Transcript ER stress and disease

ER stress and disease
introduction
• ER function: an organelle where secretory or
membrane proteins are synthesized.
----Nascent proteins are folding with the
assistance of ER chaperones located in ER, and
only correctly folded proteins are transported to
the Golgi apparatus.
----Unfolded or malfolded proteins are retained in
the ER, retrotranslocated to the cytoplasm by
ERAD.
• ER stress: When cells synthesize
secretory proteins in amounts that exceed
the capacity of the folding apparatus and
ERAD machinery, unfolded proteins are
accumulated in the ER. Unfolded proteins
exposed hydrophobic amino-acid residues
and tend to form protein aggregrates.
Final this evokes ER stress
ER stress response
The mammalian ER stress response consists of
four mechanisms
• Attenuation of unfolded proteins
• The transcriptional induction of ER chaperone
genes to increase folding capacity;
• The transcriptional induction of ERAD
component genes to increase ERAD capacity;
• Induction of apoptosis to safely dispose of cells
injured by ER stress to ensure the survival of the
organism.
ER stress- inducing chemicals
First group: glycosylation inhibitor
N-glycosylation of proteins for folding
Tunicamycin: antibiotic produced by
streptomyces lysosuperificus that
inhibitors by preventing UDP-GlcNAcdilichol phosphate GlcNAc-phosphate
transferase activity.
2-Deoxy-D-glucose is less efficient than
tunicamycin.
Second group: Ca2+ metablism disruptor
• Ca2+ ionophore (A23187) and Ca2+ pump
inhibitor (thapsigargin)
• Why: the high concentration of Ca2+ ion in
ER should be kept; and ER chaperones
such as BiP required Ca2+ ions
• The third group: reducing agents
DTT and β-mecaptoethanol
• The fourth group: hypoxia
ER chaperones
• Molecular chaperones
• Folding enzymes
Molecular chaperones
• BiP/GRP78s : belong to HSP70
BiP binds to the hydrophobic regions of
unfolded proteins via a substrate-binding
domain and facilitates folding through
conformational change evoked by the
hydrolysis of ATP by ATPase domain.
• Oxygen-regulated protein (ORP/GRP170)
HSP110 family, the mecanism is similar to
BiP.
• ERdj1, 3, 4, 5, SEC63 and p58IPK
belongs to HSP40 family, and modulate
the functions of BiP by regulating its
ATPase activity as a cocheperone.
• BiP-associated protein (BAP), which is a
member of GrpE family and also
modulates the function of BiP by
enhancing nucleotide exchange.
• GRP94 belongs to HSP90 family.
• FKBP13 is a peptidyl-prolyl isomerase
belongs to the FKBP family
• Calnexin and calreticulin are ER
chaperones specifically involved in the
folding of glycoprotein. They share a
similar molecular structure and function,
though they are transmembrane and
luminal proteins, respectively.
Folding enzymes
• Protein disulfide isomerase(PDI), ERp72,
ERp61, GRP58/Erp57, ERp44, ERp29
and PDI-P5.
Functions: oxidize cystein residues of
nascent bonds. Reduced folding enzymes
are reoxidized by ER
oxidoreductin(ERO1),which can use
molecularoxygen as teminal electron
acceptor.
ERAD
• Unfolded or malfolded proteins are
trapped by the ERAD machinery and
transported to cytoplasm.
• Retrotranslocated proteins are
ubiquitinated and degraded by
proteasome in the cytosol.
• The process of ERAD can be divided into
four step: recognition, retrotranslocation,
ubiquitination and degradation
recognition
• During the calnexin cycle, the oligosaccharide of
nascent residue is trimmed by a-mannosidase I,
nascent polypeptides with eight mannose
residues are released from calnexin or
calreticulin and bind to ER degradationenchaning a-mannosidase-like protein (EDEM),
which discriminates unfolded proteins from
folded proteins.
• EDEM1(membrane), EDEM2, EDEM3(luminal)
contain mannosidase-like domain, which
recognite mannose residues.
• Osteosarcoma 9 (OS9) specially binds to
unfolded glycoproteins containing eight or
five mannose residues and unglycosylated
unfolded proteins. OS9 and XTP3transactivated gene B(XTP3B) contain the
mannose—phosphate receptor-like
domain, which may be critical to the
recognition of mannose residues.
Retrotranslocation
• Nascent glycoproteins recognized by EDEM and
OS9 as misfolded are destined for the
retrotranslocation. Before retrotranslocation,
nascent proteins associate with PDI and BiP to
cleave disulfide bonds and to unfold the partially
folded structure, respectively.
