ptmscan hs ubiquitin sumo remnant motif  (Cell Signaling Technology Inc)

Journal: Advanced Science

Article Title: TOGR3, a Proteasome β4 Subunit, Orchestrates Sugar Homeostasis to Trade Off Growth and Thermotolerance in Rice

doi: 10.1002/advs.202516395

Figure Lengend Snippet: TOGR3 encodes the β4 subunit of 26S proteasome. a) Identification of TOGR3 . Top: Fine‐mapping of TOGR3 on rice chromosome 3. Bottom: multiple sequence alignment of the conserved proteasome β4 subunit across species, highlighting the Leu66Pro mutation in togr3 (red arrows). b) Genetic complementation of togr3 . Transformation with the native TOGR3 genomic fragment rescued plant height and tiller number ( n ≥ 15). Data: mean ± SEM (Duncan's Multiple Range Test, p < 0.05). Scale: 10 cm. COM, complementation lines. c) Thermoresponsive expression of TOGR3 . RT‐qPCR analysis of TOGR3 transcripts in seedlings after 2 h temperature treatments. d) Temperature‐dependent accumulation of TOGR3 protein. Immunoblot analysis of wildtype and togr3 seedlings grown at 25°C and shifted to 35°C for the indicated durations. cFBP served as loading control. e) Proteasome activity assays. Chymotrypsin‐like, trypsin‐like, and caspase‐like activities in wildtype and togr3 seedlings grown at either 25°C or 35°C for 2 weeks, measured using aminoluciferin‐conjugated substrates (Suc‐LLVY, Z‐LRR, and Z‐LRR, respectively). Activities were assayed at the corresponding growth temperatures. Data represent mean ± SD ( n = 3). f) Accumulation of ubiquitinated proteins under elevated temperature. Total protein extracts from seedlings grown at 25°C or 35°C were immunoblotted with Ubiquitin antibody. a‐ACTIN served as loading control. All experiments were repeated at least three times.

Article Snippet: Ubiquitin‐modified peptide were enriched by using PTMScan Pilot Ubiquitin Remnant Motif (K‐ε‐GG) Kit (Cell Signaling Technology, #14482) and performed according to Udeshi et al. (2013) [ ] with modifications.

Techniques: Sequencing, Mutagenesis, Transformation Assay, Expressing, Quantitative RT-PCR, Western Blot, Control, Activity Assay, Ubiquitin Proteomics

Journal: Advanced Science

Article Title: TOGR3, a Proteasome β4 Subunit, Orchestrates Sugar Homeostasis to Trade Off Growth and Thermotolerance in Rice

doi: 10.1002/advs.202516395

Figure Lengend Snippet: TOGR3 improves heat and drought tolerance in rice. a,b) Heat tolerance assays. Two‐week‐old seedlings were exposed to 45°C for 32 h (a) or 36 h (b) and recovered for 10 days before survival assessment ( n ≥ 24, 3–4 biological replicates). Ri, RNAi line; OE, overexpression line. c) Heat‐induced accumulation of ubiquitin conjugates after heat treatment for the indicated durations detected by immunoblotting. a‐ACTIN serves as a loading control. d) Growth responses under combined temperature and humidity conditions. Seedling height and fourth‐leaf dimensions are shown ( n ≥ 16). Fold changes between wildtype and togr3 are indicated. e,f) Drought tolerance assays. 2‐week‐old seedlings were subjected to drought for 10 (e) or 12 days (f), followed by 7 days of rewatering ( n ≥ 24 and 3 replicates). g) Drought‐induced ubiquitin accumulation. Drought treatments for the indicated durations were indicated. Scale bars: 10 cm in (a, b, d, e). Data represent as mean ± SEM and were analyzed using two‐sided Student's t ‐test (* p < 0.05, ** p < 0.01).

Article Snippet: Ubiquitin‐modified peptide were enriched by using PTMScan Pilot Ubiquitin Remnant Motif (K‐ε‐GG) Kit (Cell Signaling Technology, #14482) and performed according to Udeshi et al. (2013) [ ] with modifications.

Techniques: Over Expression, Ubiquitin Proteomics, Western Blot, Control

Journal: Advanced Science

Article Title: TOGR3, a Proteasome β4 Subunit, Orchestrates Sugar Homeostasis to Trade Off Growth and Thermotolerance in Rice

doi: 10.1002/advs.202516395

Figure Lengend Snippet: TOGR3 shapes a thermoresponsive ubiquitylome via ubiquitin‐proteasome system. a) Numbers of differentially ubiquitinated peptides and proteins in ZH11 and togr3 . Differential ubiquitination was defined as fold change ≥ 2 and Q value < 0.05. b) Comparison of upregulated ubiquitinated peptides and proteins in togr3 versus ZH11 between 25°C and 35°C. c) Heatmap showing the relative abundance of upregulated ubiquitinated peptides in togr3 . d) Shared Gene Ontology (GO) terms among upregulated ubiquitinated proteins at both temperature conditions. e) Overlapping GO biological processes of differentially ubiquitinated proteins between ambient high temperature and acute heat stress datasets in wildtype (WYJ data adapted from Li et al., 2015) . f) Relative abundance of ubiquitinated proteins involved in carbohydrate metabolism. Different Ub‐peptides (P) were labelled. g) Activities of sugar metabolism—related enzymes. Data: mean ± SEM; assessed by two‐sided Student's t ‐test (* p < 0.05, ** p < 0.01).

Article Snippet: Ubiquitin‐modified peptide were enriched by using PTMScan Pilot Ubiquitin Remnant Motif (K‐ε‐GG) Kit (Cell Signaling Technology, #14482) and performed according to Udeshi et al. (2013) [ ] with modifications.

Techniques: Ubiquitin Proteomics, Comparison

Journal: Advanced Science

Article Title: TOGR3, a Proteasome β4 Subunit, Orchestrates Sugar Homeostasis to Trade Off Growth and Thermotolerance in Rice

doi: 10.1002/advs.202516395

Figure Lengend Snippet: Proposed model illustrating the molecular mechanism by which TOGR3‐dependent ubiquitin‐proteasome system regulates rice thermotolerance through sugar homeostasis.

Article Snippet: Ubiquitin‐modified peptide were enriched by using PTMScan Pilot Ubiquitin Remnant Motif (K‐ε‐GG) Kit (Cell Signaling Technology, #14482) and performed according to Udeshi et al. (2013) [ ] with modifications.

Techniques: Ubiquitin Proteomics