basic limonoid modulates chaperone mediated proteostasis 2433  (Proteostasis Therapeutics)

Journal: iScience

Article Title: Dopaminergic neurons are vulnerable to dysregulation of YEATS2-dependent calcium homeostasis

doi: 10.1016/j.isci.2026.115855

Figure Lengend Snippet: Pharmacological attenuation of SOCE restores DAergic synaptic integrity in dYEATS2 -deficient flies (A) Experimental scheme. Flies expressing membrane-tethered mCD8-GFP and dYEATS2 RNAi specifically in dopaminergic neurons ( Ddc>mCD8-GFP>dYEATS2-IR ) were reared on standard medium supplemented with vehicle or the Orai inhibitor BTP2 (YM-58483) at 1 μM or 10 μM. Adult flies were transferred to fresh vials containing the same treatments, and heads were dissected at 5 days post-eclosion for confocal imaging or RNA extraction. (B) Representative confocal images (posterior→anterior orientation) showing functionally active DAergic neurons identified by co-localization of mCD8-GFP (membrane marker expressed under Ddc-GAL4 driver) and tyrosine hydroxylase (TH) immunoreactivity. Central brain boundaries are indicated by dotted lines; major DA clusters (PAL, PAM, PPL1, PPM3) are highlighted with red dashed circles. Scale bars, 250μm. (C) Quantification of EGFP-TH co-localization (number of co-localized puncta) was performed on 10 independent brains per condition using the colocalization module in CellSense (Olympus). Bars show mean ± SEM; BTP2 treatment at both 1 μM and 10 μM significantly increased the number of EGFP-TH co-localizing spots relative to untreated dYEATS2 -IR animals. (D) Transcript levels of selected dYEATS2 -responsive genes ( Gαq , trpL , vMAT , and DD2R ) were measured from dissected adult heads following vehicle or BTP2 treatment to assess whether SOCE inhibition modulates these transcriptional changes. Gene expression was determined by reverse transcription quantitative PCR (RT-qPCR). Data are presented as mean ± SEM. Statistical significance was assessed by one-way ANOVA with Šidák’s and Tukey’s post hoc tests, respectively; p < 0.05 was considered significant.

Article Snippet: Scale bars, 250μm. (C) Quantification of EGFP-TH co-localization (number of co-localized puncta) was performed on 10 independent brains per condition using the colocalization module in CellSense (Olympus).

Techniques: Expressing, Membrane, Imaging, RNA Extraction, Marker, Inhibition, Gene Expression, Reverse Transcription, Real-time Polymerase Chain Reaction, Quantitative RT-PCR

Journal: STAR Protocols

Article Title: Protocol for Seahorse 3D Mito Stress assay in patient-derived atypical teratoid rhabdoid tumor CHLA-05-ATRT single neurospheres

doi: 10.1016/j.xpro.2026.104515

Figure Lengend Snippet: Oxygen consumption rate (OCR) in the presence or absence of spheroids (A) Incucyte image of spheroid in the center of the microplate well at the end of the assay and (B) the corresponding OCR kinetic graph showing a response after drug injections as expected. (C) Image of a microplate well with no spheroid detected at the end of the assay and (D) corresponding OCR kinetic graph showing no response after drug injections.

Article Snippet: Incucyte Organoid Analysis Software Module , Sartorius , 9600–0034.

Techniques:

Journal: STAR Protocols

Article Title: Protocol for Seahorse 3D Mito Stress assay in patient-derived atypical teratoid rhabdoid tumor CHLA-05-ATRT single neurospheres

doi: 10.1016/j.xpro.2026.104515

Figure Lengend Snippet: Effect of coating timing on spheroid retention in microplate wells (A) Incucyte image of a poly-lysine coated microplate with spheroids transferred on the same day as the assay, taken before the assay. (B) Image of the same plate taken after the assay, showing that the spheroids were displaced and/or destroyed during the assay. (C) Image of a poly-lysine coated microplate with spheroids transferred the day before the assay, taken before the assay and (D) after the assay, showing that the spheroids were intact.

Article Snippet: Incucyte Organoid Analysis Software Module , Sartorius , 9600–0034.

Techniques: