what are the results or diseases caused by the endoplasmic reticulum dysfunction

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[Type the document title]
ENDOPLASMIC RETICULUM
DYSFUNCTION

2. 1
Table of content
ER stress signaling………………………..…………………………………………………… 2
Cerebral hypoxia…………………………………………………………………………………3
Inflammation and endoplasmic reticulum stress in obesity and diabetes……………………5
Genesis of endoplasmic reticulum stress in Huntington’s disease……………………………8
Endoplasmic reticulum and Alzheimer’s disease…………………………………………….11
Endoplasmic reticulum and Parkinson’s disease…………………………………………….14

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ROLE OF ER IN METABOLIC DISODERS AND ITS DYSFUNCTION IN
NEUROLOGICAL DISEASES
ER STRESS SIGNALLING
The endoplasmic reticulum (ER) is the cellular organelle that is critical for protein folding and
secretion, calcium homeostasis, and lipid biosynthesis. The ER is the site of multiple post-
translational modifications such as glycosylation and disulfide bond formation. It is also the
organelle in which proteins are folded into their proper conformation and in which multi-subunit
proteins are assembled. Under various conditions, called ER stress, protein folding in the ER is
impaired leading to the accumulation of misfolded proteins. Multiple cellular disturbances can
cause ER stress including disturbances in redox regulation, calcium regulation, glucose
deprivation, and viral infection.
The accumulation of misfolded proteins in the ER is harmful to cells and thus the ER has
evolved mechanisms designed to detect misfolded proteins and either refold them or target them
for degradation. The accumulation of misfolded proteins in the ER triggers an evolutionarily
conserved program called the unfolded protein response (UPR) which is designed to clear the ER
of misfolded proteins and restore ER homeostasis. This response includes the attenuation of
protein translation to lessen the protein processing load in the ER. There is also an upregulation
of genes involved in ER protein folding including chaperones such as BiP/GRP78, enzymes
mediating folding such as protein disulfide isomerase, ER structural components, and
components of the ER-associated degradation pathway (ERAD). Proteins that cannot be refolded
into their correct conformation are targeted to the ER-associated degradation (ERAD) pathway
where they are ubiquinated and degraded via the proteasomal system.
The ER stress response is initiated when the capacity of ER-resident chaperone proteins is
exceeded by the load of misfolded proteins. Protein chaperones like BiP/GRP78, Grp 94, and
calreticulin assist in proper protein folding. BiP/GRP78, a trans-membrane protein that spans the
ER lumen, associates with the UPR sensors ATF6, IRE1, and PERK and represses their activity.
The accumulation of misfolded proteins results in the saturation of ER chaperones, including
BiP/GRP78. The sequestering of BiP by misfolded proteins results in the loss of BiP-mediated
repression of the UPR sensors resulting in their activation. Prolonged ER stress typically results
in cell death by apoptosis. ER stress occurs in both normal and patho-physiological conditions.
The efficient development of plasma cells from B cells requires components of the UPR.
However, ER stress has also been implicated in multiple disorders such as type 2 diabetes,
ischemia, and neurodegenerative disorders. ER stress can also be induced by hypoxia. This has
implications for solid tumors which usually exhibit hypoxia in their cores. SCID mice implanted
with tumor-forming cells that lack UPR components generate smaller tumors than those
implanted with wild-type controls suggesting that the UPR contributes to cancer cell survival.
Insulin resistance is also associated with ER stress and the treatment of type 2 diabetic mice with
chemical chaperones which assist protein folding in the ER restored insulin sensitivity.
CEREBRAL HYPOXIA
Our brain depends on oxygen. Even just a minute or so of oxygen deprivation can cause a
cascade of reactions that damage your brain. Cerebral anoxia occurs when your brain is
completely deprived of oxygen. When oxygen flow is reduced—usually due to reduced blood
flow—but not completely eliminated, cerebral hypoxia is the result.

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In some cases, your brain responds to the loss by temporarily increasing blood flow in an attempt
to provide more oxygen. Though this can save your life if your brain is deprived of oxygen, it
can also cause cardiovascular episodes such as brain bleeding, strokes, and ruptured blood
vessels in the brain, particularly if you have other cardiovascular problems.
Cerebral hypoxia can cause both immediate and long-term brain damage, and the course of
recovery from a serious episode of cerebral hypoxia is unpredictable. Only a doctor can properly
assess you, so if you suspect your brain has been deprived of oxygen, seek immediate medical
care.
Types of Cerebral Hypoxia
Doctors classify hypoxia of the brain into four distinct categories, ranging from least to most
severe:
 Diffused cerebral hypoxia causes mild to moderate impairment in brain function due to
low blood oxygen levels. This sort of hypoxia is common among people who hold their
breaths for too long, or who participate in sports that involve choking one's opponent,
such as jiu-jitsu.
 Focal cerebral ischemia occurs when there is oxygen deprivation in a specific area of
the brain. This is usually the result of a hemorrhage, stroke, or blockage in a single blood
vessel.
 Global cerebral ischemia is a complete cessation of blood flow to the brain, and quickly
leads to cerebral anoxia. Severe strokes, traumatic injuries such as gunshot wounds,
choking, and suffocation can cause global cerebral ischemia.
 Cerebral infarction is cessation of blood flow to multiple areas of the brain, and often
causes extensive brain damage. A stroke is the most common cause.
Symptoms of Cerebral Hypoxia
Cerebral hypoxia is a medical emergency, and victims often know the cause, particularly if
they've fallen or suffered another traumatic injury. When cerebral hypoxia is due to a stroke or
other internal issue, symptoms can appear more slowly.
Some warning signs that your brain has been deprived of oxygen include:
 Feeling light-headed.
 Intense pressure in the brain or reddening of the face; this suggests the body is increasing
blood flow to compensate for hypoxia. Some people experience very painful headaches.
 Loss of consciousness.
 Changes in mood, personality, or judgment. People suffering from hypoxia may make
poor decisions, suddenly forget words, or not know where they are.
 Weakness, particularly on one side of the body. It's common for people having a stroke to
be unable to raise both arms above the head, to have a crooked smile, or to experience
paralysis on one side of the face.
 Sudden bleeding anywhere around the face, particularly if blood vessels in the eyes are
ruptured.
Short and Long-term Effects of Cerebral Hypoxia
The effects of cerebral hypoxia depend primarily on how long the brain is deprived of oxygen.
Short-term diffuse hypoxia often produces no effects at all. For instance, a wrestler who loses
consciousness with his opponent's arm around his neck will likely regain full functioning after he
regains consciousness. People who experience this sort of short-term oxygen deprivation,
though, will still experience symptoms. Those include:

