Genes responsible for autosomal dominant forms of PD.
Of utmost importance, a molecular correlation between PD and the UPR came from studies implying several autosomal-dominant (AD) and -recessive (AR) PD causative genes. Most studies regarding the implication of AD–PD causative proteins in UPR regulation are linked to α-synuclein.
α-Synuclein is a protein encoded by the
SNCA gene that accumulates in Lewy bodies and Lewy neurites. Several point mutations, duplications, and triplication of the gene have been identified, and multiple
in vitro and
in vivo studies indicate that its accumulation triggers its aggregation and thereby induces neurotoxicity [
115,
116]. The accumulation of aggregated α-syn in the brain and notably its soluble oligomeric toxic form is strongly linked to the etiology of PD [
117,
118].
The overexpression of α-syn, and thus its aggregated toxic forms, correlates with the chronic activation of multiple branches of the UPR and ER stress-mediated apoptosis. Thus, it has been shown that the overexpression of α-syn triggers the activation of UPR in yeast [
119], and that its phosphorylation at serine 129, which is associated to its aggregation and toxicity, leads to an important ATF6 regulation in dopaminergic
in vitro models [
120]. Wild-type and mutated α-syn overexpression in SH-SY5Y cells triggers an alteration in calcium metabolism and an activation of IRE1α-XBP1-signaling pathway [
121]. The treatment of differentiated SH-SY5Y cells with oligomeric but not monomeric α-syn leads to enhanced XBP1 splicing, indicating a specific activation of the IRE1-XBP1 signaling pathway by α-syn oligomers [
122]. Differentiated 3D5 human neuroblastoma-derived cells overexpressing α-syn show increased levels of GRP78/BiP and phospho-EIF2α [
123] in basal conditions, and tunicamycin-induced ER stress leads to accumulation of oligomeric α-syn [
123], indicating that ER stress may feed α-syn aggregation and toxicity. α-Syn crowding within the ER induces the activation of the PERK-dependent pathway of the UPR
in vitro and
in vivo, an activation process mediated by α-synuclein direct interaction with BiP UPR [
124]. α-Synuclein affects ATF6 processing directly via protein–protein interactions or indirectly by means of the reduced incorporation to COPII vesicles. Altered ATF6 processing leads to an impairment of ERAD and increased apoptotic response [
125].
This network of evidence does not make it possible to ascertain whether α-syn neurotoxicity is the cause or the consequence of UPR failure and thus which of them is the primary trigger of PD pathogenesis. Nevertheless, a recent work from Colla et al. in A53T α-syn transgenic mice indicates that the accumulation of α-syn toxic species in the ER is responsible for UPR activation [
129] and that the detection of ER-associated α-syn oligomers precedes ER stress response [
130], thus suggesting that UPR activation is rather the consequence of accumulation of α-syn in PD. The aggregation of α-syn in the ER has been corroborated by approaches implying (fluorescence resonance energy transfer) FRET biosensors [
131].
LRRK2 (leucine-rich repeat kinase 2 gene) is a kinase of the ROCO family [
132], the mutations on which are associated to autosomal dominant forms of PD and more than 3% of sporadic PD forms [
133,
134,
135]. The biological functions of LRRK2 remain poorly understood, but a few studies suggest that it is linked to UPR. Thus, studies on LRRK2 subcellular distribution in control versus idiopathic PD revealed that LRRK2 is mainly detected in the ER of neurons and that it co-localizes with two ER-specific markers [
136]. The analysis of the contribution of a short hairpin RNA (shRNA)-mediated LRRK2 depletion in SH-SY5Y cells leads to a downregulation of BiP in 6OHDA ER stress conditions, indicating that LRRK2 depletion promotes cytoprotection by modulating the UPR [
137].
Moreover, recently it has been demonstrated that LRRK2 may affect mitochondrial bioenergetics by modulating ER–mitochondria tethering via the PERK-mediated ubiquitination pathway [
138] and that mutated LRRK2-increased ER stress and apoptosis by disabling the sarco/endoplasmic reticulum Ca
2+-ATPase (SERCA) in astrocytes [
139]. LRRK2-mediated SERCA dysfunction leads to Ca
2+ overload in the mitochondria.
