Download Steroid Hormones and Steroid Receptors – Pt. 1

Biology 575 Winter 2010
Human Biochemical Genetics
Steroid Hormones and
Steroid Receptors – Pt. 1
Lenore K. Beitel, Ph.D.
[email protected]
2010/3/16
A. Steroid Hormones
B. Steroid Hormone Receptors
•
S
Structure/Function
/F
i
C. Classical Mutations/SNPs in Steroid Receptors
•
Androgen Receptor (AR)
•
Glucocorticoid Receptor (GR)
•
Mineralocorticoid Receptor (MR)
•
Estrogen Receptor (ER)
D. Trinucleotide Repeat Variation in the AR
Intracellular Signalling
Multicellular organisms need intercellular communication to:
- organize body plan during embryogenesis
- maintain physiology and cell functions during life span
Steroid hormones:
- derived from cholesterol
- small hydrophobic signaling molecules
- enter target cells & bind to specific steroid receptors
Steroid receptors:
- ligand-inducible transcription factors
- regulate specific genes
- integrate diverse signaling pathways as are
targets of post-translation modification
- regulate other major signaling cascades
(signal transduction crosstalk)
hydrophobic hormones (steroids):
- carried in blood by transport proteins
- enter target cell and bind to receptors in cell
hydrophilic hormones (peptides):
- bind to receptors at cell surface
- activate formation of 2nd messengers
Steroid Hormone Structure
Overview of
Steroid Hormone Synthesis
Names of Steroids: Built-in Clues to their Structures
Suffix –ol = important –OH group ( i.e. cholesterol, cortisol)
Suffix –diol = two –OH groups ( i.e. estradiol)
Suffix –one = ketone group =O (i.e. progesterone, aldosterone, testosterone)
Distinguishing features
Coritsol and aldosterone: –OH at C-11
Cortisol: –OH at C-17
Aldosterone: –HC=O at C-18
Testosterone and androstenedione: =O at C-3
Testosterone: –OH at C-17
Androstenedione: =O at C-17
Testosterone: a C-19 steroid, =O at C-3, C-19 is CH3
Estrogen: a C-18 steroid, -OH at C3, aromatized A ring
Aromatase
A
t
(CYP19)
Group
Progestin
Steroid
Carbons
Principal
Source
Target
Tissue
Cholesterol
C27
Diet, liver
Progesterone
C21
Adrenal gland Precursor
Ovary,
placenta
Uterus,
oviduct
Adrenal gland Kidney
- cortex
Metabolic
precursor of all
steroids
Mineralocorticoid
Aldosterone
C21
Glucocorticoid
Cortisol
(hydrocortisone)
C21
Adrenal gland All cells
- cortex
Androgen
Testosterone,
C19
Testis
C18
Peripheral
tissues
Ovary
5α-dihydrotestosterone (DHT)
Estrogen
17β-estradiol
Hormonal
Function
Testis, brain,
prostate,
seminal
vesicles
Uterus,
vagina,
breast,
brain, liver
Maintenance of
pregnancy
Regulation of
electrolyte
balance
Regulation of
energy utilization,
catabolic action
Development of
male sex
characteristics,
male behavior
Development of
female sex
characteristics
Steroidogenesis
Wikipedia - Estradiol
Evolutionary tree for the
vertebrate steroid hormonethyroid hormone receptor
superfamily
-complex evolutionary history
-gene duplication events
-domain
domain swapping
α
Structural and Functional Organization
of Steroid Hormone Receptors
Steroid Hormone Receptor Domains
N-terminal domain
NURSA
Structure of Nuclear Receptor Ligand-binding Domain (LBD)
- conformational change upon ligand binding
- helix 12 traps ligand in pocket
Structure of liganded GR LBD-TIF2 complex
Hormone = dexamethasone (Dex), a synthetic glucocorticoid
Hormone (dexamethasone) in ligand-binding pocket of GR
- specific amino acids in ligand-binding pocket make contact with ligand
Structure of Steroid Receptor DBD
Estrogen Receptor Dimer Bound to DNA
Structure of Steroid Receptor DBD
Steroid Response Elements (SREs) of Various Steroid Hormone Receptors
SREs are enhancer elements; usually located in promoter region of genes
Steroid Receptors Bound to Inverted Repeats
NURSA
Co-transfection Assay - Transcriptional Activation
- steroid receptor expression can be driven by
different promoters (e.g. SV40, CMV)
- MMTV-GRE = MMTV-ARE
- reporter gene can vary (luciferase, growth
hormone, chloramphenicol acetyl transferase)
Luciferase Reporter Gene Assay
NURSA
Luciferase Reporter Gene Assay
NURSA
BIOCHEMICAL GENETICS
<GS
<ES
<EG
Nuclear Localization Signal (NLS)
Translocation:
- depends on NLS
- nuclear transport
through nuclear pores
is an ATP dependent
process
Steroid Receptor Action
Coactivators and corepressors
The conformation of a steroid receptor can change depending on whether it is bound to an
agonist (activating hormone) or antagonist (anti-hormone). Agonist-bound receptors can bind
coactivators while antagonist-bound receptors can interact with corepressors.
Alternate mechanism of inhibition of gene transcription:
occupation of SRE by steroid receptor dimers interferes with binding
of factor(s) required for gene expression
Steroid Receptors Coactivator Activity
(Histone Acetyl Transferases)
NURSA
Mechanism of Transcriptional Activation by Androgen Receptor
3
Chromatin Remodeling
4
5
TBP/TATAA
Targeting
PIC Stabilization
6
Transcription Initiation
TBP
RNA Polymerase II
HAT
Activity
TAFII
30
CBP/p300
1
Androgen
pCAF
BRG-1
TIF 2
TIF-2
Androgen
AR
AR
Androgen
ARE
AR
2
Heat Shock
Complex
Highlights of Steroid Receptor Structure and
Mechanisms of Action - 1
Steroid hormones:
- small molecules derived from cholesterol
- bind to a specific receptor and alter transcriptional activity of responsive
g
genes
Steroid hormone receptors:
- in cytoplasm or nucleus of target cells
- function as ligand-dependent transcription factors
- variable N-terminal domain involved in transcriptional activation
- central DNA-binding domain (DBD) binds to specific steroid response
(
)
elements (SRE)
- C-terminal ligand-binding domain (LBD)
- other subdomains for nuclear localization (NLS), dimerization, and
association with heat shock proteins, coactivators and coregulators.
Interactions between receptor domains modulate ligand dissociation,
dimerization and transactivation
Classical Mechanism of Action of Steroid Hormone Receptors
hsps
coativators
hsps
Highlights of Steroid Receptor Structure and
Mechanisms of Action - 2
Steroid hormones enter the cell by diffusion and bind to specific
receptors
Receptors undergo a conformational change which promotes heat
shock protein dissociation, exposes the DBD & NLS & dimerization
domains and allows receptors to interact with other proteins
Receptors translocate to the nucleus and bind to SREs as a dimer
Receptors form complexes with coregulatory proteins
The nearby promoter is activated and mRNA transcription is initiated
The mRNAs are translated to produce proteins that alter cell function
Coactivator/corepressor levels may influence response to hormone
Non-classical Actions of Steroid Hormones and Receptors
Take home message:
Steroids can have regulatory effects independent of transcription!!
¾ Nongenomic steroid actions are frequently mediated through specific
membrane receptors
¾ Estrogen receptors activated by and can activate mitogen-activated
protein kinases - ER can activate cell proliferation through ERK pathway
¾ Post-translation modifications can alter steroid receptor activity i.e.
phosphorylation
¾ Steroid receptors can bind to and modulate activity of other proteins