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 Loss of sensation in one or more areas of the body.
 Confusion, memory difficulties, or impairments of judgment.
 Loss of consciousness.
 Blurred vision or difficulty focusing the eyes on a single point.
 Feeling nauseated or woozy.
 A headache during or after the period of hypoxia.
When hypoxia lasts less than 60 seconds, it is unlikely to cause lasting damage. At two minutes,
the risk of brain damage becomes more likely, while at three to four minutes, it becomes a near-
inevitability. The long-term effects of cerebral hypoxia can include:
 Damage to specific areas of the brain. The specific prognosis depends on which areas are
damaged. For instance, severe damage to regions of the brain that govern speech and
language may lead to aphasia.
 Long-term loss of consciousness in the form of a coma. Some patients also enter a
persistent vegetative state. This loss of consciousness may give the brain time to heal, but
can also be a permanent state.
 Epilepsy or persistent seizures.
 Damage to motor skills, especially fine motor skills. Sometimes this damage is localized
to just one region or one side of the body.
 Death, either immediately after the deprivation or due to the side effects of hypoxia, such
as stroke or other cardiovascular episodes.
 Birth defects; hypoxia is a relatively common birth injury, and newborns who suffer
prolonged oxygen deprivation may suffer chronic diseases such as cerebral palsy.
Treatment for Cerebral Hypoxia
The most important treatment for cerebral hypoxia involves removing the source of the oxygen
deprivation. Blood clots might need to be removed, or the patient might need the assistance of a
ventilator until the source of the oxygen deprivation can be discovered.
Thereafter, there is no specific treatment for cerebral hypoxia. The brain remains a mysterious
organ, and we do not yet know how to reverse brain damage or regenerate brain cells—though
experimental research has shown some promising results. Instead, doctors focus on addressing
the symptoms of cerebral hypoxia. This typically means extensive physical, occupational, or
speech therapy to teach your brain how to work around any damaged areas. Such therapy can be
challenging and emotionally draining, but the more committed you are to challenging your brain,
the more likely it is that you will see improvements in functioning. Some other treatments
include:
 Drugs to prevent future hypoxia episodes; this may include the use of blood thinners.
 Antibiotics to treat infections that caused or resulted from the hypoxia.
 Surgery to remove any blockages or to discover the source of the blockage.
 The use of assistive gear, such as a wheelchair, to help you work around hypoxia-related
motor skill deficits.
 Psychotherapy to help you and your family find effective ways to cope with the long and
short-term effects of your injuries.
In industrialized countries stroke is the third biggest killer and the leading cause of disability in
adults. Acute ischaemia is the main cause of neuronal loss and, although its pathology is
complex, appears to involve ER stress. During cerebral ischaemia, neuronal depolarization due

