Month Flat Week Day
November 2014
January 2015

Mon. 1 Dec, 2014

Cutting Edge Seminar - Jay J. Thelen: Phosphoproteomic analysis of seed maturation – from discovering phosphorylation sites to identifying kinase clients

Mon. 1 Dec, 2014 10:00 - 11:00

UPSC Seminar Series 2014
Cutting Edge Seminar 

Jay J. Thelen
Department of Biochemistry, 
Christopher S. Bond Life Sciences Center
University of Missouri, Columbia, MO, USA

Phosphoproteomic analysis of seed maturation – from discovering phosphorylation sites to identifying kinase clients

Host: Vaughan Hurry

Place: Lilla hörsalen, KB3A9


Although metabolic networks for storage reserve synthesis have been largely characterized in diverse plant seed we are only beginning to understand the complex regulatory processes involved in seed development. Protein phosphorylation is a major form of post-translational regulation in eukaryotes as evidenced by over 1000 protein kinases in the Arabidopsis proteome. To begin studying protein phosphorylation in developing seed we performed large-scale, mass spectrometry-based phosphoproteomic studies on seeds at five stages of development in soybean (Glycine max), rapeseed (Brassica napus), and Arabidopsis (Arabidopsis thaliana). Phosphopeptides were enriched from 0.5 mg total peptides using a combination of immobilized metal affinity and metal oxide affinity chromatography. Enriched phosphopeptides were analyzed by Orbitrap tandem mass spectrometry and spectra mined against cognate genome or cDNA databases in both forward and randomized orientations, the latter to calculate false discovery rate (FDR). We identified a total of 2001 phosphopeptides containing 1026 unambiguous phosphorylation sites from 956 proteins with an average FDR of 0.78% for the entire study (Meyer et al., 2012). The dataset was uploaded into the Plant Protein Phosphorylation Database (P3DB,, including annotated spectra, for public accession. P3DB is a portal for all plant phosphorylation data and allows for homology-based querying of experimentally-determined phosphosites (Gao et al., 2009). Comparisons to other large-scale studies revealed that 652 of the phosphoproteins are novel to this study. The unique proteins fall into several Gene Ontology categories, some of which are overrepresented in our study as well as other large scale phosphoproteomic studies including metabolic process and RNA binding; while other categories are only overrepresented in our study like embryonic development. Leveraging large-scale phosphoproteomic datasets such as these, we developed a phosphorylation prediction tool called MUSite ( that incorporates protein disorder as one of three features for prediction (Gao et al., 2010). The sensitivity and accuracy of this prediction algorithm is unmatched, and application to whole plant proteomes such as Arabidopsis TAIR10 indicates greater than 17,000 phosphorylation sites at the 99% confidence interval. Clearly, experimental and bioinformatic prediction of phosphorylation sites is rapidly becoming a facile task. However, confirmation and identification of cognate protein kinases responsible for these events remains challenging. I will also introduce a novel approach to address this problem called the Kinase Client or KiC Assay (Huang et al., 2009). Using this approach we have identified many kinase client relationships, including three different protein kinases responsible for over ten phosphorylation events on a single phosphoprotein.

Ahsan N, Huang Y, Tovar-Mendez A, Swatek KN, Zhang J, Miernyk JA, Xu D, Thelen JJ. (2013) A

Versatile Mass Spectrometry-Based Method to Both Identify Kinase Client-Relationships and Characterize Signaling Network Topology. J Proteome Res. 12:937-48

Meyer LJ, Gao J, Xu D, Thelen JJ (2012) Phosphoproteomic analysis of seed maturation in

Arabidopsis, rapeseed, and soybean. Plant Physiol. 159:517-28

Huang Y, Houston NL, Tovar-Mendez A, Stevenson SE, Miernyk JA, Randall DD, Thelen JJ (2010) A

quantitative mass spectrometry-based approach for identifying protein kinase –clients and quantifying kinase activity. Anal. Biochem. 402:69-76

Gao J, Agrawal GK, Thelen JJ, Xu D (2009) P3DB: A plant protein phosphorylation database. Nucl. Acids

Res. 37:D960-962

Gao J, Thelen JJ, Dunker AK, Xu D (2010) Musite: a tool for global prediction of general and kinase-

specific phosphorylation sites. Mol. Cell. Prot. 9:2586-25600

Wed. 3 Dec, 2014

Thesis Defence - Dmitry Kremnev: Get in tune: Chloroplast and nucleus harmony

Wed. 3 Dec, 2014 13:00 - 14:00
UPSC, Department of Plant Physiology
Thesis Defence

Dmitry Kremnev

Get in tune: Chloroplast and nucleus harmony

Faculty Examiner: Professor Francis Andre Wollman, Institut de Biologie Physico-Chimique (IBPC), Paris, Frankrike.
Supervisor: Åsa Strand.

Room: KB3B1 Stora hörsalen

Mon. 8 Dec, 2014

Seminar - Adeline Rigal: Unraveling transcriptional regulation of adventitious root formation with a small molecule

Mon. 8 Dec, 2014 10:00 - 11:00

UPSC Seminar - postdoc seminar

Adeline Rigal

Unraveling transcriptional regulation of adventitious root formation with a small molecule

Host: Stéphanie Robert

Room: KB3A9, Lilla hörsalen

Mon. 15 Dec, 2014

PhD half-time - Zsofia Reka Stangl

Mon. 15 Dec, 2014 10:00 - 11:00
UPSC PhD Halftime Seminar

Zofia Stangl

Tree growth is affected by temperature and nutrient interactions

Host: Vaughan Hurry

Room: KB3A9, Lilla hörsalen

Tue. 16 Dec, 2014

PhD-Halftime seminar - Tomas Bergström New insights in botulinum toxin analysis by mass spectrometry

Tue. 16 Dec, 2014 13:00 - 14:00
PhD Halftime Seminar
Forest Genetics and Plant Physiology, SLU/ The Swedish Defence Research Agency (FOI)

Tomas Bergström

New insights in botulinum toxin analysis by mass spectrometry

Supervisor: Gunnar Wingsle

Room: Lilla hörsalen, KB3A9, KBC

Seminar - Alison Smith: The leaf at night

Tue. 16 Dec, 2014 14:00 - 15:00
UPSC Seminar

Alison Smith
John Innes Centre, Norwich, UK

The leaf at night

Room. Lilla Hörsalen, KB3A9
Host: Eduoard Pesquet

Wed. 17 Dec, 2014

Dissertation Sabine Kunz

Wed. 17 Dec, 2014 10:00 - 11:00
Thesis title: Sugar-modulated gene expression and cell division in cell culture and seedlings of A. thaliana.
Opponent: Prof. Alison Smith (Dept. of Metabolic Biology, John Innes Centre, Norwich, UK)