L12 Nuclear Receptors Flashcards
Nuclear receptors location and types
Nuclear receptors are receptors located inside the cell. These receptors are found either in the cytoplasm (Type I) or the nucleus (Type II) of a cell.
Main effect of stimulation of a nuclear receptor
Altered gene transcription
Nuclear Receptor subfamiles
7 subfamiles
48 NRs
Nuclear receptor binds to ligands (in or out of cell) and interact with what via recognition of response elements
Intracellularly
DNA
2 xenobiotic receptors
- pregnane X receptor (PXR)
- constitutive androstane receptor (CAR)
What do xenobiotics to the CYP expression
increase expression
Orphan Receptors.
When some NRs do not have a known ligand at this point in time.
RXR meaning (nuclear receptors)
Retinoid X Receptor
Type 2: RXR heterodimer
Ligand, dimerisation,location and response element
L: fatty acids, retinoic acids, cholesterol
D: heterodimer(RXR + Class II NR).
Lo: Nucleus (main) but also cytoplasm.
RE: Direct repeats
Type 1: Steroid hormone NR
Ligand, dimerisation,location and response element
L: Glucocorticoids and estrogen.
D: homodimer.
Lo: cytoplasm (main) but also nucleus.
RE: Inverted repeats
RE for inverted repeats how many base pairs
one RE half-site spacer (bp=3 is common)
RE for direct repeats how many base pairs
one RE half-site spacer (bp=0-5)
Class I in the absence of ligands are located primarily
Cytoplasm
Structure of a nuclear receptor - 4 domains
N-terminal (A/B)
DNA-Binding Domain (C)
Hinge Region (D)
Ligand-Binding Domain (E/F)
N-terminal domain (A/B)
- least conserved - varies in length and amino acid sequence
- activation function 1 (AF1)
- binds co-regulators in a ligand-independent way to modify binding or regulatory capacity of receptor
DBD (DNA binding domain, C)
- highly conserved - responsible for DNA recognition and binding
- two zinc fingers formed by cysteine-rich loops
1. first zinc finger - recognise specific hormone response elements (HREs)
2. second zinc finger - role in receptor dimerisation
Hinge region (D)
- links the DBD and the LBD (ligand binding domain); highly flexible
- may contain a nuclear localisation signal (NLS) - role in receptor nuclear translocation
- NLS can overlap with the DNA binding domain
[NLS will guide NR to translocate from the cytoplasm to the nucleus via the binding of a ligand. ]
Why fluorescence of proteins
To monitor movement of receptor of interest
LBD (ligand binding domain, E/F)
- fairly conserved
- a hydrophobic pocket formed by 12 α-helices
o helix 12: important role in co-activator/co-repressor switching - role in receptor dimerisation
- activation function 2 (AF2): binds co-regulators in a ligand-
dependent manner - post-translational modifications
How an antagonist and agonist inhibits and activates AF2
Antag will bind to the LBD to sterically inhibit binding of AF2 and so there is no attachment of co-activator proteins.
Agonist bind to LBD to allow AF@ to bind which allows binding of co-activators which lets gene transcription proceed.
