Signal transduction

Intracellular (nuclear) receptors

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Dimension of time and solubility

protein 5’3’

Role of intracellular receptors in signal transduction

steroid-thyroid-retinoid- receptor superfamily

Nuclear hormone receptor superfamily

Steroid-thyroid-retinoid- receptor superfamily

• Development

• Differentiation

• Cell-cell interactions

• Nutrient sensing

Some ligands of the steroid-thyroid-retinoid receptor superfamily

Module 1: Figure aldosterone and cortisol biosynthesis Module 1: Figure aldosterone and cortisol biosynthesis

Cell Signalling Biology www.cellsignallingbiology.org 2007 Cell Signalling Biology www.cellsignallingbiology.org 2007

„prereceptorial” activation of hormones - biotransformation

~ 150 protein (caterpillar – human)

similar structure – regulation of transcription

ligands: hormonesvitaminsdrugs, fatty acids

lipid soluble

binding to DNAligandother transcription factors

regulation ligand dependentnon receptor factors

Steroid-thyroid-retinoid- receptor superfamily

Principal mechanism of action of steroid hormones

A/B C D E/F

Nuclear Hormone ReceptorSuperfamily

Steroid family Non-steroid family

ER ,

GR TR ,

RAR ,

PPAR ,

VDR RXR ,

PRAR

DBD

Type I family Type II family

Helix 12

AF1 LBD - AF2

CAR, SXR/PXR

MR

LXR ,, FXR

A/B C D E/F

Nuclear Hormone ReceptorSuperfamily

Steroid family Non-steroid family

ER ,

GR TR ,

RAR ,

PPAR ,

VDR RXR ,

PRAR

HRE

GRER

DBD

Type I family Type II family

Helix 12

AF1 LBD - AF2

CAR, SXR/PXR

MR

LXR ,, FXR

GR

homodimer

A/B C D E/F

Nuclear Hormone ReceptorSuperfamily

Steroid family Non-steroid family

ER ,

GR TR ,

RAR ,

PPAR ,

VDR RXR ,

PRAR

HRE

TRRXR

DBD

Type I family Type II family

Helix 12

AF1 LBD - AF2

CAR, SXR/PXR

MR

LXR ,, FXR

heterodimer

Orphan receptors

more than 40 orphan subfamilies

ligand „candidates” smalllipofilic

retinoids

terpenoidsfarnesollong chain fatty acidsPGJ2 analogues

sequence homology unknown LIGAND

unknown FUNCTION

GRGR

Steroid receptorsGR glucocorticoidMR mineralocorticoidPR progesteronAR androgenicER estrogen

RARRXR

RXR heterodimersTR thyroid hormoneRAR trans RAVDR 1,25 – (OH)2 – VD3PPAR eicosanoids

EcR ecdysonRXR 9 cisz RA, terpenoids

(peroxisome proliferator activated receptor)

Consensus sequences of DNA response elements for differentnuclear hormone receptors

Fig 11.42 Lodish et al. Molecular Cell Biology

The glucocorticoid receptor and oestrogenreceptor bind to their respective response elements as homodimers. The response element is an inverted repeat

The vitamin D receptor, the thyroid hormone receptor and the retinoic acid receptor bind to their respective response elements as heterodimers (with RXR). The response element is an direct repeat.The spacing betweenthese repeats determines the specificity ofthe interaction.

Similar structure, different length

Conserved domains of transcription factors in nuclear-hormonereceptor superfamily

Fig 11.41 Lodish et al. Molecular Cell Biology

Two non-repeating C4 Zn finger motif

A/B C E

AF-2 domainAF-1 domain

Structure of nuclear receptors 2.

A/B

connections with

variable

DBD conservativebinding to HRE

two Zn fingers DNA binding

dimerisation( helix dimerisation)

coactivatorstransactivatorsproteins of transcription

DBD structures

Zinc Finger

NK H

VRQ

HRSLAS

KE

VFS

E

C

LG

CK

YNH2

HOOC

R

Zn

H

H

C

C

Zn

H

H

C

C

ZnH

H

C

C

Zn

C2H2

C4

C5

C6

Finger type Transcription FactorsGal4 C6Steroid hormone C4 + C6

Structure of nuclear receptors 2.

