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MCB2050 > Unit 2: Nucleus > Flashcards

Unit 2: Nucleus Flashcards

1
Q

Eukaryote nucleus

A
  • membrane bund
  • nuclear envelope
  • nuclear pores
  • organized internal nuclear structure (highly regulated)
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2
Q

Characteristics of nucleus

A
  • irregular shape
  • one per cell
  • largest organelle
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3
Q

Two main functioins:

A
  • compartmentalization of cellular genome and activities (DNA replication, transcription, RNA processing)
  • coordination of cellular activities (metabolism, protein synthesis, reproduction)
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4
Q

Nuclear structure components

A
  • nuclear envelope (membrane, lamina, pores)

- nuclear content (chromatin, nucleoplasm, nuclear matrix, nucleolus)

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5
Q

3 functions of nuclear envelope

A
  • separates nuclear content from cytoplasm (genome and cytosol; transcription/translation)
  • selective barrier (regulates certain molecules, composition of nucleus and regulates gene expression)
  • binds nuclear lamina - structural framework
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6
Q

Nuclear Lamina structure

A
  • thin meshwrok of long filament-like proteins

- bound to inner surface of nuclear envelope

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7
Q

Nuclear functions

A
  • support structure for nuclear envelope

- scaffold for chromatin and nuclear matrix (?) attachment

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8
Q

Progeria

A
  • premature aging
  • point mutation
  • deletion of 150 nucleotides
  • progerin = mutation of lamin A
  • structure of lamina compromised
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9
Q

Nucleoplasm

A

highly organized fluid-filled interior of nucleus

>30 specialized regions (subdomains) w/ special functions

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10
Q

Choromosomes

A
  • during interphase organized into discrete subdomains within nucleus
  • location related to activity
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11
Q

Interchromosomal channels

A

regions between domains that serve as barriers for unwanted interactions (DNA-DNA and DNA-protein)

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12
Q

Transcriptioin factories

A

active genes (chromatin) from different subdomains extend into interchromosomal channels to form “transcription factories” - where TF are concentrated

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13
Q

Nuclear Speckles

A
  • subdomains where mRNA splicing factors are concentrated - i.e. mRNA processing
  • located in interchromosomal channels next to TF
  • numerus and highly dynamic depending on needs of cell
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14
Q

Nucleolus

A
  • most obvious subdomain: irregular, dense; NOT membrane bound
  • 1-5 nucleoli depending on metabolic activity
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15
Q

Function of the nucleolus

A
  • ribosomal biogenesis
  • site of ribosomal rDNA transcription and rRNA processing
  • INITIAL stages of ribosomal subunit assembly (final assembly in cytosol)
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16
Q

Nuclear matrix

A
  • insoluble fibrillar-like protein network (mesh) distributed throughout the nucleoplasm
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17
Q

Role of the nuclear matrix

A
  • structural role (maintain overall shape)

- scaffold (organizing nuclear subdomains and anchoring protein factors)

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18
Q

Nuclear Pores

A
  • inner and outer membranes of nuclear envelope fuse = pore

- gateway between cytoplasm and nucleoplasm

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19
Q

NPC Structure

A
  • highly conserved in euk.
  • octagonal symmetry of overal structure
  • central scaffold (anchors complex to nuclear envelope)
  • forms an aqueous central channel
  • filament like proteins - FG nucleoporins (Nups) - inner surface of channel lined
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20
Q

FG domains

A
  • FG repeats (FG nucleoporins)
  • line inner sruface of channel
  • extended and flexible organization (highly disordered sec. structure)
  • extend into central channel
  • hydrophobic mesh - sieve like gel limits macromolec
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21
Q

Cytoplasmic and Nuclear rings

A
  • cytoplasmic and nuclear side of NPC

- linked o central scaffold and cytoplasmic filament s or nuclear basket

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22
Q

Cytoplasmic filaments

A
  • long protein filaments that extend into cytosol on cytosolic side
  • nucl. receptor cargo protein recognition and import
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23
Q

Nuclear basket

A
  • basket like structure located on nucl side of NPC

- involved in nucl receptor-cargo protein import and export

24
Q

Functions of NPC

A

size exclusion

  • passive diffusion of small molecules (nucleotides etc)
  • regulate mvt (active) of large molecules (RNAs and proteins)
25
Q

What is nucleoplasmin?

A
  • nuclear protein
26
Q

Where is nucleoplasmin synthesized

A
  • cytoplasm, associates with cytoplasmic fillaments and translocates into nucleus
27
Q

Nucleocytoplasmic transport (cytosol to nuclear) requires

A
  • energy
  • specific protein receptors
  • unique targeting signals
28
Q

Nuclear Localization Signal is used in

A
  • imported proteins

- mediates targeting of protein from cytosol to nucleus

29
Q

What is a NLS?

A
  • nuclear localization signal
  • a.a. sequence necessary and sufficient for cytosol to nuclear targetting
  • recognized by nuclear receptor proteins
30
Q

Necessary? NLS

A
  • sequence cannot be mutated or else the modified protein will fail to target nucleus
31
Q

Sufficient? NLS

A
  • if sequence linked to non-nuclear passenger protein is capable of redirecting fusion protein to nucleus
32
Q

Transport receptors are also known as

A

Karyopherins

  • importin - into nucleus
  • exportin - out of nucleus
33
Q

What is the role of karyopherins?

