Phosphoproteomics: Searching for a needle in a haystack

Tichý, Aleš; Šalovská, Barbora; Řehulka, Pavel; Klimentová, Jana; Vávrová, Jiřina; Stulı́k, Jiřı́; Hernychová, Lenka

doi:10.1016/j.jprot.2011.07.018

Cited by 47 publications

(37 citation statements)

References 95 publications

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“…24 h post-transfer, cells had media changed to lineage-specific media per the manufacture specifications, with media being replaced every other day. Samples for the multilineage differentiation time-course (Study Set #1) were performed in triplicate and began with time zero (no differentiation media added), followed by 33 time points for adipocytic and osteoblastic lineages (terminal differentiation reached at day 14), and 39 for the chondrocytic lineage (terminal differentiation reached at 20 days) (Fig. 1).…”

Section: Methodsmentioning

confidence: 99%

“…CRKII has been predominantly studied in the context of cancer research and implicated in cellular functions such as proliferation, differentiation, and migration (41,42,43). One study examining synovial sarcoma cells discovered a novel signaling mechanism with CRKII activating v-src sarcoma (Schmidt-Ruppin A-2) viral oncogene homolog (SRC) and subsequently p38MAPK, with CRKII suppression leading to decreased proliferation (14). The current study found a similar relationship with CRKII and SRC, although p38MAPK was not implicated because of a lack of correlation with CRKII.…”

Section: Mst1(t183)/mst2(t180)mentioning

confidence: 99%

“…These reversible kinasedriven phosphorylation events alter protein confirmation, ultimately affecting enzymatic activity and protein-protein in-teractions leading to an array of cellular events from differentiation to gene expression, thus encompassing signal transduction. Characterization of this broad-scale signaling architecture is necessary to provide a more finite depiction of the complex signaling events directing a given cellular phenotype and aid in the understanding of the repercussions of alterations in regulation (13,14). Presently, phosphoproteomic analysis of cellular differentiation processes such as with mass spectrometry has been performed in a very limited number of independent time points that span the cellular differentiation process, and important dynamic changes in the cellular signaling architecture may have been missed (12,15,16).…”

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confidence: 99%

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Dynamic Protein Pathway Activation Mapping of Adipose-Derived Stem Cell Differentiation Implicates Novel Regulators of Adipocyte Differentiation

Wilson

Liotta

PetricoinIII

2013

Molecular & Cellular Proteomics

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Next to embryonic stem cell research, adult stem cell research is providing a promising alternative for enhanced tissue regeneration and transplantation. The key biochemical networks controlling the differentiation processes regulating stem cell biology remain largely disputed and or undefined, contributing to a lack of knowledge of the principle phosphoregulatory events propagating signal transduction. To effectively monitor these events relative to adipocyte differentiation, this study utilized a high throughput reverse phase protein microarray platform and characterized adult adipose-derived stem cell (ASC) differentiation through the monitoring of ϳ100 phosphospecific endpoints with 33 distinct time points examined across 14 days. This kinetic-based analysis showed time ordered signal transduction ultimately implicating pathways correlated with adipogenic differentiation. To further validate the causal significance of these network activations, pharmacological targeting was implemented to include the chemical inhibitors MAPK inhibitor PD169316, rapamycin, and HNMPA-(AM) 3 yielding partial or complete disruption of adipocytic differentiation, as noted by a decrease or lack of lipid formation within the mature adipocytes. Based on this analysis, v-crk sarcoma virus CT10 oncogene homolog (CRKII) and c-abl oncogene 1, non-receptor tyrosine kinase (c-ABL) were implicated as novel key regulators of adipocyte differentiation, with v-akt murine thymoma viral oncogene (AKT), mammalian target of rapamycin (mTOR), and SMAD family member (SMAD) pathways being implicated as secondary regulators. This dynamic molecular profiling provides a novel insight into the signaling architecture of mesenchymal stem cell differentiation and may be useful in the development of therapeutic modulators for clinical applications; in addition to advancing the collective understanding of key cellular processes, ultimately contributing to more confident stem cell manipulation. Molecular & Cellular

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Section: Methodsmentioning

confidence: 99%

Section: Mst1(t183)/mst2(t180)mentioning

confidence: 99%

mentioning

confidence: 99%

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Dynamic Protein Pathway Activation Mapping of Adipose-Derived Stem Cell Differentiation Implicates Novel Regulators of Adipocyte Differentiation

Wilson

Liotta

PetricoinIII

2013

Molecular & Cellular Proteomics

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“…Protein phosphorylation is a key regulatory mechanism in biological systems, being a central regulator of intracellular processes [1,2]. Reversible protein phosphorylation is involved in the regulation of various processes, such as cellular signaling and communication, cell homeostasis, differentiation, proliferation and survival, protein degradation, and metabolism, to name a few [3].…”

Section: Introductionmentioning

confidence: 99%

A scoring model for phosphopeptide site localization and its impact on the question of whether to use MSA

Fischer

Santos

Marchini

et al. 2015

Journal of Proteomics

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“…We covered in detail various MS approaches of phosphoproteomic analysis in our previous work (Tichy et al 2011 ). Nevertheless, the MS analysis of phosphoproteins is still complicated because of the relatively low abundance of phosphorylated proteins in eukaryotic cells.…”

Section: Introductionmentioning

confidence: 99%

Enrichment strategies for phosphoproteomics: state-of-the-art

Šalovská¹,

Tichý²,

Řezáčová³

et al. 2012

Reviews in Analytical Chemistry

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Protein phosphorylation is a key regulator in many biological processes, such as homeostasis, cellular signaling and communication, transcriptional and translational regulation, and apoptosis. The defects in this tightly controlled reversible posttranslational modifi cation have been described to contribute to genesis and progression of various diseases, emphasizing the importance of a systematic research of this phenomenon. Although considerable effort has been devoted to improving the analysis of phosphorylation by mass spectrometry, which is currently the method of choice to study protein phosphorylation, the detection and identifi cation of phosphorylation sites remains challenging because of the low abundance and low ionization effi cacy of phosphoproteins in comparison with nonphosphorylated proteins. To overcome this obstacle, different enrichment strategies for phosphorylated peptides/proteins have been established and optimized for subsequent mass spectrometry analysis. In this review, we will give an overview of the methods currently available for the enrichment of phosphorylated proteins and peptides including immunoprecipitation, chemical derivatization and affi nity enrichment techniques.

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Phosphoproteomics: Searching for a needle in a haystack

Cited by 47 publications

References 95 publications

Dynamic Protein Pathway Activation Mapping of Adipose-Derived Stem Cell Differentiation Implicates Novel Regulators of Adipocyte Differentiation

Dynamic Protein Pathway Activation Mapping of Adipose-Derived Stem Cell Differentiation Implicates Novel Regulators of Adipocyte Differentiation

A scoring model for phosphopeptide site localization and its impact on the question of whether to use MSA

Enrichment strategies for phosphoproteomics: state-of-the-art

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