A/B

connections with

variable

DBD conservative

binding to HRE

two Zn fingers DNS binding

dimerisation

( helix dimerisation)

coactivators

transactivators

proteins of transcription

LBDligand binding with high affinity (KM > 1 nM)selective, stereospecific, reversible

C terminal part protein binding hsp

transactivatordimerisationtranslocation

D possible change of conformation „folds” hinge function translocation

Hsp90 - GR Steroid type

Steroid or hsp90

Steroid or hsp90

Grouping according to the localisation of the receptor (1)

A. steroid typeGR, AR, PR, MR, ER

longer A/B domainassociated to hsp when no ligand is bound

no HRE binding without ligand no silencer effect but repressor effects

importance of LBD

must dimerise for HRE bindingactivation domainsagonist, antagonist binding sites (different)

Grouping according to the localisation of the receptor (2)

B. Thyroid type

TR, RAR, VDR, RXR, PPAR, orphan

short A/B domenno hsp associationbinds to HRE without ligand silencer effect

can bind as a monomer

thyroid type

Modulating factors

1. presence of ligands

2. activation without ligands

3. receptor phosphorylation

4. structure of binding site – chromatin structure

5. nuclear non-receptor transcription factors

Modulating factors

presence of ligandsligand metabolism(e.g. metamorphosis, prostate: dihydrotestosterone production

enhanced AR effect)

activation without ligands

effect of neurotransmitters on sexual behaviourdopamine PR, ER, VDR activating effect

effect of growth factorsEGF activates ER – it can be prevented by antiestrogens

„crosstalk” membrane receptornuclear receptor

Modulating factors

receptor phosphorylationmultiple sitespresumably (also) after DNA bindingnuclear DNA-dependent protein kinases

roles needed for transactivation?needed for receptor transport?activation without ligands?

size of the „interface”

structure of binding site – structure of chromatinthe order of heterodimer according to the binding site

RXR - TRTR - VDR

„half binding site”hormone-induced changes in DNA conformation

alteration of nucleosome structure

Chromatin-based mechanisms

Histone deacetylases (HDAC) corepressors since they don’t recognize DNA directly but are recruited by association with sequence-specific, DNA-binding proteins. HDAC cleaves the acetyl moiety from histone tails.

Thyroid hormone receptor (TR) provides an example of a DNA binding protein that switches activity by changing its associated cofactor.In the absence of thyroxin, TR associates with a target gene but inhibits transcription because TR recruits an HDAC complex.Thyroxin induces a conformational change upon binding TR and causes dissociation of the HDAC and association of a HAT complex. This contributes to transcriptional activation.

General Scheme for Activation of Gene Transcription by Nuclear Hormone Receptors

Robyr, Wolffe, Wahli Mol. Endocrinol 2000

Therapeutic implications

GR+1

Gene Transcription

RNA Polymerase II

CYTOPLASM

NUCLEUS

GLUCOCORTICOID HORMONE (DEX) RECEPTOR (GR) ACTIVATION OF A GR RESPONSIVE GENE

Hormone Regulated GeneGRE

DEX

DEX

GR HSP90

p23HSP70

GR

DEX DEX

GR GR

DEX DEX

ER +1

Gene Transcription

RNA Polymerase II

CYTOPLASM

NUCLEUS

ESTROGEN (E) RECEPTOR (ER) ACTIVATION OF AN ER RESPONSIVE GENE

Hormone Regulated GeneERE

ER HSP90

p23HSP70

ERE E

E

E

E ER EER

T3

T3

T3

T3

T3R +1

Gene Transcription

RNA Polymerase II

CYTOPLASM

NUCLEUS

THYROID HORMONE RECEPTOR ACTIVATION OF A T3 RESPONSIVE GENE

T3 Regulated GeneTRE

RXR

VDR+1

Gene Transcription

RNA Polymerase II

CYTOPLASM

NUCLEUS

VITAMIN D (V) RECEPTOR (VDR) ACTIVATION OF A VDR RESPONSIVE GENE

Hormone Regulated GeneVDRE

V

VDR

RXR

RXR VDR

V

V

V

Receptor deficiency

Nuclear receptor mutations (LBD, DBD)

familiar diseases

X-linked AR mutationtesticular feminisation

no androgen response

vitamin-D-resistent rachitis

TR mutations in LBDrare syndromes ?

glucocorticoid resistancehypercortisolism without Cushing symptomes

Therapeutic significance of receptor detection

mamma carcinoma ER antiestrogen therapy

leukemiaslymphoid tumorsuterus tumors

GR

antisteroids: no dissociationcompetition

inhibition of dimerisation

tamoxifenantiestrogen

VDR ligand research: osteoporosis, prostate carcinoma, ER ligand research: osteoporosis, mamma carcinoma, menopausa