Modelové interakce jaderných receptorů a
enzymových systémů
JADERNÉ RECEPTORY
http://www.ens-lyon.fr/LBMC/laudet/nurebase/nurebase.html
General design of transcription factors in nuclearreceptor
superfamily. The centrally located DNAbinding
domain exhibits considerable sequence
homology among different receptors and has the
C4 zinc-finger motif. The C-terminal hormonebinding
domain exhibits somewhat less
homology. The N-terminal regions in various
receptors vary in length, have unique sequences,
and may contain one or more activation domains.
This general pattern has been found in the
estrogen receptor (553 amino acids [aa]),
progesterone receptor (946 aa), glucocorticoid
receptor (777 aa), thyroid hormone receptor (408
aa), and retinoic acid receptor (432 aa). [See R. M.
Evans, 1988, Science 240:889.]
Vazba na DNA:Vazba na DNA:
We can divide the receptors into
subgroups on the basis of their pattern
of dimerization. One group consists of
the steroid receptors, all of which
appear to function as homodimers.
This group includes receptors for
estradiol (ER), progesterone (PR),
androgens (ARs), glucocorticoids
(GRs) and mineralocorticoids (MRs). A
second major group contains receptors
that form heterodimers with retinoid X
receptor (RXR) ­ the receptor for 9-cis
retinoic acid. Members of this group
include the receptors for all-trans
retinoic acid (RAR),vitamin D3 (VDR)
and thyroid hormone (TR), as well as
liver X receptor (LXR), peroxisome
proliferator activated receptor (PPAR)
and others. A third group consists of
receptors that can bind DNA as
monomers, such as NGFI-B, RevErb,
ROR and SF-1.
A: Unliganded heterodimerizing
receptors, exemplified here by VDR,
exist as weakly associated heterodime
with RXR, presumably bound
nonspecifically to DNA [Haussler et al.,
1998]. Binding of the 1,25(OH)2D3
ligand to VDR (1) promotes high-affinity
heterodimerization with RXR
accompanied by binding of the
heterodimer to its direct repeat VDRE
(2).
B: Unliganded GR, like other receptors
in group (d) (see Fig. 2), exists as a
complex with heat shock proteins in the
cytoplasm. Upon binding its cortisol
ligand (1), GR dissociates from the
cytoplasmic complex, translocates to th
nucleus and forms a homodimer on its
palindromic GRE (2). Triggered by a
ligand-mediated change in GR
conformation, the AF1 and AF2 domain
then synergize to promote a series of
events (3­6) involving the recruitment o
coregulatory complexes similar to those
described for the VDR-RXR
heterodimer, but with some distinctive
features.
Evoluce jaderných receptorEvoluce jaderných receptorůů
Many family members have been identified by DNA sequencing only, and their
ligand is not yet known; these proteins are therefore referred to as orphan
nuclear receptors. The importance of such nuclear receptors in some animals
is indicated by the fact that 1­2% of the genes in the nematode C. elegans
code for them, although there are fewer than 50 in humans.
Syntéza ligandů
Jaderné receptory
Transaktivace
Metabolismus ligandů
Cílové geny
(metabolismus,
kontrola buněčné
proliferace,
diferenciace a
apoptózy)
Toxické látky, farmaka
??????????
Nízkomolekulární lipofilní sloučeniny jako ligandy = aktivita jaderných receptorů je
do značné míry závislá na syntéze a degradaci ligandů a naopak
Úloha jaderných receptorů a enzymů katalyzujících
degradaci nebo syntézu jejich ligandů:
* endokrinní regulace - steroidní hormony, thyroidní
hormony;
* regulace signálních drah ­ eikosanoidy,
metabolismus kyseliny retinové, vitamínu D3;
* metabolismus lipidů;
* metabolismus xenobiotik;
Modelový příklad 1:
Metamorfóza hmyzu
Regulation of insec
metamorphosis. (A
Structures of
juvenile hormone,
ecdysone, and the
active molting
hormone 20-
hydroxyecdysone.
