Our research is interdisciplinary. We develop and utilize proteomics tools to study cell-surface N-glycoproteins that play important roles in stem-cell development. Through various model systems, we want to understand how extra-cellular microenvironment influences stem-cell function and behavior through these proteins and their N-glycosylation.
LC-MS based shortgun proteomics is a powerful technique that utilizes HPLC to deliver nanoliter per minute constant flow under hundreds to thousands fold of atmosphere pressure for extremely high resolution separation and for downstream accurate mass spectrometry analysis, in which millions of peptides from thousands of proteins can be measured in less than an hour. Doing so, the technique is able to detect and quantify the entire proteome of any biological samples and build global protein interaction networks for systematic study of human health and diseases.
1. Development of sensitive proteomic methods for cell surface proteins.
Life fascinated us by turning a single fertilized embryo to a complex body plan, in which each tissue and organ has its unique morphology, function, and molecular composition. Together organisms display remarkable skills in survival and prosperity by orchestrating intricate interactions between different body parts. Using mouse as a model and by studying both mouse embryonic stem cells and their terminally differentiated organs and tissues, we would like to track molecular roots of the observed changes and interactions occurred in development, homeostasis, and pathological conditions. The obtained information will help our better understanding of stem-cells fate commitment, and how our body functions as a whole in disease diagnosis and treatment through organ specific biomarkers. Recommended reading: (1) Sun, B.*, Hood, L., “Protein-centric proteomics analysis of membrane and plasma membrane proteins.”, J. Proteome Res., 2014, 13(6), 2705-2714. (2) Sun, B.*, “Proteomics and glycoproteomics of pluripotent stem-cell surface proteins”, Proteomics, 2015, 15, 1152-1163. (3) Lee, G.; and Sun, B.*, “Glycopeptide capture for cell surface proteomics”, JoVE, 2014, 87, doi: 10.3791/51349. (4) Lee, G.; Wu, K.; Tam, N.; Sun, B.*, “Sodium dodecyl sulfate polyacrylamide gel electrophoresis for direct quantitation of protein adsorption”, Anal. Biochem., 2014, 102-104. (5) Sun, B.*; Li, M.; Yan, X.; Lee, D.; et al., “N-glycoproteome of the E14.Tg2a mouse embryonic stem cells”, PLOS ONE, 2013, 8(2): e55722. DOI:10.1371. (6) Zhang, Y.; Akintola,S. O.; Liu, J. K.; and Sun, B*, “Membrane gene ontology bias in sequencing and microarray obtained by housekeeping-gene analysis”, 2016, Gene, 575, 559-566.
Life fascinated us by turning a single fertilized embryo to a complex body plan, in which each tissue and organ has its unique morphology, function, and molecular composition. Together organisms display remarkable skills in survival and prosperity by orchestrating intricate interactions between different body parts. Using mouse as a model and by studying both mouse embryonic stem cells and their terminally differentiated organs and tissues, we would like to track molecular roots of the observed changes and interactions occurred in development, homeostasis, and pathological conditions. The obtained information will help our better understanding of stem-cells fate commitment, and how our body functions as a whole in disease diagnosis and treatment through organ specific biomarkers. Recommended reading: (1) Sun, B.*, Hood, L., “Protein-centric proteomics analysis of membrane and plasma membrane proteins.”, J. Proteome Res., 2014, 13(6), 2705-2714. (2) Sun, B.*, “Proteomics and glycoproteomics of pluripotent stem-cell surface proteins”, Proteomics, 2015, 15, 1152-1163. (3) Lee, G.; and Sun, B.*, “Glycopeptide capture for cell surface proteomics”, JoVE, 2014, 87, doi: 10.3791/51349. (4) Lee, G.; Wu, K.; Tam, N.; Sun, B.*, “Sodium dodecyl sulfate polyacrylamide gel electrophoresis for direct quantitation of protein adsorption”, Anal. Biochem., 2014, 102-104. (5) Sun, B.*; Li, M.; Yan, X.; Lee, D.; et al., “N-glycoproteome of the E14.Tg2a mouse embryonic stem cells”, PLOS ONE, 2013, 8(2): e55722. DOI:10.1371. (6) Zhang, Y.; Akintola,S. O.; Liu, J. K.; and Sun, B*, “Membrane gene ontology bias in sequencing and microarray obtained by housekeeping-gene analysis”, 2016, Gene, 575, 559-566.
2. Stem-cell and organismal proteomics to decoding tissue and organ networks.
Due to their unique position at interface of external environment and intracellular molecular machinery, cell surface proteins are an important protein class. They not only provide the fingerprint of cell identity, but also function in a number of crucial biological processes such as adhesion and migration, nutrient and waste transport, sensing, signaling, as well as defence. In stem cells, surface proteins interact with extracellular environment (i.e. niche) to coax stem-cell self renewal, differentiation, lineage specification, and homing. In silico analysis suggests that one third to half of the genome encodes membrane proteins, yet only a small number of these proteins have been characterized and less than 5% proteins have their structures resolved. The reason is because these proteins are lowly expressed and many of them have fewer than 100 copies in a cell. In addition, they are difficult to dissolve in aqueous solution, and likely to get lost in sample preparation due to non-specific adsorption to solid surfaces. Global proteomics and transcriptomics including RNA-seq often miss them. We are using sensitive LC-MS proteomics and developing various enrichment approaches to better detect and characterize these proteins. Recommended reading: (1) Zhang, Y.; Li, D.; Sun, B.*; “Do housekeeping genes exist?” , PLoS ONE, 2015, 10(5): DOI: 10.1371. (2) Sun, B.*; Ranish, J.A.; Utleg, A.G.; et al. “Glycocapture-Assisted Global Quantitative Proteomics (gagQP) reveals multiorgan responses in blood toxicoproteome” , J. Proteome Res., 2013, 12 (5), 2034–2044. (3) Zhang, Y.; Akintola,S. O.; Liu, J. K.; and Sun, B*, “Detection bias in microarray and sequencing transcriptomic analysis identified by housekeeping genes”, 2015, Data In Brief, 6, 121-123.