A
  • mobile proteins responsible for moving (ferrying) protein cargo across nuclear envelope
34
Q

G proteins

A
  • molecular switch in transport process

- i.e. Ran

35
Q

How are G proteins activated?

A
  • weak intrinsic GTPase activity
  • conformational change
  • GTP binding and hydrolysis regulates activity and conformation
36
Q

Ran-GTP and Ran-GDP(active/inactive)

A

Ran GTP = active

RanGDP = inactive

37
Q

Where is there a high concentration gradient of Ran GTP?

A
  • high in nucleus
  • low in cytoplasm
  • gradient maintained high Ran-GTP in nucleus maintained by accessory proteins
38
Q

How is Ran-GDP yielded?

A
  • Ran-GDP is yielded in the cytoplasm (deactivated)
  • GTPase Activating Protein (GAP)
  • HYDROLYSIS of GTP of Ran-GTP to GDP = Ran-GDP
  • Ran-GDP then moves (translocates) to the nucleus
39
Q

How is GTP produced?

A
  • produced in the nucleus
  • Guanine nucleotide Exchange factor (GEF)
  • GDP is EXCHANGED by GEF
  • to product Ran-GTP
  • NOT a PHOSPHORYLATION event (the whole GDP is replaced)
40
Q

What are the accessory proteins involved in maintaining the [Ran-GTP] gradient?

A
  • RCC1 (promotes conversion of Ran-GDP to Ran-GTP (high [Ran-GTP] in nucleus]
  • Ran-GAP-1 cytosolic protein promotes hydrolysis of Ran-GTP to Ran-GDP (maintains low [Ran-GTP] in cytosol)
41
Q

Ran-GTP gradient determines what?

A

directionality of nucleocytoplasmic transport

42
Q

What is required for nucleoplasmic transport (energy)?

A

GTP hydrolysis

43
Q

Step one of Nuclear import

A
  • nascent NLS-containing cargo protein
  • recognized in cytosol by importin
  • importin a recognizes and binds basic residues of cargo NLS
44
Q

Importin is a ____ proteins

A

heterodimeric

- containing a and b subunits (two distinct)

45
Q

Step Two Nuclear Import

A
  • cargo protein- importer complex moves through cytosol towards nucleus
  • cytoskeleton elements serves as highway for intracellular transport
  • surface of nucleus (outside) importin b subunit binds cytoplasmic filament at NPC
  • importin ability to bind to cytoskeleton
46
Q

Step Three Nuclear Import

A
  • cargo protein- importin complex translocaated through aqueous central channel of NPC
  • cargo-receptor complex interacts with FG-domains
  • interactions dissolve/untangle FG domain network and allow translocation through channel
47
Q

Step Four Nuclear Import

A
  • cargo-receptor complex associates with nuclear basket on inner NPC
  • cargo-receptor complex binds toRan-GTP via importin b
  • released from NPC and disassembled in nucleoplasm
  • success import NLS - containing cargo into the nucleus
48
Q

Step Five Nuclear Import

A
  • Ran-GTP bound importin b subunit moves back to cytosol due to [Ran-GTP] gradient
  • in cytosol, Ran-GTP hydrolyzed
49
Q

What hydrolyzes Ran-GTP in cytosol?

A

RanGap1

50
Q

What happens after RanGTP is hydrolyzed in cytosol?

A
  • Ran-GDP released from importin b
  • importin b used for another round of protein import
  • Ran-GDP moves back to nucleus due to gradient
51
Q

What converts Ran-GDP back to RanGTP?

A
  • accessory protein RCC1 in the nucleus
52
Q

What happens to importin a (and cargo proteins) in the nucleus (after import)?

A
  • need to move back into the cytosol - nuclear export

- importin a binds exportin (karyopherin)

53
Q

How are other cargo proteins brought back to the cytoplasm?

A
  • Nuclear export signal (NES)
  • specific sequence of a.a.
  • LxxLxxL
  • leucine motif
54
Q

How does the imporitina- exportin complex move into the nucleus?

A
  • binds Ran-GTP
  • high RanGTP conc in nucleus
  • RanGTP promotes stable assembly and transports to cytosol
55
Q

What happens to Ran-GTP in the cytosol after nuclear export?

A
  • GTP on Ran-GTP is hydrolyzed by Ran-Gap1

- Ran-GDP hydrolyzed releases exportin and releases importin a

56
Q

What happens to importin a and exportin once they are brought back into the cytosol?

A
  • importin a is used for another round of nuclear import Ran-GDP
  • exportin moves back into the nucleus for another round of export
57
Q

How do some proteins move into the nucleus without a NLS?

A
  • piggyback nuclear protein import
  • nascent protein lacking NLS binds NLS containing protein in cytosol
  • targeting and import of protein-protein complex mediated by importin receptors

MCB2050 (10 decks)

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