adeno-associated viral vectors expressing excitatory receptors and a fluorescent marker of expression aav8-hsyn-dio-hm3d(gq)-mcherry, plasmid #44361  (Addgene inc)

Journal: Life Science Alliance

Article Title: CTPS2 regulates CTP synthetase activity by interacting with CTPS1

doi: 10.26508/lsa.202403117

Figure Lengend Snippet: (A) CTPS1+2-KO HEK cells were transiently co-transfected with plasmids coding GFP-CTPS2 (in green) and pLVX vectors for CTPS1, CTPS2, or CTPS1 H355A (expressing mCherry as a reporter of transfection in red) in the absence of any treatment. Data from one representative experiment of two independent experiments. Bar graphs on the right panel correspond to the filament–cytoophidium/cell quantification from the images on the left with indicated numbers of cells and filaments–cytoophidia counted (n=) in three independent fields/images for each condition independently of the mCherry staining. Magnification 40x, scale bar: 25 μm. (B, C) CTPS1+2-KO cells stably expressing GFP-CTPS1, GFP-CTPS2, GFP-CTPS1 H355A , or GFP-CTPS2 H355A . Cells were then maintained in culture without cytidine. (B) Histograms of GFP expression of different cell lines. Data in the right and left panels are from the same experiment with the same “nontransfected” control histogram, which is representative of two independent experiments. (C) Confluency curves as percentages (%) showing the proliferation of different cell lines. Confluency was measured using an IncuCyte Zoom system. Cells were seeded for 24 h, then treated or not (untreated) with the indicated concentrations of 3-deazauridine (3-DU) or 200 μM cytidine. (A) Data as means ± SD. Mann–Whitney statistical tests, * P < 0.05 and ** P < 0.01. n = is indicated in the panel.

Article Snippet: The cDNAs were verified by sequencing and inserted into a bicistronic lentiviral expression vector encoding mCherry as a reporter (pLVX-EF1α-IRES-mCherry Vector; Clontech).

Techniques: Transfection, Expressing, Staining, Stable Transfection, Control, MANN-WHITNEY

Journal: Life Science Alliance

Article Title: CTPS2 regulates CTP synthetase activity by interacting with CTPS1

doi: 10.26508/lsa.202403117

Figure Lengend Snippet: (A) FACS dot-plots of GFP and mCherry expression. (A, B) Bar graphs from flow-cytometry data as presented in (A), showing the percentage (%) of GFP+ alone, mCherry+ alone, and double-positive GFP+mCherry+cells.

Article Snippet: The cDNAs were verified by sequencing and inserted into a bicistronic lentiviral expression vector encoding mCherry as a reporter (pLVX-EF1α-IRES-mCherry Vector; Clontech).

Techniques: Expressing, Flow Cytometry

Journal: Life Science Alliance

Article Title: CTPS2 regulates CTP synthetase activity by interacting with CTPS1

doi: 10.26508/lsa.202403117

Figure Lengend Snippet: Histograms of mCherry reporter expression profiles of CTPS1-KO Jurkat cells transduced with CTPS1 WT or CTPS1 H355A after or not cytidine deprivation (no cytidine) during 3, 6, and 24 d. The CTPS1 H355A profiles are similar to those of CTPS1 WT.

Article Snippet: The cDNAs were verified by sequencing and inserted into a bicistronic lentiviral expression vector encoding mCherry as a reporter (pLVX-EF1α-IRES-mCherry Vector; Clontech).

Techniques: Expressing, Transduction