Embedded Files
Reyes AV, Shrestha R, Grimser TS, Byun D, Xu SL. (2024). Impact of alternative splicing on Arabodpsis proteome. bioRxiv. DOI: 10.1101/2024.02.29.582853.
Impact of Alternative Splicing on the Arabidopsis Proteome
One of the first questions explored in high school biology classes is how humans and c. elegans can have a similar number of genes, yet humans are the more complex organism. One answer is that alternative splicing contributes to protein diversity. But how much protein diversity is caused by AS is still debated.
Objectives
To explore the global impact of alternative splicing (AS) on the Arabidopsis proteome, with a specific emphasis on Intron Retention (IR), the predominant AS event in plants.
Results
By employing Tandem-mass tag (TMT) proteomics in conjunction with extensive fractionation, I have achieved deep proteome coverage, quantifying 13,850 protein groups between wild-type and acinus-pnn mutant. Using bioinformatics we dissect the regulation at protein level and reveal various expression patterns as a result of alternative splicing, in addition to discovering a potential truncated protein in the acinus-pnn mutants.
Project was initiated and designed by me. Currently being led by Dr. Wenxuan Zhai in Xu Lab
Nanobody Engineered O-Sugar Modification in A. Thaliana
O-linked sugar modification in humans has effects on hunger, mice with a knocked down O-GlcNAc Transferase (OGT) become fat from overeating. What kind of implications can this have in plants? Currently, the main methods of studying this PTM in plants is by global knockdown or over expression. Having a more targeted method could help unravel the functions of this PTM. Potential discoveries could potentially be utilized for future genetic engineering.
Objectives
To engineer plant O-Linked sugar transferases, SPY, and SEC, to be selective using nanobody driven approach. With this tool we could dissect the function this modification has any of its target proteins.
Results
Preliminary results using AP-MS indicate that our system is selectively boosting O-Fucose PTM on a selected target protein. Further validation and functional studies are underway.
Reyes AV, Shrestha R, Baker PR, Chalkley RJ, Xu SL. (2022). Application of Parallel Reaction Monitoring in 15N labeled Samples for Quantification. Front. Plant Sci., DOI: 10.3389/fpls.2022.832585.
Parallel Reaction Monitoring of Labeled Samples
During my time at the Carnegie Mass Spectrometry Facility, I dedicated substantial efforts to the advancement of proteomic methodologies within the plant biology field. My primary focus was the development and refinement of cutting-edge tools, particularly the utilization of 14 N and 15 N metabolic labeling for relative protein quantification. My efforts were to ultimately make these proteomic tools more easily assessable to plant scientists.
Objectives
To provide a detailed method on how to preform a targeted analysis of 14 N and 15 N metabolic labeled protein samples to address ambiguous quantifications from DDA acquisition.
Results
My published works provide a targeted approach using the Protein Prospector and the Skyline platform. Using the methods, you can deconvolute low confidence data through the use of 14 N/ 15 N labeling with Parallel Reaction Monitoring.
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