Comprehensive quantitative comparison of the membrane proteome, phosphoproteome, and sialiome of human embryonic and neural stem cells

Marcella Nunes Melo-Braga, Melanie Schulz, Qiuyue Liu, Andrzej Swistowski, Giuseppe Palmisano, Kasper Engholm-Keller, Lene Jakobsen, Xianmin Zeng, Martin Røssel Larsen

Research output: Contribution to journalArticle

Abstract

Human embryonic stem cells (hESCs) can differentiate into neural stem cells (NSCs), which can further be differentiated into neurons and glia cells. Therefore, these cells have huge potential as source for treatment of neurological diseases. Membrane-associated proteins are very important in cellular signaling and recognition, and their function and activity are frequently regulated by posttranslational modifications such as phosphorylation and glycosylation. To obtain information about membraneassociated proteins and their modified amino acids potentially involved in changes of hESCs and NSCs as well as to investigate potential new markers for these two cell stages, we performed large-scale quantitative membrane- proteomic of hESCs and NSCs. This approach employed membrane purification followed by peptide dimethyl labeling and peptide enrichment to study the membrane subproteome as well as changes in phosphorylation and sialylation between hESCs and NSCs. Combining proteomics and modification specific proteomics we identified a total of 5105 proteins whereof 57% contained transmembrane domains or signal peptides. The enrichment strategy yielded a total of 10,087 phosphorylated peptides in which 78% of phosphopeptides were identified with ≥99% confidence in site assignment and 1810 unique formerly sialylated N-linked glycopeptides. Several proteins were identified as significantly regulated in hESCs and NSC, including proteins involved in the early embryonic and neural development. In the latter group of proteins, we could identify potential NSC markers as Crumbs 2 and several novel proteins. A motif analysis of the altered phosphosites showed a sequence consensus motif (R-X-XpS/T) significantly up-regulated in NSC. This motif is among other kinases recognized by the calmodulin- dependent protein kinase-2, emphasizing a possible importance of this kinase for this cell stage. Collectively, this data represent the most diverse set of post-translational modifications reported for hESCs and NSCs. This study revealed potential markers to distinguish NSCs from hESCs and will contribute to improve our understanding on the differentiation process. Molecular & Cellular Proteomics 13: 10.1074/mcp.M112.026898, 311-328, 2014.

Original languageEnglish
Pages (from-to)311-328
Number of pages18
JournalMolecular and Cellular Proteomics
Volume13
Issue number1
DOIs
Publication statusPublished - Jan 2014

Fingerprint

Neural Stem Cells
Proteome
Stem cells
Membranes
Proteomics
Proteins
Post Translational Protein Processing
Peptides
Phosphotransferases
Phosphorylation
Human Embryonic Stem Cells
Phosphopeptides
Calcium-Calmodulin-Dependent Protein Kinases
Glycopeptides
Consensus Sequence
Protein Sorting Signals
Glycosylation
Neuroglia
Embryonic Development
Membrane Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Analytical Chemistry

Cite this

Comprehensive quantitative comparison of the membrane proteome, phosphoproteome, and sialiome of human embryonic and neural stem cells. / Melo-Braga, Marcella Nunes; Schulz, Melanie; Liu, Qiuyue; Swistowski, Andrzej; Palmisano, Giuseppe; Engholm-Keller, Kasper; Jakobsen, Lene; Zeng, Xianmin; Larsen, Martin Røssel.

In: Molecular and Cellular Proteomics, Vol. 13, No. 1, 01.2014, p. 311-328.