6. 5
to energy depletion causes the uncontrolled release of glutamate. The consequent activation of
NMDA receptors on nearby neurons generates further glutamate release and causes an ischemic
depolarization wave to spread outward from the initial site of damage leading to widespread
disturbance of calcium homeostasis. Unsurprisingly, many studies implicate ER calcium store
release in the resulting excitotoxic death.
The ER is the main site for calcium storage within the cell and so its chaperones have evolved to
function efficiently in this high calcium environment; indeed many require high calcium to
function. Calcium is pumped into the ER by the sarco-endoplasmic reticular calcium-ATPase
(SERCA) and released back into the cytosol by the inositol trisphosphate receptor (IP3R) in
response to extracellular signals and by the ryanodine receptor during calcium-induced calcium
release. Inhibition of the SERCA pump by thapsigargin induces apoptosis in many cell types
including neuroblastoma cells and is often used to induce ER stress in the experimental setting.
During cerebral ischemia, energy depletion leads to failure of the SERCA pump and thus
redistribution of ER calcium into the cytosol. This delivers a double blow with combined toxicity
from uncontrolled cytosolic calcium and ER stress due to chaperone dysfunction. The UPR is
activated in many rodent models of cerebral ischemia. In these models, protein synthesis is
rapidly inhibited.
When taken together, these observations suggest that ER stress is induced by cerebral ischemia
and that the UPR limits cerebral infarct size. There is good evidence to suggest that manipulation
of ER stress signaling by increasing may have important therapeutic effects as an acute -
phosphorylation of eIF2 intervention for cerebral ischemia.
INFLAMMATION AND ENDOPLASMIC RETICULUM STRESS IN OBESITY AND
DIABETES
Obesity is associated with chronic low-grade inflammation. Inflammatory signals interfere with
insulin action and disrupt metabolic homeostasis. The c-Jun N-terminal kinase (JNK) has been
identified as a central mediator of insulin resistance. Recent studies showed that in obesity
compromising endoplasmic reticulum (ER) function results in insulin resistance and type 2
diabetes that are dependent on JNK activation.
Obesity is associated with chronic, low grade, inflammatory responses in metabolically active
sites, most notably, adipose tissue. This increased chronic inflammatory status triggered by
metabolic cues, which differs from the classic inflammation, is a critical link between obesity
and other associated pathologies, such as insulin resistance and type 2 diabetes. The principal
mechanism by which the inflammatory signals interfere with insulin action involves
posttranslational modification of insulin receptor substrate molecules, particularly through serine
phosphorylation. This modification is essentially universal to all forms of insulin resistance
whether they are chemically or genetically induced in cells, animal models, or human disease. In
efforts to identify the mechanisms leading to insulin resistance in general and serine
phosphorylation of insulin receptor substrate molecules in particular, we have previously
identified c-Jun N-terminal kinase (JNK) as a central mediator of insulin resistance in cultured
cells and obese animal models. These studies showed that obesity results in marked JNK
activation in insulin-sensitive tissues, such as fat and liver,
Genetic deletion of JNK1 gene in mice resulted in marked protection against insulin resistance,
type 2 diabetes and fatty liver disease. Furthermore, blocking JNK activity using chemical,
biochemical, or molecular strategies in obese mice also results in enhanced insulin sensitivity
and correction of hyperglycemia, indicating a potential for utilizing JNK inhibition as therapeutic

7. 6
strategy against type 2 diabetes. The ER is a vast membranous network responsible for the
trafficking of a wide range of proteins. The ER is a principal site of protein synthesis, maturation
and, together with the Golgi apparatus, the transportation and release of correctly folded proteins.
As the ER plays a central role in integrating multiple metabolic signals critical in cellular
homeostasis, it is of paramount importance to the cell to maintain proper ER function and to
adapt organelle capacity to manage metabolic and other adverse conditions.Therefore, under
conditions that challenge ER function, particularly its folding capacity, the organelle has evolved
an adaptive response system known as the unfolded protein response. Conditions that may
trigger unfolded protein response activation include increased protein synthesis, the presence of
mutant or misfolded proteins, inhibition of protein glycosylation, imbalance of ER calcium
levels, glucose and energy deprivation, hypoxia, pathogens or pathogenassociated components
and toxins Given that the unfolded protein response is closely integrated with stress signaling,
inflammation and JNK activation, as well as the fact that obesity features many conditions to
challenge ER (from increase in synthetic demand to energy availability and fluxes), we
hypothesized that obesity may lead to a condition of ER stress in metabolically active tissues and
organs.
ER stress is a critical mechanism underlying obesity-induced JNK activity, inflammatory and
stress responses, and insulin resistance and offer potential new therapeutic opportunities against
obesity, insulin resistance and type 2 diabetes.
Metabolic disorders
The endoplasmic reticulum (ER) is the largest multifunctional organelle inside cells with
distinctstructure elements containing both smooth and rough ER. Some of the best-known ER
functionsinclude the production of lipids like cholesterol, glycerophospholipids, and ceramide,
the synthesis ofprotein, and the regulation of calcium storage and dynamics.
Metabolic stress is a condition characterized by insufficient or excessive nutrient supply
comparedto cellular bioenergetic needs. Excessive supply is characteristic of over nutrition that
representsa public health concern in developed and developing countries. Chronic metabolic
stress leads toa variety of metabolic diseases, including obesity, insulin resistance, diabetes, fatty
liver disease,and cardiovascular diseases. The two main types of nutrients that exert cytotoxic
effects via theiraction on ER are fatty acids (FA) and glucose.
ER Stress Induces IR and Diabetes
Chronic metabolic stress induces both ER and oxidative stress and is invariably associated with
Inflammation, an element of cellular stress response considered as a major cause of obesity,
insulinresistance (IR), and type 2 diabetes. These pathologies are characterized by a general
multi-organdysfunction, including liver, muscle, adipose tissue, brain, and pancreas, and ER
stress is associatedwith the dysfunction of these tissues. Some of the ER-linked mechanisms are
common to all these tissues, while others are cell type-specific.
Lipotoxicity
ER is involved in lipid metabolism, and lipotoxicity is one of the most important triggers of ER
stress in peripheral tissues. Although lipid toxicity on various cell types has been described for a
longtime, the mechanisms are still unclear, and ER stress might represent a relatively new
explanation forthese deleterious effects. Damage of beta cells by prolonged exposure to
circulating FA is consideredas a main cause of diabetes. High fat diet by itself is not sufficient to

