Journal: bioRxiv
Article Title: The elongation of Mest transcript into MestXL sustains, but does not initiate, the maternal allele bias of its convergent gene Copg2 during neurogenesis
doi: 10.64898/2026.03.13.711300
Figure Lengend Snippet: A) Heatmap illustrating the expression levels of specific markers, as measured by RT-qPCR, during the differentiation of brain organoids. Pluripotency markers ( Nanog , Pou5f1 ), neural progenitor markers ( Nestin , Pax6 ), neuronal markers ( Elavl3 , Tbr1 , Tubb3 ), and the astrocyte marker Gfap were analyzed across B/J embryonic stem cells (ESCs) and at days 7, 14, and 21 of brain organoid differentiation. Data are presented for each independent experiment, with sample sizes of n=4 for ESCs and n=3 for days 7, 14, and 21. The color scale represents Z-score expression levels. B) Genome browser view of the Mest and Copg2 loci, showing allelic RNA-seq signals (upper panel) and H3K36me3 enrichment obtained by Cut&Run (lower panel) in merged B/J and J/B newborn brain tissue (NBB)( n = 2 for RNA-seq; n = 4 for C&R). For each condition, the quantitative and merged parental allelic signals are at the top and bottom, respectively. Maternal and paternal expression levels are shown in red and blue, respectively. C) Genome browser view at the Klhdc10 and Kfl14 genes, respectively, to show the allelic-oriented RNA-seq signals in B/J ESCs (n=4) and at day 7 ( n = 3), day 14 ( n = 3), and day 21 ( n = 3) of in vitro brain organoid differentiation, as well as in merged B/J and J/B newborn brain tissue (NBB) ( n = 2). For each condition, the quantitative and merged parental allelic RNA-seq signals are at the top and bottom, respectively. Maternal and paternal expression levels are shown in red and blue, respectively.
Article Snippet: Cut&Run (C&R) was performed using the CUTANA CUT&RUN Kit (Epicypher) and non-fixed nuclei from ESCs and NPCs (for each cell type: n = 1 in the B/J and n = 1 in the J/B background, respectively), according to the manufacturer’s instructions.
Techniques: Expressing, Quantitative RT-PCR, Marker, RNA Sequencing, In Vitro
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![Soluble LacY permease can be reconstituted into liposomes that catalyze lactose active transport monitored by [ 14 C]lactose uptake in response to a membrane potential induced by valinomycin (interior negative) The soluble LacY was purified and reconstituted into proteoliposomes as described under and legend to figure (A). A schematic representation of the LacY activity assay is shown. Dilution of liposomes containing K + into Na + in the presence of valinomycin generates a positive outward membrane potential (ΔΨ) (B). To measure membrane potential-driven uphill transport, 10 μL of proteoliposomes were diluted into NaP i (pH 7.5) containing 0.1 mM [ 14 C] lactose and 10 μm valinomycin. At the times indicated, the reactions were quenched and counted as described in . Uphill transport of lactose is driven by DY (interior negative) when [K + ]in > [K + ]out at the time of dilution. Bottom set of time points represents dilution into 50 mM KP i containing 10 μm nigericin (negative control), which was used to dissipate the membrane potential and perform measurements with de-energized proteoliposomes. The radioactivity taken up by the proteoliposomes was converted to lactose concentration and normalized to the amount of LacY in the sample. A background value was determined by assaying protein-free liposomes lacking LacY and was subtracted from all measurements. All incubations were at 30°C. Average results representative of three experiments are shown. Data are represented as mean ± SEM.](https://pub-med-central-images-cdn.bioz.com/pub_med_central_ids_ending_with_5257/pmc13015257/pmc13015257__gr6.jpg)