Research output: Contribution to journalArticle

Melo-Braga, MN, Schulz, M, Liu, Q, Swistowski, A, Palmisano, G, Engholm-Keller, K, Jakobsen, L, Zeng, X & Larsen, MR 2014, 'Comprehensive quantitative comparison of the membrane proteome, phosphoproteome, and sialiome of human embryonic and neural stem cells', Molecular and Cellular Proteomics, vol. 13, no. 1, pp. 311-328. https://doi.org/10.1074/mcp.M112.026898
Melo-Braga, Marcella Nunes ; Schulz, Melanie ; Liu, Qiuyue ; Swistowski, Andrzej ; Palmisano, Giuseppe ; Engholm-Keller, Kasper ; Jakobsen, Lene ; Zeng, Xianmin ; Larsen, Martin Røssel. / Comprehensive quantitative comparison of the membrane proteome, phosphoproteome, and sialiome of human embryonic and neural stem cells. In: Molecular and Cellular Proteomics. 2014 ; Vol. 13, No. 1. pp. 311-328.
@article{415d42845ba0450a89a8a98d00226c72,
title = "Comprehensive quantitative comparison of the membrane proteome, phosphoproteome, and sialiome of human embryonic and neural stem cells",
abstract = "Human embryonic stem cells (hESCs) can differentiate into neural stem cells (NSCs), which can further be differentiated into neurons and glia cells. Therefore, these cells have huge potential as source for treatment of neurological diseases. Membrane-associated proteins are very important in cellular signaling and recognition, and their function and activity are frequently regulated by posttranslational modifications such as phosphorylation and glycosylation. To obtain information about membraneassociated proteins and their modified amino acids potentially involved in changes of hESCs and NSCs as well as to investigate potential new markers for these two cell stages, we performed large-scale quantitative membrane- proteomic of hESCs and NSCs. This approach employed membrane purification followed by peptide dimethyl labeling and peptide enrichment to study the membrane subproteome as well as changes in phosphorylation and sialylation between hESCs and NSCs. Combining proteomics and modification specific proteomics we identified a total of 5105 proteins whereof 57{\%} contained transmembrane domains or signal peptides. The enrichment strategy yielded a total of 10,087 phosphorylated peptides in which 78{\%} of phosphopeptides were identified with ≥99{\%} confidence in site assignment and 1810 unique formerly sialylated N-linked glycopeptides. Several proteins were identified as significantly regulated in hESCs and NSC, including proteins involved in the early embryonic and neural development. In the latter group of proteins, we could identify potential NSC markers as Crumbs 2 and several novel proteins. A motif analysis of the altered phosphosites showed a sequence consensus motif (R-X-XpS/T) significantly up-regulated in NSC. This motif is among other kinases recognized by the calmodulin- dependent protein kinase-2, emphasizing a possible importance of this kinase for this cell stage. Collectively, this data represent the most diverse set of post-translational modifications reported for hESCs and NSCs. This study revealed potential markers to distinguish NSCs from hESCs and will contribute to improve our understanding on the differentiation process. Molecular & Cellular Proteomics 13: 10.1074/mcp.M112.026898, 311-328, 2014.",
author = "Melo-Braga, {Marcella Nunes} and Melanie Schulz and Qiuyue Liu and Andrzej Swistowski and Giuseppe Palmisano and Kasper Engholm-Keller and Lene Jakobsen and Xianmin Zeng and Larsen, {Martin R{\o}ssel}",
year = "2014",
month = "1",
doi = "10.1074/mcp.M112.026898",
language = "English",
volume = "13",
pages = "311--328",
journal = "Molecular and Cellular Proteomics",
issn = "1535-9476",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "1",

}

TY - JOUR

T1 - Comprehensive quantitative comparison of the membrane proteome, phosphoproteome, and sialiome of human embryonic and neural stem cells

