Proteomic and functional profiles of a follicle‐stimulating hormone positive human nonfunctional pituitary adenoma

Abstract

Nonfunctional pituitary adenoma (NFPA) is highly heterogeneous with different hormone-expressed subtypes in NFPA tissues including follicle-stimulating hormone (FSH) positive, luteinizing hormone-positive, FSH/luteinizing hormone-positive, and negative types. To analyze in-depth the variations in the proteomes among different NFPA subtypes for our long-term goal to clarify molecular mechanisms of NFPA and to detect tumor biomarker for personalized medicine practice, a reference map of proteome of a human FSH-expressed NFPA tissue was described here. 2DE and PDQuest image analysis were used to array each protein. MALDI-TOF PMF and human Swiss-Prot databases with MASCOT search were used to identify each protein. A good 2DE pattern with high level of between-gel reproducibility was attained with an average positional deviation 1.98 ± 0.75 mm in the IEF direction and 1.62 ± 0.68 mm in the SDS-PAGE direction. Approximately 1200 protein spots were 2DE-detected and 192 redundant proteins that were contained in 141 protein spots were PMF-identified, representing 107 nonredundant proteins. Those proteins were located in cytoplasm, nucleus, plasma membrane, extracellular space, and so on, and those functioned in transmembrane receptor, ion channel, transcription/translation regulator, transporter, enzyme, phosphatase, kinase, and so on. Several important pathway networks were characterized from those identified proteins with DAVID and Ingenuity Pathway Analysis systems, including gluconeogenesis and glycolysis, mitochondrial dysfunction, oxidative stress, cell-cycle alteration, MAPKsignaling system, immune response, TP53-signaling, VEGF-signaling, and inflammation signaling pathways. Those resulting data contribute to a functional profile of the proteome of a human FSH-positive NFPA tissue, and will serve as a reference for the heterogeneity analysis of NFPA proteomes.

The differentially mitochondrial proteomic dataset in human ovarian cancer relative to control tissues

“…DAVID Bioinformatics Resources 6.7 (https://david.abcc.ncifcrf.gov/home.jsp) was used to analyze the mitochondrial DEPs, including biological process, cellular compartment, molecular function, and KEGG pathway enrichments [6].…”

Revival of 2DE-LC/MS in Proteomics and Its Potential for Large-Scale Study of Human Proteoforms

“…In review of all those 2DE publications, four main contributions of 2DGE in the field of proteomics in the past years ( Fig. 1) are: (i) 2DGE-based comparative proteomics that identified differentially expressed proteins (DEPs) in a given condition relative to its control [43,44] , (ii) 2DGE-based reference map that established the database of a proteome [45,46] , (iii) 2DGE-based Western blot in combination with a specific protein antibody that visually detected variants/proteoforms of that given protein in a proteome [47,48] , and (iv) 2DGE-based Western blot in combination with a specific post-translational modification (PTM) antibody that visually identified a kind of PTM in a proteome [49,50] , and relatively quantified a differential level of a protein in a given condition relative to its control [51,52] . Furthermore, in the past 10-15 years, 2DE protocols have been highly refined in the analysis of a proteome with the use of postfractionation strategy [53] , the change of the detergent composition of the solubilization buffer [54] , or the pre-extraction of sample handling with automated frozen disruption [55] to resolve those traditional 2DE problems such as proteins stacked at pH extremes, the 2D-gel-area obscured by highabundance proteins, unresolved peptides that migrated at the front of separations, and hydrophobic membrane proteomes [53][54][55] .…”

“…Almost all these publications described 2DE as a very high-resolution separation technique with only one or two proteins detected in each 2D spot in a proteome analysis. 2DE-separated proteins commonly identified with MALDI-MS PMF, MALDI-MS/MS, or LC-MS/MS with determination of the firstor second-ranked proteins in the list of searched proteins as the result proteins in a 2D spot [8][9][10][11][12][13] . These types of 2DE-based data are collected in many 2DGE resources such as the World-2DPAGE Constellation (http://world-2dpage.expasy.org) [14] , which include the World-2DPAGE list (http://world-2dpage.expasy.…”

“…Thus, if maximizing the total number of identified proteins in a proteome is the objective, then 2DE-MS is commonly seen as a seemingly "low-throughput" approach, compared to seemingly "highthroughput" bottom-up non-gel methods [15][16][17] such as isobaric tags for relative and absolute quantification (iTRAQ) [18,19] , peptide tandem mass tags (TMT) [20,21] , stable isotope labeling of amino acids in cell culture (SILAC) [22,23] , or label free [24,25] based quantitative proteomics. When one studies very precious, hard-to-obtain human brain samples such as, for example, pituitary tissue, then 2D gels provide a crucially important method to collect, store, and archive that human proteome [3,6,[10][11][12] . 2DLC-MS methods can never provide that necessary archival storage advantage.…”

Editorial: Molecular Network Study of Pituitary Adenomas

“…We strongly recommend the use of multiomics to study pituitary adenoma invasiveness, which might be the right way to resolve its clinical invasiveness challenge for clarification of its molecular mechanisms, discovery of effective therapeutic targets, and determination of effective biomarkers for diagnosis and prognostic assessment. Some proteomics and transcriptomics between invasive and non-invasive pituitary adenomas have been performed to understand molecular mechanism and discover biomarkers of pituitary adenoma invasiveness (24)(25)(26). (iv) Proteome is the final functional performer of genome and transcriptome.…”