8. 7
induce diabetes, but it contributesto the worsening of the diabetic condition in genetically
predisposed individuals.
GENESIS OF ER STRESS IN HUNTINGTON’S DISEASE
Recent research has identified ER stress as a major mechanism implicated in cytotoxicity in
many neurodegenerative diseases, among them Huntington’s disease. This genetic disorder is of
late-onset, progressive and fatal, affecting cognition and movement. There is presently no cure or
any effective therapy for the disease.
HUNTINGTON’S DISEASE
Huntington’s disease (HD) is a neurodegenerative disease arising from an expanded CAG repeat,
coding for a polyglutamine (polyQ) tract in the huntingtin (Htt) protein. It is a member of a quite
large family of polyQ diseases, such as several spinocerebellar ataxias and Machado- Joseph
Disease. HD is a genetic, autosomal dominant disease that causes motor dysfunction and
cognitive decline. These symptoms are progressive and usually of late onset, the age of onset
being inversely correlated with the number of glutamine repeats, from a minimum of about 35
repeats for Htt to be pathogenic, going up to over 100 repeats in early onset patients. The
mutation causes Htt aggregation and there is accumulating evidence that the toxic species are
intermediate oligomeric associations of Htt and not the final large aggregates. In a cell-protective
pathway, the aggregates can be cleared by autophagy, and interestingly, it was recently
determined that wild type Htt participates in the process of protein targeting to autophagy.
Mutant Htt was reported to interfere with the autophagic process in several ways; one being
through a deleterious effect on mTorc1 .The process of aggregation of mutant Htt interferes in
several other ways with normal cell metabolism and leads to cell death through a still unclear
mechanism. One of the implicated pathways has been glutamate receptor overstimulation, so-
called excitotoxicity, which activates calcium influx and cAMP response element binding protein
(CREB), leading to mitochondrial dysfunction. Mitochondrial damage can also come about from
oxidative stress in HD. Mutant Htt was also recently reported to inhibit protein import to
mitochondria. There is, in addition, interference with transport on microtubules, which affects
endoplasmic reticulum (ER)-Golgi traffic and axonal transport. There is also sequestration of
transcription factors and importantly, interference with the ubiquitin proteasome system (UPS) as
seen in cells in culture, in mouse HD models and in HD patients. The turnover of Htt is then
affected and may lead to its accumulation and aggregation .Interference with the UPS by mutant
Htt inhibits cytosolic protein degradation as well as ER-associated protein degradation (ERAD) .
ERAD is a pathway that normally reduces the protein load in the ER and inhibition of this
pathway leads to the accumulation of unfolded and misfolded proteins in the ER, which is
termed ER stress. ER stress causes activation of the unfolded protein response (UPR).Despite the
expression of mutant Htt in most cell types in HD patients, HD initially affects medium spiny
neurons in the brain striatum and only later regions of the brain cortex. This is a common feature
of many neurodegenerative diseases, where there is an unexplained high sensitivity of certain
specific regions or cell types of the central nervous system. The reasons for the special sensitivity
of striatal cells in HD are unclear; mechanisms have been proposed, involving enhanced
expression of proteins in these cells and also, recently, of long non-coding RNAs. As we will see
later, striatal neurons have distinct features in their UPR.

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GENESIS AND IMPACT OF ER STRESS IN HUNTINGTON’S DISEASE
As mentioned above, interference with the UPS by mutant Htt inhibits degradation of proteins
from the cytosol as well as from the ER, which are targeted to ERAD Inhibition of ERAD leads
to the accumulation of unfolded or misfolded proteins in the ER, or in other words, causes ER
stress, which in turn activates the UPR. UPR induction was observed by expression of mutant
Htt in yeast and mammalian cells. It has also been reported in animal models of HD. P97
depletion by mutant Htt appears to be a major cause in the inhibition of ERAD, which causes ER
stress, as p97 overexpression was sufficient for complete compensation in mammalian cells.
Overexpression of the p97 cofactors Npl4 and Ufd1 also reduced mutant Htt toxicity in yeast. It
was also reported that mutant Htt interacts with the ER E3 ligase gp78, inhibiting ERAD.
However, gp78 is not the major pathway to ERAD, whereas p97 is an essential factor for the
process. Interactions of Htt with ER membrane-bound p97 and with transmembrane gp78 may
explain the finding of mutant Htt associated with the ER membrane. Other mechanisms were
also suggested that could lead to ER stress in HD, such as impaired ER-Golgi traffic, inhibition
of autophagy and calcium deregulation , but evidence is scarce. Besides the products of the
classic UPR-induced genes (ATF6, BiP, protein disulfide isomerase, CHOP, etc), other proteins
that are induced by ER stress have recently been linked to HD pathology and areupregulated in
HD patients, Rrs1 and SCAMP5, the latter especially upregulated in the striatum. We recently
showed that the onset of ER stress is due to soluble Htt forms and correlates with the formation
of Htt oligomers, preceding the formation of visible inclusions. ER stress levels did not increase
in response to the presence and growth of large aggregates, but ER stress was actually reduced
with time, implying a protective role for these aggregates. Htt regions that bind and sequester
cellular factors may be exposed in the oligomeric state. These aggregation-prone regions could
be hidden and protected inside the structure of the large amyloid aggregates, similar to what has
been found in other neurodegenerative diseases. This is consistent with increasing evidence that
Htt oligomers and not aggregates are the cytotoxic species in HD and the reports of UPS
inhibition before Htt inclusion into large aggregates. This might explain why clinical trials of
anti-aggregating molecules in HD have been so far unsuccessful. The presence of toxic
oligomeric forms (detected with specific antibodies), was found to predict neuro-degeneration.
Apoptotic pathways induced through ER stress have been suggested in HD pathology through
induction of CHOP and also of ASK1, leading finally to caspase activation. Mutant Htt causes
altered calcium signaling and apoptosis, possibly by its interference with the ER IP3R. This
effect may be downstream of the UPR or by direct interaction with IP3R. Interestingly, as
mentioned before, the IP3R is mostly located at the MAM, and autophagy, which can be induced
by the UPR, has also been reported to initiate at this region. Apoptotic and autophagic pathways
might then be induced in parallel by mutant Htt at the MAM. Sigma-1R, also at the MAM, was
recently reported to have a protective effect in cells expressing mutant Htt, increasing UPS
function and Htt degradation.
We recently showed that striatal neurons are especiallysensitive to ER stress . Their PERK
pathway is altered, with very reduced PERK activity and low phosphorylationof eIF2α, a
characteristic that we also found in WT mousebrain striatum. In contrast, a knock-in striatal cell
line expressing mutant Htt and the striatum of HD model mice showed higher levels of eIF2α-P.
Huntingtin toxicity in the mutant Htt expressing cells could be strongly reduced by inhibiting
PERK. This suggests on one hand a reason for the special sensitivity of the striatum in HD, and
on other it underscores the importance of ER stress for Htt cytotoxicity. As the dephosphorylated