3.3.1. Genes Responsible for Autosomal Recessive forms of PD (AR PD)
Most studies linking AR PD genes to ER function and UPR are centered around parkin (PRKN), PINK1 (PTEN (Phosphatase and tensin homolog) -induced putative kinase 1) and DJ-1.
Parkin is an E3-ligase [
140,
141] and transcription factor [
142] involved in multiple cellular processes that are affected in PD. Parkin protein is encoded by the PRKN gene, the mutations of which are responsible for most of autosomal recessive juvenile PD [
140]. One of the first pieces of evidence linking parkin to ER stress came from
in vitro studies showing that parkin induced neuroprotection against ER stress [
141] and that the Pael (parkin-associated endothelin receptor-like) receptor that is involved in ER stress-mediated apoptosis is a parkin substrate [
143]. These studies also demonstrate that the potent ER stress inducer, tunicamycin, leads to an upregulation of parkin mRNA and protein levels that correlates to increased neuroprotection [
141]. Moreover, parkin overexpression was found to be protective against Pael ER stress-mediated apoptosis [
143].
Interestingly, it has been shown that parkin expression may be induced by either ER or mitochondrial stress via its transcriptional regulation by ATF4. An upregulation of parkin levels protects against mitochondrial failure and cell death, suggesting a functional link between parkin, ER stress, and mitochondrial homeostasis [
144]. Moreover, it was shown that salubrinal, an ER stress inhibitor, prevents rotenone-induced apoptosis in SH-SY5Y, corroborating the neuroprotective role of the ATF4–parkin pathway in ER stress triggered by PD inducers [
145]. Corroborating the link between ER and mitochondria via parkin, researchers showed that the increase of parkin levels facilitated the crosstalk between these organelles and granted the calcium mitochondrial load to assure cell bioenergetics [
146].
We have shown that endogenous and overexpressed parkin are induced by ER stress and that parkin impacts the UPR response via a p53-dependent transcriptional control of XBP1 [
147]. These data provide a direct evidence of a role of parkin in neuronal control of the UPR. Of note, parkin-mediated control of ER stress is not restricted to neurons since astrocytes depleted in parkin show increased levels of spliced XBP1, ATF6, ATF4, CHOP, and Ccl2 in response to thapsigargin [
148]. Interestingly, it has been shown that the induction of parkin levels may vary according to the cell type since an increased expression of parkin was observed in astrocytes and not primary hippocampal neurons submitted to ER stress [
149]. The contrasting data between SH-SY5Y cells and hippocampal neurons may suggest a preponderant function of parkin in dopaminergic neurons. Moreover, in corroborating a cell type-specific induction of parkin by ER stress, it was shown that 2-mercaptoethanol and tunicamycin increased the expression of parkin in SH-SY5Y (H) cells, Neuro2a cells, Goto-P3 cells, but not in SH-SY5Y (J) cells and IMR32 cells [
150].
Several
in vivo models corroborate the impact of parkin to UPR control. Thus, parkin mutant flies show an activation of the PERK branch of the UPR through the establishment of mitofusin bridges between defective mitochondria and the ER [
151]. Moreover, drosophila models of parkin overexpression show an enhancement of K48-linked polyubiquitin and reduced levels of protein aggregation during aging [
152].
A few studies have implicated PINK1 in the UPR response. PINK1 is a mitochondrial serine/threonine kinase that, in conjunction with parkin, is strongly implicated in the control of mitophagy [
153,
154]. Mutations of PINK1 are associated to both genetic and sporadic PD cases [
155,
156] and perturbed mitochondrial homeostasis. Further, the overexpression of the deletion mutant of OTC (ornithine transcarbamylase) (ΔOTC), which induces mitochondrial UPR in mammalian cells [
157], leads to an increase of PINK1 protein levels, parkin recruitment, and mitophagy firing without dissipation of mitochondrial potential in HeLa cells [
158]. These data indicate that mitochondrial UPR leads to the induction of PINK1–parkin-dependent mitophagy followed by reduced misfolded protein load. Interestingly, PINK1 modulation was also shown to regulate mitochondrial UPR. Thus, mutations in both human and fly PINK1 result in higher levels of misfolded components of respiratory complexes and accumulation of HSP60 [
159].