(III) dimeric orphan NR (L,D,Lo,RE)
L: unknown
D: homo or hetero
Lo: nucleus
RE: direct repeat
(IV) monomeric orphan NR (L,D,Lo,RE)
L: unknown
D: monomer
Lo: nucleus
RE: binds to one extended RE
half-site
The HPA axis
- release of glucocorticoids (GCs; cortisol in human)
- negative feedback mechanisms
Glucocorticoids (class 1)
- maintain homeostasis and respond to stress
- regulate metabolic and immune responses
- anti-inflammatory and immunosuppressive
- synthetic GCs to treat inflammatory and autoimmune diseases
Glucocorticoids can treat inflammation and immune disease bc of
its ability to
* maintain homeostasis and respond to stress
* regulate metabolic and immune responses
Glucocorticoid receptor (encoded by the NR3C1 gene)
- alternative splicing of exon 9 → GR(a) and GR(ß)
- GR(y) contains an arginine insertion in the DBD
- other splice variants
NLS1 and NLS2 (Hinge region)
- NLS1 - overlaps with the end of DBD and the hinge region
- NLS2 - overlaps with the LBD
GR-mediated signaling + hsp90
- unliganded monomeric GR resides in the cytoplasm and forms a complex with HSP90 (heat shock protein 90) and other chaperone proteins
- ligand binding→ GR dissociates from the complex→ expose nuclear localisation signals
- GR is translocated into the nucleus
increase or drecrease
1.transactivation: gene transcription
2.transrepression: gene transcription
1.↑
2.↓
GR-mediated regulation of gene transcription
GR dimer will interact with its own response element to regulate gene transcription.
GR form homodimer will interact with own to increase expression of target gene
and react with negative RE like a monomer to inhibit the transcription of target gene
Direct (direct binding of GR to glucocorticoid response element [GRE])
- GR dimer - binds to GRE (GGAACAnnnTGTTCT) - ↑ gene transcription
- bind to negative GRE (IR nGRE, CTCC(n)0-2GGAGA) - ↓ gene transcription
activator protein 1 (AP1) and nuclear factor kB (NF-kB)
- transcription factors - ↑ gene transcription in response to stimuli, e.g., cytokines, growth factors
- pro-inflammatory - important role in inflammatory diseases
interaction between GR and AP-1/NF-kB → transrepression
- tethering - GR binds directly to DNA-bound AP-1 or NF-kB (TSF)
- composite - GRE-bound GR interacts with neighbouring DNA-bound AP-1 or NF-kB
Non-permissive RXR heterodimer
- e.g., thyroid hormone receptor or vitamin D receptor
- heterodimer is activated by
ligands of the partner NR
while RXR is silenced
activated by RXR ligand attached to the partner RN not the RXR
Permissive RXR heterodimer
- e.g., peroxisome proliferator-
activated receptor - heterodimer is activated by
ligands of either NR, and
synergistic effects when both
NRs are activated
activated by retinoic acid or partner’s ligand meaning that ligand can bind to either the partner or the RXR and will activate NR
PPAR(y) y=gamma
white and brown adipose tissue - adipogenesis, lipid metabolism and insulin sensitivity
What type of NRs:
Androgen, Oestrogen, glucocorticoid, progesterone
Type 1
Cyto, homo, translocate to nucleus, recruit co-activators, TSF and other proteins
What type of NRs:
ReinoidX, retinoic acid, thyroid hormone, peroxisome proliferator, pregnaneX
Type 2 (usually lipids)
Nuclear, Hetero with RXR, complexed with co-reperssors that are displaces for co-activators
PPAR(y) agonists (e.g., thiazolidinediones) are used clinically to treat what
type 2 diabetes
PPAR(y) and insulin sensitisation
- regulate adipocyte differentiation and mature adipocyte function
- lipid metabolism
- glucose homeostasis
o Glut4 ↑
o PI3K ↑
o CAP ↑
o IRS-1/IRS2 ↑
4 Post-translational modifications of NRs
acetylation (A)
phosphorylation (P) - change transcriptional activity
SUMOylation (S; small ubiquitin related modifiers)
ubiquitination (U) - facilitate degradation
PXR and CAR as xenobiotic sensors
nuclear receptors and form RXR heterodimers
* PXR - pregnane X receptor
* CAR - constitutive androstane receptor
↑ CYP enzymes / Phase 2 enzymes / drug transporters
phenobarbital-mediated CYP induction
- EGFR-mediated signaling phosphorylates CAR to inhibit its transcriptional activity
- binding of phenobarbital to EGFR attenuates the signaling
- CAR is monomerised and dephosphorylated
- CAR undergoes nuclear localisation and forms a CAR/RXR heterodimer - ↑ target gene transcription