(B) General pathwa
of insect
metamorphosis.
Ecdysone and
juvenile hormone
together cause
molts to keep the
status quo and form
another larval insta
When there is a
lower concentration
of juvenile hormon
the ecdysoneinduced
molt
produces a pupa.
When ecdysone
acts in the absence
of juvenile hormon
the imaginal discs
differentiate, and th
molt gives rise to
the adult
Formation of the ecdysone receptors. Alternative mRNA splicing of the ecdysone
receptor (EcR) transcript creates three types of EcR mRNAs. These generate
proteins having the same DNA-binding site (blue) and hydroxyecdysone-binding site
(red), but with very different amino termini.
Three isoforms of EcR have been identified in insects, each with a different,
stage-specific role in regulation of molting and development. This allows for one
steroid hormone to induce a variety of different tissue responses. In general,
EcR A is predominant when cells are undergoing a maturation response (from
juvenile to adult) and is predominant in imaginal discs, whereas EcR B1
predominates in juvenile cells during proliferation or regression. Little is known
about the function of the EcR B2 isoform.
DNA and hormone binding are similar in the three isoforms of EcR. Little is
known about the crustacean EcR isoforms and how they change during the molt
cycle. However, the EcR that has been cloned from the crab, Uca pugilator
(U31817, GenBank), shares 85­87% homology with that of Drosophila (M74078,
GenBank). The differences are primarily in the region of the molecule involved
with dimerization. Similar sequence similarities are found between the
heterodimeric partner, USP.
There are several ecdysteroids which bind EcR, including 20-hydroxyecdysone,
turkesterone, makisterone A, ponasterone A, and muristerone A. Some
arthropods may use specific ecdysteroids as their principal molting hormone, but
often several ecdysteroids are found within one group. The primary molting
hormone for a range of organisms, including some insects and crustacea, is 20OH
ecdysone (20 HE). Among other examples, makisterone A is an important
hormone for some crustacea and hemipteran insects.
Cholesterol (from diet- a vitamin for insects)
Prothoracic
gland
20-HydroxyecdysoneEcdysone
mono-oxygenase
(fat body, epidermis)
Conjugates
(storage)
Metabolites
(excretion)
Synthesis of molting hormones
SyntSyntééza ekdysteroidza ekdysteroidůů aa úúloha cytochromloha cytochromůů P450:P450:
Modelový příklad 2:
Metabolismus xenobiotik
http://drnelson.utmem.edu/CytochromeP450.htm
Aktivace
promutagenů:
Metabolismus léčiv:
JadernJadernéé receptory a dalreceptory a dalšíší proteiny:proteiny:
ABC TRANSPORTÉRY: MULTIDRUG
RESISTANCE (MDR) SYSTEM
Transport
ipidů,
xenobiotik
aj. látek
přes buněčné
membrány
3. FÁZE BIOTRANSFORMACE
(ABC TRANSPORTÉRY)
Modelový příklad 3:
Steroidní hormony
PPěět hlavnt hlavníích skupin steroidnch skupin steroidníích hormonch hormonůů::
ˇ Cortisol stimulates the release of amino acids from muscle. These are taken
up by the liver and converted to glucose.
Ťhe increased circulating concentration of glucose stimulates insulin release.
Cortisol inhibits the insulin-stimulated uptake of glucose in muscle via the
GLUT4 transporter.
Čortisol has mild lipolytic effects. These are overpowered by the lipogenic
action of insulin secreted in response to the diabetogenic action of cortisol.
Čortisol also has varied actions on a wide range of other tissues
The glucocorticoid receptor and activation by cortisolThe glucocorticoid receptor and activation by cortisol
1) Unbound, lipophilic cortisol readily crosses
cell membranes and in target tissues will
combine with the glucorticoid receptor (GR).
