Next‐generation technologies for spatial proteomics: Integrating ultra‐high speed MALDI‐TOF and high mass resolution MALDI FTICR imaging mass spectrometry for protein analysis

Spraggins, Jeffrey M.; Rizzo, David G.; Moore, Jessica L.; Noto, Michael J.; Skaar, Eric P.; Caprioli, Richard M.

doi:10.1002/pmic.201600003

Cited by 147 publications

(142 citation statements)

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“…Notably, recent technological advances are merging omics with histological analysis; these approaches are known as ‘spatial genomics’ [39] or ‘spatial proteomics’ [40, 41]. Single-cell RNA sequencing or mass spectrometry in situ (preserving the spatial location of the studied cells) promises to expose new layers of beta cell biology and heterogeneity.…”

Section: Multihormonal Islet Cellsmentioning

confidence: 99%

Beta cell heterogeneity: an evolving concept

Avrahami

Klochendler²,

Dor³

et al. 2017

Diabetologia

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Beta cells are primarily defined by their ability to produce insulin and secrete it in response to appropriate stimuli. It has been known for some time, however, that beta cells are not functionally identical to each other and that the rates of insulin synthesis and release differ from cell to cell, although the functional significance of this variability remains unclear. Recent studies have used heterogeneous gene expression to isolate and evaluate different subpopulations of beta cells and to demonstrate alterations in these subpopulations in diabetes. In the last few years, novel technologies have emerged that permit the detailed evaluation of the proteome (e.g. time-of-flight mass spectroscopy, [CyTOF]) and transcriptome (e.g. massively parallel RNA sequencing) at the single-cell level, and tools for single beta cell metabolomics and epigenomics are quickly maturing. The first wave of single beta cell proteome and transcriptome studies were published in 2016, giving a glimpse into the power, but also the limitations, of these approaches. Despite this progress, it remains unclear if the observed heterogeneity of beta cells represents stable, distinct beta cell types or, alternatively, highly dynamic beta cell states. Here we provide a concise overview of recent developments in the emerging field of beta cell heterogeneity and the implications for our understanding of beta cell biology and pathology.

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Section: Multihormonal Islet Cellsmentioning

confidence: 99%

Beta cell heterogeneity: an evolving concept

Avrahami

Klochendler²,

Dor³

et al. 2017

Diabetologia

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“…However, a challenge in the construction of protein microarrays is that the proteins must be produced and purified from biological systems that allow proteins undergo posttranslational modifications and folding so that they retain their functions on the chip (Popescu et al, 2007). Proteins can also be separated using 1-dimensional (based on size only) or 2-dimensional (based on charge and size) protein gel electrophoresis (Gallagher, 2006; Rabilloud and Lelong, 2011), or chromatography (e.g., ultra-high speed MALDI-TOF and high mass resolution MALDI FTICR imaging MS) (Spraggins et al, 2016). A comparative proteomic analysis of two B. cinerea strains 1.11 and 2100 identified proteins that play crucial roles in their differential virulence, including housekeeping enzymes, such as malate and glyceraldehyde dehydrogenases (Fernández-Acero et al, 2007).…”

Section: Proteomicsmentioning

confidence: 99%

‘Omics’ and Plant Responses to Botrytis cinerea

2016

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Botrytis cinerea is a dangerous plant pathogenic fungus with wide host ranges. This aggressive pathogen uses multiple weapons to invade and cause serious damages on its host plants. The continuing efforts of how to solve the “puzzle” of the multigenic nature of B. cinerea’s pathogenesis and plant defense mechanisms against the disease caused by this mold, the integration of omic approaches, including genomics, transcriptomics, proteomics and metabolomics, along with functional analysis could be a potential solution. Omic studies will provide a foundation for development of genetic manipulation and breeding programs that will eventually lead to crop improvement and protection. In this mini-review, we will highlight the current progresses in research in plant stress responses to B. cinerea using high-throughput omic technologies. We also discuss the opportunities that omic technologies can provide to research on B. cinerea-plant interactions as an example showing the impacts of omics on agricultural research.

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“…The analysis of their spatial distribution can provide additional information about their role in physiological and pathological processes [10]. Numerous studies have been published on MS imaging of proteins, examples include references [11–13]. Lipids are usually removed by a series of washing steps prior to analysis [14].…”

Section: Introductionmentioning

confidence: 99%

Approaching cellular resolution and reliable identification in mass spectrometry imaging of tryptic peptides

et al. 2018

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On-tissue digestion has become the preferred method to identify proteins in mass spectrometry (MS) imaging. In this study, we report advances in data acquisition and protein identification for MS imaging after on-tissue digestion. Tryptic peptides in a coronal mouse brain section were measured at 50 μm pixel size and revealed detailed histological structures, e.g., the ependyma (consisting of one to two cell layers), which was confirmed by H&E staining. This demonstrates that MS imaging of tryptic peptides at or close to cellular resolution is within reach. We also describe a detailed identification workflow which resulted in the identification of 99 proteins (with 435 corresponding peptides), based on comparison with LC-MS/MS data and in silico digest. These results were obtained with stringent parameters, including high mass accuracy in imaging mode (RSME < 3 ppm) and at least two unique peptides per protein showing consistent spatial distribution. We identified almost 50% of proteins with at least four corresponding peptides. As there is no agreed approach for identification of proteins after on-tissue digestion yet, we discuss our workflow in detail and make the corresponding mass spectral data available as “open data” via ProteomeXchange (identifier PXD003172). With this, we would like to contribute to a more effective discussion and the development of new approaches for tryptic peptide identification in MS imaging. From an experimental point of view, we demonstrate the improvement due to the combination of high spatial resolution and high mass resolution/mass accuracy on a measurement at 25 μm pixel size in mouse cerebellum tissue. A whole body section of a mouse pub imaged at 50 μm pixel size (40 GB, 230,000 spectra) demonstrates the stability of our protocol. For this data set, we developed a workflow that is based on conversion to the common data format imzML and sequential application of freely available software tools. In combination, the presented results for spatial resolution, protein identification, and data processing constitute significant improvements for the field of on-tissue digestion. Graphical abstractMS imaging of coronal mouse brain cerebellum with a pixel size of 25 μm: A Optical image, B myelin staining, C H&E staining, and D MS image overlay (RGB) of tryptic peptides m/z = 726.4045 ± 0.005, HGFLPR + H+ (red), m/z = 536.3173 ± 0.005, AKPAK + Na+ (green), and m/z = 994.5436 ± 0.005, WRQLIEK + Na+ (blue) Electronic supplementary materialThe online version of this article (10.1007/s00216-018-1199-z) contains supplementary material, which is available to authorized users.

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Next‐generation technologies for spatial proteomics: Integrating ultra‐high speed MALDI‐TOF and high mass resolution MALDI FTICR imaging mass spectrometry for protein analysis

Cited by 147 publications

References 50 publications

Beta cell heterogeneity: an evolving concept

Beta cell heterogeneity: an evolving concept

‘Omics’ and Plant Responses to Botrytis cinerea

Approaching cellular resolution and reliable identification in mass spectrometry imaging of tryptic peptides

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