• Retrotranslocation machinery is elusive
• Sec61 , Derlin-1associated with p97 through an
adaptor protein, valocin-containing
protein(VCP)-interacting membrane protein1.
• P97/cdc48/VCP is a cytosolic AAA-ATPase and
recruits unfolded ER proteins to the cytosol.
Ubiquitin fusion degradation protein 1 (Ufd1) and
nuclear protein localization 4 (Npl4) bind to p97
as cofactor and help p97 to extract unfolded
proteins. The polypeptide portion of unfilded
proteins interacts with p97, whereas the poly-Ub
chains attached to them are recognized by
bothe p97 and Ufd1 and may activate the
ATPase activity of p97.
Ubiquitination
Retrotranslocated proteins are ubiquitinated by E1-E2-E3
ubiquitin system. Ub is first conjugated to E2 by E1, and then
transferred to ERAD substrates by E3.
E1:Ub activating enzyme that is ubiquitously involved in
protein degradation.
E2: UBC2 and UBC7.
ERAD-L and READ-C
HMG-CoA reductase degradation protein 1(HRD1,preference
for substrate that contain misfolded luminal domain), gp78
and TEB4/Doa10 (prefer transmembrane proteins containing
misfolded cytosolic) are membrane-anchored E3;FBX2 (F-box
only protein 2) spescially recognizes N-glycosylated proteins
located in cytosol.
Parkin: both ERAD-L and ERAD-C
Degradation
Retrotranslocated and ubiquitinated proteins are
deglycosylated by peptide-N-glycanase before their
degradation by the proteasome,
Why: bulky glycan chains hampering effect
Peptide-N-glycanase is associated with Derlin-1
(cotrotranslocationally)
DSK2 and Rad23 facilited the delivery of substrates into
proteasome
Response pathways for ER stress
The mammalian ER stress response has four
mechanisms: (1) translational attenuation; and
enhanced expression of (2) ER chaperones
and (3) ERAD components; (4) induction of
apoptosis.
These responses are regulated by the
regulatory pathways.
PERK pathway
PERK is transmembrane protein located in ER, which sense the
acculumation of unfolded proteins in ER luminal.
PERK: luminal portion
cytosolic portion contains a kinase domains
ER stress(-): BiP binds to luminal PERK luminal domain.
ER stress(+): BiP release from PERK, and PERK is activated through
oligomerization and trans-phosphorylation.
Activated PERK phosphorylates and inactivates the a-subunit of eIF2
ATF6 pathway
• ATF6, on ER membrane, its luminal domain responsible
for the sensing of unfolded proteins, its cytosolic domain
has bZIP and a transcriptional domain.
ER stress(-): BiP bind to the luminal domain of ATF6 and
by hinders the Golgi-localization signal, leading to
inhibition of ATF6 translocation
ER stress (+):BiP dissociates from ATF6, and ATF6 is
moved to Golgi apparatus by vasicular transport, then
cleaved by S2P and S1P. The resultant cytoplasmic
portion of ATF6 translocates into nucleus.
pATF6(N) : BiP, GRP94 and calreticulin
CCAAT(NF-Y)-(N9)-CCACG (pATF6(N)
IRE1 pathway
• The third sensor molecule is IRE1 (inositol requirement 1): IRE1a
and IRE1β
• Cytosolic domain contains a kinase domain and Rnase domain
• ER stress (-): BiP bind to its luminal domian
• ER stress (+): BiP suppression of IRE1 activation is released, IRE1
is activated through dimerization and transphosphorylation.
• Activated IRE1a convert XBP1 pre-mRNA into mature mRNA by
cytosolic splicing.
• pXBP1(S) not pXBP1(U) : activation of ERAD component genes
such as EDEM, HRD1, Derlin-2 and -3 through a cis-acting element;
induce the expression of protein involved in lipid synthesis and ER
biogensis, as well as ER chaperons, such as BiP, p581PK, ERdj4,
PDI-P5 and HEDJ.
Apoptosis-inducing pathway
• If PERK, ATF6 and IRE1 pathways can
not suppress ER stress, an apoptotic
pathway is triggering to ensure survival of
organism as a last line of defense.
---CHOP/GADD153 pathway
---IRE1-TRAF2-ASK1 pathway
---Caspase 12 pathway
ER stress-related disease
• Neurodegenerative disease
AD, PD, polyQ disease, Prion disease,ALS
• Bipolar disorder
Expression of XBP1 and BiP wane
• Diabetes mellitus
• Atherosclerosis
• Inflammation
• Ischemia
• Heart disease
• Liver disease
• Kidney disease
• Viral infection