q

What are the two classes of importins?

A

Importin-α and importin-β

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7
Q

How can the importins be recognised / carry different cargo proteins?

A

Each have a slightly different import sequences so will be recognised by different cargo proteins

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8
Q

What are the two structural ways importin-β can function?

A
  1. can function as a monomer
  2. heterodimer with importin-α
    - importin-α acts as an adaptor protein
    - importin-α is bound to the cargo, and importin-β is bound to importin-α
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9
Q

What is an example of a importin-α cargo?

A

cellular cargo: BRCA1

viral cargo: HIV-1 integrase

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10
Q

What sort of repeats do nucleoporins contain?

A

FG (Phe-Gly) repeats

- repeats line the channel of the nuclear pore complex

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11
Q

How are importin-β’s thought to transport their cargo through the nucleus?

A

They transport through the nuclear pore via weak transient interactions with FG repeats

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12
Q

What is nuclear import and export controlled by?

A

Ran GTPases

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13
Q

What is the Ran cycle?

A

Cycle between the two states:

  • The switching on of Ran involves the displacement of GDP with GTP
  • Conformation change causes the released of GDP
  • Hydrolysis of GTP make it into GDP
  • Turns it from its ON state into OFF state
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14
Q

What protein and exchange factor is needed during the Ran cycle? Inc locations.

A

Guanine Nucleotide exchange factor (GEF) needed to replace GDP with GTP - turning it ON
- located in the nucleus, bound to chromatin
- Ran-GEF
GTPase Accessory protein (GAP) helps the process of turning it OFF, converting GTP to GDP
- located in the cytosol
- Ran-GAP

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15
Q

What is the process during the importing of cargo into the nucleus?

A
  1. Protein with NLS / cargo is binding to an importin
  2. Trafficked into the nucleus
  3. Ran-GTP (active site) binds to the importin
    - Will trigger a conformation change
  4. Cargo released into the nucleus
  5. Importin bound to Ran-GTP is imported back out of the nucleus into cytosol
  6. Ran-GAP switches off Ran (converts Ran-GTP to Ran-GDP)
    - Hydrolysis of GTP
    - Ran can no longer take part in importing and releases the importin
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16
Q

What state is Ran when bound to GTP and where in the cell would it be located?

A

ON state

- nucleus

17
Q

What is the process during the exporting of cargo out of the nucleus?

A
  1. Cargo that needs to be exported has an exporting signal
  2. Recognised by the Exportin protein
  3. Bound to Ran-GTP
  4. Complex transported out of nucleus into the cytosol
  5. After exiting the complex, Ran-GTP converted into Ran-GDP
  6. Exportin released as conformational change has occurred
18
Q

What transcription factor controls expression of the genes involved in T cell activation?

A

NF-AT

19
Q

Where is NF-AT found in its inactive state?

A

NF-AT is found in the cytosol

20
Q

How is NF-AT expression controlled in T cells?

A
  1. Exposure of T cells to antigen results in a rise in intracellular Ca
  2. When there is high intracellular [Ca], calcineurin binds to NF-AT masking a nuclear export signal
    - calcineruin also dephosphorylates NF-AT, exposing a nuclear import signal
  3. NF-AT is then imported into the nucleus
  4. Activation of gene transcription
  5. Once T-cell is a resting T-cell this will lead to low Ca conc
    - calcineurin no longer binds to NF-AT exposing the nuclear export signal
  6. NF-AT is phosphorylated once more and exported from the nucleus, switching off gene transcription
21
Q

What are cyclosporin A and FK506?

A

Cyclosporin A & FK06:

immunosuppressant drugs taken for rheumatoid arthritis, psoriasis, and in organ transplants to prevent rejection.

22
Q

How does cyclosporin A and FK506 work?

A

Inhibition of the action of calcineurin. It forms a complex to block the phosphatase activity of calcineurin, resulting in a failure to activate the genes regulated by the NF-AT transcription factor. These genes include those required for B-cell help such as interleukin.

23
Q

What happens during high cholesterol levels to stop transcription of more cholesterol?

A

SREBP = transcription factor
- Needs to be moved into the nucleus
High levels of cholesterol:
- SREBP is found in the cytosol associated with SCAP
- SCAP is binds to cholesterol
- This retains SREBP in the ER membrane
- Remains inactive (cannot function as transcription factor)

24
Q

What happens during low cholesterol levels to encourage transcription of more cholesterol?

A

Cholesterol wont bind to SCAP

  • When cholesterol not bound, SCAP has a conformational change
  • Changes into a vesicle and enters the Golgi membrane
  • Encounters 2 proteases
  • They both cleave SREBP
  • releases cytoplasmic domain of SREBP
  • also releases a release factor which goes into the nucleus
  • Can undergo transcription
25
Q

How is the sex determined in mammals?

A

SRY is a transpiration factor that inhibits female anatomical structures in males
promotes formation of tested from undifferentiated gonadal tissue

26
Q

What is the Swyer Syndrome?

A
  • rare disorder where there is a failure of gonads to develop
  • females with syndrome are XY, are externally female but lack ovaries or testes
27
Q

What is the cause of Swyer Syndrome?

A
  • mutations in the SRY gene on the Y chromosome
  • can affect ability of the transcription factor to develop
  • mutated SRY cannot interact with importin so cannot be trafficked into the nucleus and interact with the target gene
28
Q

What other diseases are linked to nuclear transport?

A
  • Tau Interferes with Nuclear Transport in Alzheimer’s Disease
  • impaired nucleocytoplasmic trafficking has emerged as a mechanism contributing to ALS

Cell and Developmental Biology (16 decks)

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