2) Like the androgen and progesterone
receptors, unliganded GRs are located in the
cytoplasm attached to heat shock proteins (hsp-
90, hsp-70 and hsp-56).
3) When hormones bind to these receptors hsps
are released and the hormone receptor
complexes translocate to the nucleus.
4) These complexes form homo- or heterodimers
and the zinc fingers of their DNA-binding
domains slot into the glucocorticoid response
elements (GREs) in the DNA helix.
5) Together with other transcription factors, such
as NF-B or c-jun and c-fos, they initiate RNA
synthesis (activation of RNA polymerase)
downstream of their binding.
Diagrammatic outline of the synthesis of cortisol from cholesterol in the adrenal corte
Cholesterol is either obtained from the diet or
synthesized from acetate by a CoA reductase
enzyme. Approximately 300 mg cholesterol is
absorbed from the diet each day and about 600
mg synthesized from acetate. Cholesterol is
insoluble in aqueous solutions and its transport
from the main site of synthesis, the liver, requires
apoproteins to form a lipoprotein complex. In the
adrenal cortex, about 80% of cholesterol required
for steroid synthesis is captured by receptors
which bind low-density lipoproteins (LDL)
although recent evidence has shown that highdensity
lipoprotein (HDL) cholesterol may also be
taken up by adrenal cells. The remaining 20% is
synthesized from acetate within the adrenal cells
by the normal biochemical route.
BiosyntBiosyntééza steroidnza steroidníích hormonch hormonůů::
aromatáza
Modelový příklad 4:
Metabolismus mastných kyselin
Receptory aktivovanReceptory aktivovanéé
peroxizperoxizóómovými prolifermovými proliferáátorytory
(PPAR)(PPAR)
The peroxisome proliferator-activated receptors (PPAR , , ) are activated by
polyunsaturated fatty acids, eicosanoids, and various synthetic ligands. Consistent with
their distinct expression patterns, gene-knockout experiments have revealed that each
PPAR subtype performs a specific function in fatty acid homeostasis.
PPAR is a global regulator of fatty acid catabolism. PPAR activation up-regulates the
transcription of liver fatty acid­binding protein, which buffers intracellular fatty acids and
delivers PPAR ligands to the nucleus. In addition, expression of two members of the
adrenoleukodystrophy subfamily of ABC transporters, ABCD2 and ABCD3, is similarly upregulated
to promote transport of fatty acids into peroxisomes where catabolic enzymes
promote -oxidation. The hepatocyte CYP4A enzymes complete the metabolic cascade
by catalyzing -oxidation, the final catabolic step in the clearance of PPAR ligands.
PPAR was identified initially as a key regulator of adipogenesis, but it also plays an
important role in cellular differentiation, insulin sensitization, atherosclerosis, and cancer.
Ligands for PPAR include fatty acids and other arachidonic acid metabolites, antidiabetic
drugs (e.g., thiazolidinediones), and triterpenoids. In contrast to PPAR, PPAR promotes
fat storage by increasing adipocyte differentiation and transcription o a number of
important lipogenic proteins.
Ligands for PPAR include long-chain fatty acids and carboprostacyclin.
Pharmacological activation of PPAR in macrophages and fibroblasts results in upregulation
of the ABCA1 transporter, and because of its widespread expression, PPAR
may affect lipid metabolism in peripheral tissues can be antagonized by other small
lipophilic agents, including 22(S)-hydroxycholesterol, certain unsaturated fatty acids, and
Syntéza ligandů
Jaderné receptory
Transaktivace
Metabolismus ligandů
Cílové geny
(metabolismus,
kontrola buněčné
proliferace,
diferenciace a
apoptózy)
Toxické látky, farmaka
??????????
Nízkomolekulární lipofilní sloučeniny jako ligandy = aktivita jaderných receptorů je
do značné míry závislá na syntéze a degradaci ligandů a naopak