Due to their unique position at interface of external environment and intracellular molecular machinery, cell surface proteins are an important protein class. They not only provide the fingerprint of cell identity, but also function in a number of crucial biological processes such as adhesion and migration, nutrient and waste transport, sensing, signaling, as well as defence. In stem cells, surface proteins interact with extracellular environment (i.e. niche) to coax stem-cell self renewal, differentiation, lineage specification, and homing. In silico analysis suggests that one third to half of the genome encodes membrane proteins, yet only a small number of these proteins have been characterized and less than 5% proteins have their structures resolved. The reason is because these proteins are lowly expressed and many of them have fewer than 100 copies in a cell. In addition, they are difficult to dissolve in aqueous solution, and likely to get lost in sample preparation due to non-specific adsorption to solid surfaces. Global proteomics and transcriptomics including RNA-seq often miss them. We are using sensitive LC-MS proteomics and developing various enrichment approaches to better detect and characterize these proteins. Recommended reading: (1) Zhang, Y.; Li, D.; Sun, B.*; “Do housekeeping genes exist?” , PLoS ONE, 2015, 10(5): DOI: 10.1371. (2) Sun, B.*; Ranish, J.A.; Utleg, A.G.; et al. “Glycocapture-Assisted Global Quantitative Proteomics (gagQP) reveals multiorgan responses in blood toxicoproteome” , J. Proteome Res., 2013, 12 (5), 2034–2044. (3) Zhang, Y.; Akintola,S. O.; Liu, J. K.; and Sun, B*, “Detection bias in microarray and sequencing transcriptomic analysis identified by housekeeping genes”, 2015, Data In Brief, 6, 121-123.
3. Structure and function characterization of membrane proteins.
Membrane proteins possess unique structures as they reside in lipid bilayer and relay signals across membrane or function as tunnels to transport nutrients, drugs, and metabolic waste. More than 50% drugs target to membrane proteins, yet less than 5% membrane proteins have crystal structures due to their low abundance, heavy post-translational modification, large size, and low solubility in aqueous solutions. Using high throughput LC-MS proteomics and bioinformatics, we are developing methods to characterize membrane protein structure at global scale. Recommended reading: (1) Sun, B.*; Li, M.; Yan, X.; Lee, D.; et al., “N-glycoproteome of the E14.Tg2a mouse embryonic stem cells” , PLOS ONE, 2013, 8(2): e55722. DOI:10.1371.
Membrane proteins possess unique structures as they reside in lipid bilayer and relay signals across membrane or function as tunnels to transport nutrients, drugs, and metabolic waste. More than 50% drugs target to membrane proteins, yet less than 5% membrane proteins have crystal structures due to their low abundance, heavy post-translational modification, large size, and low solubility in aqueous solutions. Using high throughput LC-MS proteomics and bioinformatics, we are developing methods to characterize membrane protein structure at global scale. Recommended reading: (1) Sun, B.*; Li, M.; Yan, X.; Lee, D.; et al., “N-glycoproteome of the E14.Tg2a mouse embryonic stem cells” , PLOS ONE, 2013, 8(2): e55722. DOI:10.1371.
4. Environmental modulation of N-glycosylation.
N-glycosylation is the dominant protein post-translational modification. It functions as an anchorage signal in protein trafficking, and also assists protein folding and increases protein stability at cell surface or extracellular milieu. Defects in N-glycosylation can cause severe congenital disorders, and are also linked to cancer, cardiovascular and immunological diseases among others. Environmental factors are known to influence the degree of glycosylation, i.e. the occupancy of N-glycosylation sites (macroheterogeneity). The regulation mechanism is complex and involves the bioavailability of sugar, nucleotides, enzymes, chaperones and transporters. Proteomics therefore is an ideal approach to elucidate these molecular changes holistically. We are investigating how the external sugar supplements can affect cellular protein glycosylation, and revealing modulation mechanism to control stem-cell behavior and function. Recommended reading: (1) Akintola, S. O. and Sun, B*, “N-acetylgucosamine as a regulant of CHO-K1 cell migration”, Annual Canadian National Proteomics Network Symposium, Montreal, QC
N-glycosylation is the dominant protein post-translational modification. It functions as an anchorage signal in protein trafficking, and also assists protein folding and increases protein stability at cell surface or extracellular milieu. Defects in N-glycosylation can cause severe congenital disorders, and are also linked to cancer, cardiovascular and immunological diseases among others. Environmental factors are known to influence the degree of glycosylation, i.e. the occupancy of N-glycosylation sites (macroheterogeneity). The regulation mechanism is complex and involves the bioavailability of sugar, nucleotides, enzymes, chaperones and transporters. Proteomics therefore is an ideal approach to elucidate these molecular changes holistically. We are investigating how the external sugar supplements can affect cellular protein glycosylation, and revealing modulation mechanism to control stem-cell behavior and function. Recommended reading: (1) Akintola, S. O. and Sun, B*, “N-acetylgucosamine as a regulant of CHO-K1 cell migration”, Annual Canadian National Proteomics Network Symposium, Montreal, QC