Expression
ix) See TSR (GTEX등 included)
x) Gray matter vs White matter
a. {Suridjan, 2015 #2001}: NLRP3 자체는 못 찾겠고) TSPO signal은 WM 이 GM 의 ~70%임.
xi) NLRP3 is predominantly expressed in microglia/macrophages
xii) (https://doi.org/10.3389/fncel.2017.00063) NLRP3 inflammasome is is present in microglia and astrocytes in the CNS (Cho et al., 2014; Zendedel et al., 2016). It remains debated whether neurons express NLRP3 (Fann et al., 2013a; Yang et al., 2014; Kaushal et al., 2015).
xiii) {von Herrmann, 2018 #1772} expression of the NLRP1,13 AIM2,14 and NLRP315 NLRs has been detected in neurons
| Astrocyte | Neuron | |
|---|---|---|
| Human | {Gustin, 2015 #1814} 2nd, in human astrocytes IL-1β mRNA was not translated to protein, probably by an active repression mechanism [40]. | It remains debated whether neurons express NLRP3 (Fann et al., 2013a; Yang et al., 2014; Kaushal et al., 2015). |
| mouse |
{Gustin, 2015 #1814} the capacity to form a functional NLRP3 inflammasome and secretion of IL-1β is limited to the microglial compartment in the mouse brain. We were not able to observe IL-1β secretion from astrocytes, nor do they express all NLRP3 inflammasome components. Brain microglia, but not astrocytes, express NLRP3 inflammasome components and substrates |
GWAS
| (von Herrmann, 2018 #596) |
multiple SNPs) including rs7525979 that was associated with a significantly reduced risk of developing PD. Mechanistic studies conducted in HEK293 cells indicated that : rs7525979 → ↓ efficiency of NLRP3 translation impacting NLRP3 → ↓ protein, ↑ ubiquitination state, and ↑ solubility. → accumulation of a ubiquitinated, insoluble form of NLRP3 (protein state that is suggestive of protein inactivation) → ↓ PD risk (js: so this is LOF mutation) |
Table 1. Parkinson disease risk in the Parkinson’s Progression Markers Initiative (PPMI) study associated with the NLRP3 SNP rs7525979
| Cases n = 402, n (%) | Controls n = 182, n (%) | OR (95% CI) | P value | |
|---|---|---|---|---|
| Age, mean (sd) | 62.2 (9.7) | 61.6 (11.0) | 1.006 (0.99-1.02) | 0.53 |
| Sex | ||||
| Male | 260 (64.7) | 116 (63.7) | 1.0 (Reference) | |
| Female | 142 (35.3) | 66 (36.3) | 0.94 (0.65-1.36) | 0.73 |
| rs7525979 (C/C) | ||||
| C/C | 357 (88.8) | 148 (81.3) | 1.0 (Reference) | |
| C/T+T/T | 45 (11.2) | 34 (18.7) | 0.55 (0.34-0.89) | <0.02 |
Model adjusted for all variables in the table
Neuronal
KO
NLRP3 KO mice we have used: https://www.jax.org/strain/021302 (B6.129S6-Nlrp3tm1Bhk/J)
Caspase 1
Caspase 1 expression: GTEX: HC cbll=cortex, {Ko, 2020 #2484} AD mouse: fig5, no signal in cerebellum?
aSyn splicing
{Ma, 2018 #2046} Caspase-1 cleaves full-length α-Syn to generate α-Syn121.
α-Syn121 (ie 1-121) activated apoptosis pathway more efficiently than α-Syn FL, and 3.3. α-Syn121 showed strong cytotoxicity (fig 5) (but weak membrane disruption ability)
α-Syn121 were expressed and purified as we reviously reported [22]. C-terminal 19-residues peptide (C19) was synthesized with standard solid phase peptide synthesis (SPPS) and purified by HPLC. → α-Syn121 + C19 amorphous aggregates rather than typical β-sheet-rich fibril (TEM) → ↑ aggregation (ThT assay)
in vitro
To confirm cleavage site: {Wang, 2016 #2086} i) we used matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to determine the size of the major caspase-1-cleaved fragment (Fig. 5 C and D). The truncated fragment of aSyn (1-121) was found to have a molecular mass of 13,167 Da, which corresponds to the molecular mass of an aSyn fragment ending at residue aspartic acid 121 (theoretical MW, 13,166 Da). Cf: aSyn FL MW is 14.5 kDa. ii) To confirm that we identified the correct cleavage site, we introduced a point mutation substituting Asp121 for glutamic acid, which we expressed and purified as described previously (31, 32) (Fig. S3). Introducing the D121E mutation completely abolished truncation of aSyn by caspase-1 in our in vitro experiments (Fig. 5A, lane 121E),
{Wang, 2016 #2086} (normally, fig5) Caspase-1 directly cleaves aSyn at Asp121. casp1 activation → aSyn truncation (wb),
- the concentration of Casp1 to cleave α-Syn needs μM order of casp1 (fig 5a).
- VX765 inhibits aSyn truncation compared with control (wb), (fig3, in vitro)
- Fig. 6. Truncated aSyn is aggregation-prone (ThT aggregation assay).
- Fig7. VX765 and genetic knockdown with anti-caspase-1 shRNA (RNAi), → partially rescues menadione-induced toxicity in neuronal cells (by ~half)
{Wang, 2016 #2086} the presence of caspase-1 in Lewy bodies in PD patients (staining, fig4),
Js: What about the presence of 1-121 and 122-140 (in Lewy bodies) in PD patients 이거 증거문헌 아직 없네??
Catalytic cleft
the P1 position of natural caspase-1 substrates have 100% stringently conserved amino acid Asp (which we have identified as Asp121 in aSyn), the cleavage efficiency is determined by the surrounding residues (P10-P10’). divergent from the sequence YVHD found to be cleaved in caspase-1’s natural substrate pro-IL-1b.
Uncertain Spans
| location | transcription | uncertainty |
|---|---|---|
| KO row | B6.129S6-Nlrp3<sup>tm1Bhk</sup>/J | Superscript allele tag is partly stylized; reading is consistent with JAX strain 021302 page text. |
| Catalytic cleft | (P10-P10') | Subscript/prime distinction in source text is hard to verify; the prime is rendered as '. |
| Caspase 1 line | {Ko, 2020 #2484} AD mouse: fig5 | Reference number #2484 may be off by one digit; numerals are small. |