AU - Melo-Braga, Marcella Nunes

AU - Schulz, Melanie

AU - Liu, Qiuyue

AU - Swistowski, Andrzej

AU - Palmisano, Giuseppe

AU - Engholm-Keller, Kasper

AU - Jakobsen, Lene

AU - Zeng, Xianmin

AU - Larsen, Martin Røssel

PY - 2014/1

Y1 - 2014/1

N2 - Human embryonic stem cells (hESCs) can differentiate into neural stem cells (NSCs), which can further be differentiated into neurons and glia cells. Therefore, these cells have huge potential as source for treatment of neurological diseases. Membrane-associated proteins are very important in cellular signaling and recognition, and their function and activity are frequently regulated by posttranslational modifications such as phosphorylation and glycosylation. To obtain information about membraneassociated proteins and their modified amino acids potentially involved in changes of hESCs and NSCs as well as to investigate potential new markers for these two cell stages, we performed large-scale quantitative membrane- proteomic of hESCs and NSCs. This approach employed membrane purification followed by peptide dimethyl labeling and peptide enrichment to study the membrane subproteome as well as changes in phosphorylation and sialylation between hESCs and NSCs. Combining proteomics and modification specific proteomics we identified a total of 5105 proteins whereof 57% contained transmembrane domains or signal peptides. The enrichment strategy yielded a total of 10,087 phosphorylated peptides in which 78% of phosphopeptides were identified with ≥99% confidence in site assignment and 1810 unique formerly sialylated N-linked glycopeptides. Several proteins were identified as significantly regulated in hESCs and NSC, including proteins involved in the early embryonic and neural development. In the latter group of proteins, we could identify potential NSC markers as Crumbs 2 and several novel proteins. A motif analysis of the altered phosphosites showed a sequence consensus motif (R-X-XpS/T) significantly up-regulated in NSC. This motif is among other kinases recognized by the calmodulin- dependent protein kinase-2, emphasizing a possible importance of this kinase for this cell stage. Collectively, this data represent the most diverse set of post-translational modifications reported for hESCs and NSCs. This study revealed potential markers to distinguish NSCs from hESCs and will contribute to improve our understanding on the differentiation process. Molecular & Cellular Proteomics 13: 10.1074/mcp.M112.026898, 311-328, 2014.

AB - Human embryonic stem cells (hESCs) can differentiate into neural stem cells (NSCs), which can further be differentiated into neurons and glia cells. Therefore, these cells have huge potential as source for treatment of neurological diseases. Membrane-associated proteins are very important in cellular signaling and recognition, and their function and activity are frequently regulated by posttranslational modifications such as phosphorylation and glycosylation. To obtain information about membraneassociated proteins and their modified amino acids potentially involved in changes of hESCs and NSCs as well as to investigate potential new markers for these two cell stages, we performed large-scale quantitative membrane- proteomic of hESCs and NSCs. This approach employed membrane purification followed by peptide dimethyl labeling and peptide enrichment to study the membrane subproteome as well as changes in phosphorylation and sialylation between hESCs and NSCs. Combining proteomics and modification specific proteomics we identified a total of 5105 proteins whereof 57% contained transmembrane domains or signal peptides. The enrichment strategy yielded a total of 10,087 phosphorylated peptides in which 78% of phosphopeptides were identified with ≥99% confidence in site assignment and 1810 unique formerly sialylated N-linked glycopeptides. Several proteins were identified as significantly regulated in hESCs and NSC, including proteins involved in the early embryonic and neural development. In the latter group of proteins, we could identify potential NSC markers as Crumbs 2 and several novel proteins. A motif analysis of the altered phosphosites showed a sequence consensus motif (R-X-XpS/T) significantly up-regulated in NSC. This motif is among other kinases recognized by the calmodulin- dependent protein kinase-2, emphasizing a possible importance of this kinase for this cell stage. Collectively, this data represent the most diverse set of post-translational modifications reported for hESCs and NSCs. This study revealed potential markers to distinguish NSCs from hESCs and will contribute to improve our understanding on the differentiation process. Molecular & Cellular Proteomics 13: 10.1074/mcp.M112.026898, 311-328, 2014.

UR - http://www.scopus.com/inward/record.url?scp=84891777324&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84891777324&partnerID=8YFLogxK

U2 - 10.1074/mcp.M112.026898

DO - 10.1074/mcp.M112.026898

M3 - Article

C2 - 24173317

AN - SCOPUS:84891777324

VL - 13

SP - 311

EP - 328

JO - Molecular and Cellular Proteomics

JF - Molecular and Cellular Proteomics

SN - 1535-9476

IS - 1

ER -