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state of eIF2αwas linked to memory and long term potentiation, possibly to maintain high
translation rates, the appearance of ER stress upon expression of mutant Htt and the consequent
increase in eIF2α-P levels suggest the intriguing possibility that they are linked to the cognitive
impairment observed in HD. 7
Therapeutic Targeting
Several therapeutic strategies have been proposed for HD, but so far with no successful resulting
therapy. Gene therapy approaches have been suggested for the silencing of mutant Htt
expression, but they are far from implementation. Another strategy proposed recently is the
targeting of Htt cleavage by caspase 6, which gave positive results in a BACHD mouse model.
As a general strategy for reducing ER stress, there are reports that chemical chaperones,
including 4-PBA and TUDCA, hindered disease progression in HD mouse models, and
decreased ER stress levels in HD and other disease models . Reduction of eIF2α-P , or treatment
with a compound that restores translation downstream of eIF2α, thwarted prion-related disease in
mouse models. This is consistent with our results of PERK inhibition in knock-in striatal neurons
expressing mutant Htt, which considerably reduced cytotoxicity. Inhibition of the PERK
pathway could be a promising therapeutic strategy.
As explained above, Sigma-1R expression has a general cell protective effect and Sigma-1R
agonists have proven effective in mouse models of brain disease. Sigma-1R activation was
reported to induce neuronal re-growth and functional recovery following experimental stroke in a
rat model and a phase II trial was conducted with the Sigma-1R agonist cutamesine (SA4503) in
patients with ischemic stroke. A recent study showed that another Sigma-1R agonist, PRE084,
improved behavioral symptoms in a Parkinson’s disease mouse model. In another study, this
same agonist promoted cell viability, reduced oxidative stress and decreased cleavage of
caspases in mutant Htt-expressing cells, suggesting a possible therapeutic benefit of Sigma-1R
agonists in HD.
Many pathways could partake in mutant huntingtin cytotoxicity, but the fact that UPR
modulation, such as PERK inhibition or Sigma-1R activation, reduces significantly the toxicity,
implicates ER stress as a main factor. UPR modulation with novel drugs could thus be a
promising therapeutic approach for HD. The onset of ER stress, as a consequence of ERAD
inhibition through p97 depletion and other interferences, is linked to the formation of Htt
oligomers, whereas the formation of Htt large aggregates was shown to be protective. Therefore,
caution is advised in the development of inhibitors of aggregation, which might have an overall
detrimental effect.
ENDOPLASMIC RETICULUM AND ALZHEIMER'S DISEASE
Mitochondria participate in the pathogenesis of several neurodegenerative diseases, including
Parkinson's, Huntington's and the heterogeneous group of mitochondrial diseases of
mitochondrial and nuclear origin. Indeed, most of the metabolic processes that are performed in,
associated with, or controlled by mitochondria, are known to be perturbed in neurodegeneration
(Schon and Przedborski, 2011). However, mitochondria seem to have a less defined role in the
pathogenesis of AD, one of the most devastating neurodegenerative conditions (Morais and De
Strooper, 2010). In particular, while mitochondrial dysfunction has been extensively reported in
models of AD, as well as in cells from AD patients, the prevailing consensus is that this
represents an epiphenomenon of a dysfunctional neuron, secondary to well-characterized

