mcitrine usp19  (Addgene inc)

Journal: bioRxiv

Article Title: Common molecular mechanisms underlie the transfer of alpha-synuclein, Tau and huntingtin and modulate spontaneous activity in neuronal cells

doi: 10.1101/2021.07.18.452825

Figure Lengend Snippet: (A) Immunoblots showing protein levels in cell lysates and released to the cell media of cells expressing the different proteins. HEK cell stably expressing 25QHtt-EGFP, 103QHtt-EGFP, aSyn-EGFP and EGFP-Tau were transfected with USP19 or with the catalytic inactive form USP19 C506S. Quantifications were normalized to total protein levels using MemCode. (B) LDH measurements confirm the absence of cell toxicity and cell death in the experiments. (C) Tau is more strongly internalized by naïve cells. Percentage of EGFP positive cells after incubation with media from cells co-expressing 25QHtt-EGFP, 103QHtt-EGFP, aSyn-EGFP or EGFP-Tau together with USP19 or USP19 C506S for 24 hours. Cell counting was performed using flow cytometry. Data from at least three independent experiments for each condition. Significant differences were assessed by one-way ANOVA followed by multiple comparisons with significance between groups corrected by Bonferroni procedure. Differences were considered to be significant for values of p<0.05 and are expressed as mean ± SD, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.

Article Snippet: The following plasmids used for the transfection protocol: pcDNA 3.1, pcDNA 3.1-aSyn, pcDNA 3.1-Tau (4R2N), pcDNA 3.1-22QHtt exon 1 (1-90, CAG, ID CHDI-90000027, Coriell Institute), pcDNA 3.1-72QHtt exon 1 (1-90, CAG, ID CHDI-90001882-1, Coriell Institute), mCitrine-USP19 (Plasmid #78593, Addgene), mCitrine-USP19 C506S (Plasmid #78594, Addgene).

Techniques: Western Blot, Expressing, Stable Transfection, Transfection, Incubation, Cell Counting, Flow Cytometry