SCoPE-MS: mass spectrometry of single mammalian cells quantifies proteome heterogeneity during cell differentiation

Budnik, Bogdan; Levy, Ezra; Harmange, Guillaume; Slavov, Nikolai

doi:10.1186/s13059-018-1547-5

Cited by 701 publications

(782 citation statements)

References 57 publications

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“…Dorus et al, 2006;Baker et al, 2008;Poland et al, 2011;Skerget et al, 2013;Amaral et al, 2014;Ma et al, 2014;Whittington et al, 2015;Degner et al, 2019). As methods for single-cell analysis improve (Budnik et al, 2018), single-cell proteomics could be used to inventory sperm composition across the entire life history of sperm in the FRT. The consequent determination of gains, losses and modifications of sperm (or sperm-associated seminal or female) proteins will provide molecular precision to our understanding of the sequence of changes in sperm as they reside in the FRT.…”

Section: Future Directionsmentioning

confidence: 99%

Post‐ejaculatory modifications to sperm (PEMS)

Pitnick¹,

Wolfner

Dorus³

2019

Biological Reviews

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Mammalian sperm must spend a minimum period of time within a female reproductive tract to achieve the capacity to fertilize oocytes. This phenomenon, termed sperm ‘capacitation’, was discovered nearly seven decades ago and opened a window into the complexities of sperm–female interaction. Capacitation is most commonly used to refer to a specific combination of processes that are believed to be widespread in mammals and includes modifications to the sperm plasma membrane, elevation of intracellular cyclic AMP levels, induction of protein tyrosine phosphorylation, increased intracellular Ca2+ levels, hyperactivation of motility, and, eventually, the acrosome reaction. Capacitation is only one example of post‐ejaculatory modifications to sperm (PEMS) that are widespread throughout the animal kingdom. Although PEMS are less well studied in non‐mammalian taxa, they likely represent the rule rather than the exception in species with internal fertilization. These PEMS are diverse in form and collectively represent the outcome of selection fashioning complex maturational trajectories of sperm that include multiple, sequential phenotypes that are specialized for stage‐specific functionality within the female. In many cases, PEMS are critical for sperm to migrate successfully through the female reproductive tract, survive a protracted period of storage, reach the site of fertilization and/or achieve the capacity to fertilize eggs. We predict that PEMS will exhibit widespread phenotypic plasticity mediated by sperm–female interactions. The successful execution of PEMS thus has important implications for variation in fitness and the operation of post‐copulatory sexual selection. Furthermore, it may provide a widespread mechanism of reproductive isolation and the maintenance of species boundaries. Despite their possible ubiquity and importance, the investigation of PEMS has been largely descriptive, lacking any phylogenetic consideration with regard to divergence, and there have been no theoretical or empirical investigations of their evolutionary significance. Here, we (i) clarify PEMS‐related nomenclature; (ii) address the evolutionary origin, maintenance and divergence in PEMS in the context of the protracted life history of sperm and the complex, selective environment of the female reproductive tract; (iii) describe taxonomically widespread types of PEMS: sperm activation, chemotaxis and the dissociation of sperm conjugates; (iv) review the occurence of PEMS throughout the animal kingdom; (v) consider alternative hypotheses for the adaptive value of PEMS; (vi) speculate on the evolutionary implications of PEMS for genomic architecture, sexual selection, and reproductive isolation; and (vii) suggest fruitful directions for future functional and evolutionary analyses of PEMS.

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Section: Future Directionsmentioning

confidence: 99%

Post‐ejaculatory modifications to sperm (PEMS)

Pitnick¹,

Wolfner

Dorus³

2019

Biological Reviews

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“…Next to the development of dedicated low-input sample preparation methods [1][2][3][4], which aim at reducing sample losses prior to analysis, changes to the chromatographic support material have recently been identified key to the sensitive profiling of low-input proteomics samples by nano-HPLC-ESI-MS/MS [5,6].…”

Section: Technical Briefmentioning

confidence: 99%

Improved Sensitivity in Low-Input Proteomics using Micro-Pillar Array-based Chromatography

Stadlmann

Hudecz

Krššáková

et al. 2019

Preprint

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Capitalizing on the massive increase in sample concentrations which are produced by extremely low elution volumes, nano-LC-ESI-MS/MS is currently one of the most sensitive analytical technologies for the comprehensive characterization of complex protein samples.However, despite tremendous technological improvements made in the production and the packing of monodisperse spherical particles for nano-flow HPLC, current state-of-the-art systems still suffer from limits in operation at the maximum potential of the technology.With the recent introduction of the µPAC system, which provides perfectly ordered micro-pillar array based chromatographic support materials, completely new chromatographic concepts for optimization towards the needs of ultra-sensitive proteomics become available.Here we report on a series of benchmarking experiments comparing the performance of a commercially available 50 cm micro-pillar array column to a widely used nano-flow HPLC column for the proteomics analysis of 10 ng tryptic HeLa cell digest.Comparative analysis of LC-MS/MS-data corroborated that micro-pillar array cartridges provide outstanding chromatographic performance, excellent retention time stability, increase sensitivity in the analysis of low-input proteomics samples, and thus repeatedly yielded almost twice as many unique peptide and unique protein group identifications when compared to conventional nano-flow HPLC columns.

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“…38 A comprehensive review of LC-MS/MS QC and optimization tools has been published by Bittremieux et al 39 We found these tools useful in developing Single-Cell Proteomics by Mass Spectrometry (SCoPE-MS), which combines TMT-labeled peptides from single cells with a TMT-labeled carrier channel to enable quantifying proteins across many single cells. [25][26][27]40 However, none of these tools provided all of the metrics needed for optimizating our SCoPE-MS analysis. 40 This motivated us to develop DO-MS, a highly modular and interactive environment for optimizing ultra-sensitive LC-MS/MS methods.…”

Section: Introductionmentioning

confidence: 99%

DO-MS: Data-Driven Optimization of Mass Spectrometry Methods

Huffman¹,

Specht²,

Chen³

et al. 2019

Preprint

Self Cite

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The performance of ultrasensitive LC-MS/MS methods, such as Single-Cell Proteomics by Mass Spectrometry (SCoPE-MS), depends on multiple interdependent parameters. This interdependence makes it challenging to specifically pinpoint bottlenecks in the LC-MS/MS methods and approaches for resolving them. For example, low signal at MS2 level can be due to poor LC separation, ionization, apex targeting, ion transfer, or ion detection. We sought to specifically diagnose such bottlenecks by interactively visualizing data from all levels of bottom-up LC-MS/MS analysis. Many search engines, such as MaxQuant, already provide such data, and we developed an open source platform for their interactive visualization and analysis: Data-driven Optimization of MS (DO-MS). We found that in many cases DO-MS not only specifically diagnosed bottlenecks but also enabled us to rationally optimize them. For example, we used DO-MS to diagnose poor sampling of the elution peak apex and to optimize it, which increased the efficiency of delivering ions for MS2 analysis by 370%. DO-MS is easy to install and use, and its GUI allows for interactive data subsetting and high-quality figure generation. The modular design of DO-MS facilitates customization and expansion. DO-MS is available for download from GitHub: github.com/SlavovLab/DO-MS

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SCoPE-MS: mass spectrometry of single mammalian cells quantifies proteome heterogeneity during cell differentiation

Cited by 701 publications

References 57 publications

Post‐ejaculatory modifications to sperm (PEMS)

Post‐ejaculatory modifications to sperm (PEMS)

Improved Sensitivity in Low-Input Proteomics using Micro-Pillar Array-based Chromatography

DO-MS: Data-Driven Optimization of Mass Spectrometry Methods

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