11. 10
pathogenetic pathways at other cellular sites. Indeed, according to the currently prevalent
pathogenetic theory, AD results from a cascade of proteolytic events including PS1 and PS2, the
catalytic subunits of the γ-secretases, a collection of protease complexes involved in the
proteolysis of integral membrane proteins including the APP (De Strooper, 2010). Cleavage of
APP yields the well-known Aβ peptides of amyloid plaques, which provide, together with the
neuronal tangles, the characteristic pathological signature of AD. Aβ is upstream in a
pathological cascade that induces plaques, tangles, and finally leads to neurodegeneration and
dementia (De Strooper, 2010). In this hypothesis, PS1 and PS2 are linked to AD via their well-
established roles in Aβ generation and mitochondria may play a role downstream of Aβ,
providing for instance the signals that trigger caspase activation to cause apoptosis of the
neuronal spines (D'Amelio et al, 2011) or degeneration of the whole neuron (Hardy and Selkoe,
2002). However, the picture is complicated by a spatial paradox: how can this organelle be a
primary target of Aβ produced by the γ-secretase at the plasma membrane and in intracellular
vesicles? In this issue, Schon and colleagues try to solve this conundrum, showing that the γ-
secretase complex is enriched at the interface between mitochondria and the ER and that the
communication between the two organelles becomes dysfunctional in models of AD .
Schematic representation of the changes in the ER–mitochondria interface in models of AD.
Left: the normal ER–mitochondria interface; the known molecular tethers and PS1 and PS2 are
shown. Right: the closer juxtaposition observed in models of AD, where PS is lacking. Note that
the MAM has increased in size. Mfn, mitofusin; Drp1, dynamin-related protein 1; IP3R, inositol
triphosphate receptor; VDAC, voltage-dependent anion channel; GRP75, heat-shock protein 75;
PS, presenilin; the orange region on the ER illustrates the MAM.
Area-Gomez et al (2009) build on their previous findings that PS might be localized in
mitochondria-associated membranes (MAMs). MAMs are specialized subdomains of the ER,
originally identified by Vance (1990) as regions of the ER that are in close contact with
mitochondria. Mitochondria and ER are known to engage in close connections that are essential
for several functions shared by the two organelles: transfer of Ca2+, lipid metabolism,
morphology of mitochondria and the control of apoptosis and autophagy (de Brito and Scorrano,
2010). The functional importance of the ER–mitochondria interface was originally recognized by
laboratories studying the biosynthesis of lipids, often performed by pathways that are shared by
the two organelles and that require the transfer of intermediates between the two (Vance, 1990).
Subsequently, the interface was shown to be a crucial site for Ca2+ signalling, where
microdomains of high concentration of this ion are generated upon release by the ER and taken
up by the neighbouring mitochondria (Rizzuto et al, 1993). Despite the increasing awareness of
the importance of this region in physiology, its structural composition remains largely
uncharacterized. Physical bridges between mitochondria and ER have been imaged by electron
tomography (Csordas et al, 2006) and several proteins are known to be enriched in this interface.
So far the only structural component identified in mammalian cells to physically link ER and
mitochondria is the mitochondria-shaping protein mitofusin 2 (Mfn2), which is essential for the
establishment of a close interface that functions in Ca2+ and lipid transfer (de Brito and
Scorrano, 2008).
While MAMs are operationally defined as a membrane fraction that can be isolated following
specific differential centrifugation procedures, their nature remains largely elusive. Schon and
colleagues provide evidence that these regions float in sucrose gradients when extracted in
specific detergents, which characterizes them as raft-like domains. Lipid rafts are specialized

12. 11
domains enriched in cholesterol and sphingolipids (Lingwood and Simons, 2010). Lipid rafts
have been characterized extensively in the plasma membrane, where they are crucial platforms
for the assembly of signalling complexes. Interestingly, proteins are targeted to MAMs by
certain lipidation modifications that are also known to facilitate targeting of proteins to plasma
membrane lipid rafts (Lynes et al, 2012). Notably, the components of the γ-secretase complex
are known to be enriched in lipid rafts (Vetrivel et al, 2004); however, the activity of this
complex at the plasma membrane is low, suggesting that other intracellular membranes with
similar biophysical properties could be the major physiologically relevant location for γ-
secretase. Area-Gomez et al found that the active complex can be retrieved with MAMs, along
with the structural components of the γ-secretase, including PS.
Since mutations in PS are the major cause of inherited AD, this raises the interesting questions
whether PS is involved in MAM function and whether this function is disturbed in familial AD.
One function of MAM is intermediary lipid metabolism and a series of remarkable observations
were made in PS knockout fibroblasts in this regard. The cells accumulate lipid droplets, display
increased cholesterol and cholesterylester levels, and synthesize phospholipids at increased rates
(Area-Gomez et al, 2012). At the morphological level, ER–mitochondrial contacts were two- to
three-fold larger, showing that genetic deletion of PS results in dramatic effects on MAM
organization. Interestingly, in a set of genetic experiments, Schon and colleagues found that PS
and Mfn2, the master regulator of ER–mitochondria juxtaposition, work in the same pathway.
Upon ablation of Mfn2, the activity of the γ-secretase complex was reduced, despite PS still
being localized in MAMs. Downregulation of Mfn2 in PS-deficient cells corrected the increased
juxtaposition between ER and mitochondria and complemented all the lipid metabolism defects
that characterize cells lacking PS. Conversely, ablation of PS in an Mfn2-deficient background
restored the juxtaposition between the two organelles, indicating that in the absence of PS,
mitochondria and ER can come in close contact even when Mfn2 is lacking. These results are
important for the elucidation of the cell biology of the ER–mitochondrial interface, as they
suggest that the presence of PS itself is a repulsive factor. Likely, other tethers are unveiled upon
PS ablation: maybe at steady state the products of γ-secretase activity at the MAMs somehow
interfere with the function of these uncharacterized tethers, in a mechanism that uses for instance
rhomboid proteases to control this crucial cellular process.
The importance of the observations of Area-Gomez et al for AD was accentuated by
demonstrating that fibroblasts derived from patients with mutations in PS genes display similar
phenotypes. To a certain extent, it is puzzling that similar phenotypes were also retrieved in
fibroblasts derived from patients suffering from sporadic AD. These patients have no known
mutations in PS, and the MAM dysfunction must therefore be caused by other molecular
pathways. How the different causes of AD (PS mutations, APP mutations, APOE4, deficient
clearance of Aβ and other unknown factors) all converge on MAM is indeed an intriguing and
important question to be addressed in the future.
When Schon and colleagues investigate to what extent the proteolytic function of PS is involved
in MAM functions, the answers remain unclear. On the one hand, overexpression of catalytically
inactive PS mutants could not rescue the morphological and lipid metabolism defects of the
knockout cells, which suggests the involvement of the enzymatic activity of the complex.
However, the authors point out this correctly, experiments with such dominant-negative forms of
PS can be difficult to interpret. High overexpression will not only lead to the generation of

13. 12
catalytically inactive enzyme but also to an altered stoichiometry of complex subunits and thus
potentially to the generation of partially assembled complexes. Thus, such dominant-negative
mutants can affect the function of γ-secretase at several levels and the interpretation of the results
can become very complicated. The problem can be controlled by performing native gel
electrophoresis and other experiments to monitor the effects of overexpression, but the authors
have chosen an alternative approach using a γ-secretase inhibitor to evaluate the role of
proteolytic function in MAM. The inhibitor affects lipid metabolism in wild-type cells in a
similar way as that observed in the knockout cell line, suggesting that the proteolytic activity of
γ-secretase is involved in that aspect of MAM function. However, morphological alterations of
the ER–mitochondria contacts (MAM) were not observed in the treated cells, showing separation
of effects on morphology and on lipid metabolism. Thus, how precisely PS is involved in MAM
integrity, and how it genetically interacts with Mfn2 remains unclear.
The observations of Schon et al are for the time being difficult to integrate with the canonical
knowledge in the AD field. At the cell biological level for instance it is quite difficult to
understand how fully glycosylated γ-secretase complex would become integrated in the ER–
mitochondrial interface. The nicastrin subunit of the active γ-secretase in particular is heavily
glycosylated in the trans-Golgi. How the mature complex returns from there to the ER–
mitochondrial interface cannot be explained at the moment. The same holds true for highly
glycosylated APP, which appears to be at least partially processed by the γ-secretase at the
MAMs (Area-Gomez et al, 2009). However, in yeast, an ER component of the ERMES complex
that tethers ER and mitochondria is also glycosylated, suggesting that unknown pathways exist to
allow MAM localization of glycosylated proteins (Kornmann et al, 2009).
Since alterations in the ER–mitochondrial interface are also observed in fibroblasts from
sporadic AD patients, they might have a broad relevance in AD pathogenesis. Further work is
however necessary to consolidate some of the conclusions proposed in the paper. For example,
what happens in neurons and in the brain of AD patients at the level of the MAM? Can the
alterations in MAM functions be subject to drug targetting? For example, selective inhibitors of
Mfn2 could revert the increased ER–mitochondria juxtaposition and ameliorate AD if the
observations in the current manuscript hold true in the intact organism. At the end, finding
effective drugs is the ultimate goal of AD research and the paper of Schon and coworkers might
indicates a possible new horizon in the biology and pharmacology of AD.
ER AND PARKINSON’S DISEASE
Mitochondria form close physical contacts with a specialized domain of the endoplasmic
reticulum (ER), known as the mitochondria-associated membrane (MAM). This association
constitutes a key signaling hub to regulate several fundamental cellular processes. Alterations in
ER–mitochondria signaling have polytrophic effects on a variety of intracellular events resulting
in mitochondrial damage, Ca2+ dyshomeostasis, ER stress and defects in lipid metabolism and
autophagy. Intriguingly, many of these cellular processes are perturbed in neurodegenerative
diseases. Furthermore, increasing evidence highlights that ER–mitochondria signaling
contributes to these diseases, including Parkinson’s disease (PD).
PARKINSON’S DISEASE:
PD is the second most common neurodegenerative disorder, for which effective mechanism-
based treatments remain elusive. Several PD-related proteins localize at mitochondria or MAM

14. 13
and have been shown to participate in ER–mitochondria signaling regulation. Likewise, PD-
related mutations have been shown to damage this signaling.
 Endoplasmic reticulum (ER) and mitochondria form close associations that constitute key
signaling hubs to regulate many cellular processes.
 ER–mitochondria contacts regulate many different pathways, which are damaged in
Parkinson’s disease (PD).
 ER–mitochondria associations are altered in PD.
Parkinson’s disease (PD) is the most common movement disorder and the second most common
neurodegenerative disease after Alzheimer’s disease (AD). PD patients typically experience
difficulties with slowness of movements (bradykinesia), involuntary shaking (tremor), increased
resistance to passive movement (rigidity) and postural instability. The cardinal motor symptoms
of PD are attributable to the progressive degeneration of dopaminergic neurons in the pars
compact of the substantia nigra (SNpc DA). PD is also characterized by the presence of
intraneuronal proteinaceous inclusions called Lewy bodies (LB) and abnormal dystrophic
neuronal processes termed Lewy neurites in the surviving neurons.
ENDOPLASMIC RETICULUM–MITOCHONDRIA ASSOCIATIONS:
In the eukaryotic cell, communication and cooperation between the different membrane-bound
organelles must take place to integrate cellular physiology. This integration depends upon
effective crosstalk and one way in which this is achieved is through direct membrane contact.
Thus, proper endoplasmic reticulum (ER)–mitochondria communication requires the formation
of specialized membrane micro domains at the contact sites, defining short distances between
membranes to connect them6. The ER and mitochondria association is the most studied and the
first described inter-organelle contact7. The ER is closely opposed to 5–20% of the
mitochondrial surface. The ER domain specialized in this association is known as mitochondria-
associated membranes (MAMs) and can be smooth or ribosome-containing rough ER
membranes.
ER–MITOCHONDRIA TETHERING COMPLEXES:
The presence of structures that appear to tether the two organelles has been observed by electron
microscopy in many different cell types. Early studies revealed the proteinaceous nature of the
tethers between the two membranes6, 14. Studies in yeast revealed the presence of a protein
complex, known as ERMES (ER–mitochondria encounter structure) 15. However, no
mammalian orthologues of ERMES proteins have been identified yet; on the contrary, several
different protein complexes have been proposed as ER–mitochondria tethers16. One of these
complexes is based on the interaction between the ER Ca2+ channel IP3R (inositol 1,4,5-
trisphosphate receptor) and the OMM VDAC1 (voltage-dependent anion channel 1), that is the
major mitochondrial Ca2+ transport channel, in a ternary binding complex with the
mitochondrial chaperone GRP75 (glucose-regulated protein 75)17. The ER sorting molecule
PACS-2 (phosphofurin acidic cluster sorting protein-2) has also been shown to be involved in
ER–mitochondria associations. Similarly, the interaction between the ER protein Bap31 (B-
cell receptor associated protein 31) and the mitochondrial fission protein Fis1 has been shown
to bridge the mitochondria and the ER and promote apoptosis.