PINK1 was shown to prevent ER-induced apoptosis in mice primary cortical neurons [
160], and transcriptomic studies performed in PINK1 knockout aged mice indicated a downregulation of ER stress response genes [
161]. Finally,
in vivo studies in
Drosophila show that PINK1 mutations are associated with PERK modulation [
151].
DJ-1 (PARK7) is a multifunctional protein [
162] considered as a mitochondrial oxidative stress cellular sensor that interestingly harbors chaperone properties [
163]. In addition to its key mitochondrial function, downregulation of DJ-1 was shown to affect ER mitochondria contacts in SH-SY5Y differentiated cells [
164]. Corroborating these data, DJ-1 overexpression was shown to overcome the p53-induced mitochondrial calcium uptake failure and the perturbations in ER–mitochondria tethering [
165]. Overexpressed and endogenous DJ-1 proteins protect against ER stress induced by thapsigargin and tunicamycin in Neuro 2a cells [
166].
DJ-1 regulates and is regulated by UPR signaling pathway members. Thus, DJ-1 regulates the UPR and apoptotic response through the increase of ATF4 signaling in stress conditions [
167] and is transcriptionally regulated by XBP1. Thus, we have shown that XBP-1 directly binds to its promoter, leading to its upregulation [
147]. Finally, it has been shown that oxidized DJ-1 binds to R-HSP5 and favors the elimination of misfolded cargo proteins by autophagy in oxidative stress conditions [
168].
Among genetic PD, PARK20 is a rare autosomal recessive juvenile Parkinson’s form due to mutations in the phosphatidylinositol phosphatase, synaptojanin1 (Synj1) [
169,
170]. PARK20 fibroblasts show alterations in the exit machinery of the ER and Golgi trafficking. These alterations lead to the activation of the PERK branch of UPR due to the accumulation of cargo proteins in the ER [
171].
Finally, mutations in PLA2G6 (calcium-independent phospholipase A2), which are linked to PARK14-linked young-onset dystonia-parkinsonism syndrome with recessive inheritance [
172] were shown to upregulate GRP78, IRE1, PERK, and CHOP protein levels
in vivo [
173].
3.3.2. PD Risk Factors
Glucocerebrosidase (GCase, GBA) is a lysosomal enzyme encoded by the GBA gene that is considered an important risk factor to PD [
174]. Mutations in GCase are associated to α-syn accumulation due to an impairment of its CMA (chaperone-mediated autophagy)-mediated degradation [
175].
Post-mortem analysis of brains of Lewy bodies dementia (LDB) patients carrying GBA1 mutations show alterations on protein levels BiP and HERP, indicating abnormal UPR response [
176]. Horowitz’s team has shown that mutations of GCase lead to their retention in the ER and subsequent activation of the UPR in the
Drosophila model [
177]. Moreover, they showed that the activation of UPR, illustrated by increased mRNA levels of XBP1s and Hsp-70, may be reversed by ambraxol, a GCase chaperone [
178].
Even if it is still debated, high-temperature requirement A2 (HTRA2/Omi/PARK13) is often considered as a PD risk factor [
179,
180]. HTRA2 is a serine protease with strong homology to the
Escherichia coli HTRA2, that are important to bacterial survival at high temperatures. Considering that bacterial HTRA2 is involved in the elimination of misfolded aggregated proteins, it is not surprising that HTRA2 is functionally linked to the UPR. Thus, it has been shown that HTRA2 depletion/invalidation in SH-SY5Y and immortalized mouse embryonic fibroblasts (MEFs) triggers a decrease of the pro-apoptotic CHOP protein in 6OHDA stress conditions [
181,
182]. Interestingly, it has been shown that HTRA2 is induced by tunicamycin
in vitro, indicating that Omi is activated by ER stress [
183].