Journal: bioRxiv

Article Title: Phase separation of S-RNase promotes self-incompatibility in Petunia hybrida

doi: 10.1101/2023.09.07.556770

Figure Lengend Snippet: ( A - B ) Detections of GFP-S 3L -RNase degradation after self ( S 3L pollen) (A) and non-self-pollen ( S V pollen) (B) tube extract treatments with or without MG132 in vitro. α-GFP, GFP antibody; CBB, coomassie brilliant blue. The curves show the time course of the remaining of GFP-S 3L -RNase. Data are presented as mean ± SD. ( n = 3). ( C )Pre-formed SRCs can still be degraded by non-self-pollen tube extracts. The curves show the time course of the remaining of GFP-S 3L -RNase. Data are presented as mean± SD. ( n = 3). α-GFP, GFP antibody; CBB, coomassie brilliant blue. ( D ) Fluorescence microscopy images of N. benthamiana leaf cells that express S 3 -RNase-GFP or S 3L -RNase-GFP with cLUC-PhSSK1. Scale bars, 20 μm. ( E - J ) Detections of S 3 -RNase-GFP or S 3L -RNase-GFP using N. benthamiana leaf cells that co-express cLUC-PhSSK1 (E and F), PhS 3L -SLF1-mCherry (G and H), or cLUC-PhSSK1 and PhS 3L -SLF1-mCherry (I and J) with or without MG132. α-GFP, GFP antibody; α-mCherry, mCherry antibody; α-cLUC, cLUC antibody; α-Actin, Actin antibody.

Article Snippet: Analysis of S3L-RNase-GFP, S3-RNase-GFP, cLUC-PhSSK1, and PhS3L-SLF1-mCherry proteins in N. benthamiana leaf cells, leaves 2 d after agrobacteria infiltration were treated with 40 µM MG132 (MBL, catalogue no. D153-11) for 1 d, and then analyzed using a Zeiss 980 microscope.

Techniques: In Vitro, Fluorescence, Microscopy

Journal: Scientific Reports

Article Title: Stability of tuberous sclerosis complex 2 is controlled by methylation at R1457 and R1459

doi: 10.1038/s41598-020-78274-6

Figure Lengend Snippet: Arginine methylation of TSC2 blockes Akt-dependent TSC2 phosphorylation at T1462. ( A ) Phosphorylation assays were conducted using the unmodified TSC2 peptide (cont) and the modified arginine methylated peptide (Methyl). Peptides were incubated with recombinant Akt and [γ- 32 P] ATP and phosphorylation was detected using a scintillation counter. Error bars indicate standard deviation (SD) from three independent experiments; ** p < 0.01. ( B/C ) Effect of TSC2 methylation on TSC2 phosphorylation measured by inhibiting ( B ) [ 3 H]-SAM using cycloleucine (100 mM) with MG132 (10 µM) or ( C ) PRMT1 activity using AMI-5 (2.0 µM). Phosphorylated TSC2 (T1462) and AKT (S473) levels were analysed by western blotting. Error bars indicate the SD from two independent experiments; * p < 0.05. All experiments were repeated three times.

Article Snippet: The proteasome inhibitor MG132 was obtained from Funakoshi (Cat. #133407-82-6).

Techniques: Methylation, Phospho-proteomics, Modification, Incubation, Recombinant, Standard Deviation, Activity Assay, Western Blot