15. 14
Multiple structures that tether mitochondria with the mitochondria-associated membranes
(MAMs) of endoplasmic reticulum (ER) have been described. Inositol 1,4,5-trisphosphate
receptor (IP3R) and voltage-dependent anion channel (VDAC1) interact via GRP75.
Synaptojanin 2 binding protein (SYNJ2BP) interacts with the ribosome-binding protein 1
(RRBP1). The outer mitochondrial protein tyrosine phosphates-interacting protein 51 (PTPIP51)
interacts with vesicle-associated membrane proteins-associated protein B (VAPB) or oxysterol-
binding protein-related proteins (ORP5/8) at the ER. B-cell receptor associated protein 31
(BAP31) binds to mitochondrial fission 1 protein (Fis1). ER-located mitofusin 2 (MFN2)
interacts with mitochondrial MFN1/MFN2. Other proteins, such the ER sorting molecule
phosphofurin acidic cluster sorting protein-2 (PACS-2), have been involved in ER–mitochondria
association integrity. Yeasts specific proteins have also been described: the ER–mitochondria
encounter structure (ERMES) complex composed of four proteins: the outer mitochondrial
membrane proteins Mdm10 and Mdm34, the ER protein Mmm1, and the cytosolic protein
Mdm12.
ER–MITOCHONDRIA SIGNALING IN NEURODEGENERATION:
Neurodegenerative diseases including PD, AD, and ALS/FTD (front temporal dementia) share
several obvious features: they are characterized by progressive nervous system dysfunction,
affect millions of people worldwide and there is still no cure for any of them. Furthermore,
despite affecting different brain regions PD, AD, and ALS/FTD also share other characteristics
suggesting that common cellular processes may converge.
Thus, whilst the precise mechanisms remain to be determined, a variety of cellular processes are
damaged in all of them, including Ca2+ dysregulation, defects in axonal transport,
neuroinflammation, loss of cellular proteostasis and mitochondrial dysfunction. Remarkably,
ER–mitochondria associations, regulates all of those processes. The findings that alterations in
ER–mitochondria associations occur in neurodegenerative diseases have given rise to the
hypothesis that damaged ER–mitochondria signaling is a common potential therapeutic target
amongst distinct age-dependent neurodegenerative disorders.

16. 15
ER–mitochondrial axis appears to be essential for the healthy neurons. Conversely, the
disruption of this interaction may involve the develop of some processes as: mitochondrial
dysfunction, induction of oxidative stress, calcium (Ca2+) dyshomeostasis, autophagy defects or
neuro-inflammation, which induce neuronal damage and trigger neurodegenerative diseases as
PD.
CONCLUSIONS AND FUTURE:
Although the exact pathological mechanisms underlying PD remain largely unclear a plethora of
cellular pathways are known to be damaged. The discovery that ER–mitochondria signaling,
which regulates many of those pathways, are also damaged in PD has highlight the possibility of
a common link among them. Therefore, ER–mitochondria signaling may represent a possible
drug target upstream of those pathways. However, more research should be done before gaining
a clearer understanding of the links between ER–mitochondria signaling and the pathogenesis of
PD. Hence, many questions remain unclear. Although the evidence discussed here supports the
hypothesis that deregulation of ER–mitochondria signaling has an important role in PD
pathogenesis, it is still unclear as to whether ER–mitochondria associations are either up
regulated or disrupted upon PD-related insults. Combined, the findings reviewed above highlight
the complexity of studying ER–mitochondria associations. Therefore, additional research is
needed to gain further insight into the mechanisms of tethering of both organelles, especially in
relation to neurons.
Furthermore, investigating whether other PD-related proteins also alter the mitochondria–ER
axis or if this is altered in sporadic cases would be useful to address a possible general pathway
for PD. Mutations in LRRK2 are related to both familial and sporadic PD134. Autosomal-
dominant mutations in LRRK2 have been shown to cause deficits in intracellular Ca2+ handling,
mitochondrial depolarization and increased mitophagy, which can be prevented by L-type Ca2+
channel inhibitors.
REFERENCES:
 https://www.ncbi.nlm.nih.gov/pubmed/17288551
 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3290993/
 https://smw.ch/resource/jf/journal/file/view/article/smw/en/smw.2002.09861/smw.2002.0986
1.pdf/
 http://www.hal.inserm.fr/inserm-01296824/document