A hybrid spectral library combining DIA-MS data and a targeted virtual library substantially deepens the proteome coverage

Lou, Ronghui; Tang, Pan; Ding, Kang; Li, Shanshan; Tian, Changlin; Li, Yunxia; Zhao, Suwen; Zhang, Yaoyang; Wang, Shui

doi:10.1101/2020.01.16.909952

Cited by 8 publications

(13 citation statements)

References 25 publications

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“…2C). Although we previously demonstrated this strategy in processing data from a few brain regions as a proof of a concept (21), our current study proved that this hybrid spectral library can deepen the GPCR subproteome coverage to surpass a large-scale project-specific DDA library.…”

Section: Profiling the Transmembrane Proteome In Multiple Mouse Brain...mentioning

confidence: 63%

Multiregional profiling of the brain transmembrane proteome uncovers novel regulators of depression

Luo

Lou

et al. 2021

Sci. Adv.

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Transmembrane proteins play vital roles in mediating synaptic transmission, plasticity, and homeostasis in the brain. However, these proteins, especially the G protein–coupled receptors (GPCRs), are underrepresented in most large-scale proteomic surveys. Here, we present a new proteomic approach aided by deep learning models for comprehensive profiling of transmembrane protein families in multiple mouse brain regions. Our multiregional proteome profiling highlights the considerable discrepancy between messenger RNA and protein distribution, especially for region-enriched GPCRs, and predicts an endogenous GPCR interaction network in the brain. Furthermore, our new approach reveals the transmembrane proteome remodeling landscape in the brain of a mouse depression model, which led to the identification of two previously unknown GPCR regulators of depressive-like behaviors. Our study provides an enabling technology and rich data resource to expand the understanding of transmembrane proteome organization and dynamics in the brain and accelerate the discovery of potential therapeutic targets for depression treatment.

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Section: Profiling the Transmembrane Proteome In Multiple Mouse Brain...mentioning

confidence: 63%

Multiregional profiling of the brain transmembrane proteome uncovers novel regulators of depression

Luo

Lou

et al. 2021

Sci. Adv.

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“…First of all, we needed to determine KRT17 expression. DIA-MS is a proteomic methodology for deep and proteome-wide pro ling with high accuracy and reproducibility in proteomic quanti cation [22,23], we used DIA-MS to investigate the DEPs, and the result showed that the expression of KRT17 was up-regulated in ESCC tumor tissues, the differential expression was further determined by the data obtained from GEPIA. Also, the overall survival of ESSC patients was also closely related with KRT17 levels.…”

Section: Discussionmentioning

confidence: 99%

KRT17 Facilitates Proliferation, Migration and Invasion in Esophageal Squamous Cell Carcinoma by Regulating mTOR/S6K1 pathway

Liu

Shao

Wang

et al. 2021

Preprint

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BackgroundEsophageal squamous cell carcinoma (ESCC) is one of the most aggressive malignancies worldwide which originates from the malignant transformation of esophageal epithelial cells. Dysregulated expression of Keratin17 (KRT17) has been claimed in a variety of malignancies, while its role in ESCC remains unclear. Therefore, our study aimed to explore the potential function and underlying molecular mechanism of KRT17 in ESCC.MethodsData-independent acquisition-mass spectrometry (DIA-MS) workflow was used to analysis KRT17 expression between ESCC and adjacent non-cancerous esophageal tissues. The online database gene expression profiling interactive analysis (GEPIA) was used to further determine the differential expression of KRT17 in tissues. The function of KRT17 in ESCC was tested on two human esophageal cancer cell lines (EC9706 and ECA109). Small interfering RNA (siRNA) was used to inhibit KRT17 expression. Cell proliferation was examined by cell counting kit 8 (CCK8) reagent, colony formation assay, cell cycle distribution analysis and apoptosis. Cell migration was examined by transwell and wound healing assay. Cell invasion was also examined by transwell assay. Western blot and quantitative real-time PCR (qRT-PCR) was used to evaluate protein and mRNA levels, respectively.ResultsKRT17 expression was higher in cancer tissues compared with normal tissues. Transfected with siKRT17 attenuated protein and mRNA levels of KRT17, inhibited proliferation, migration and invasion, and decreased mTOR/S6K1 phosphorylation levels in EC9706 and ECA109.ConclusionKRT17 facilitates proliferation, migration and invasion in ESCC cells, and these cell viability functions were mediated by mTOR/S6K1 pathway.

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“…While this increased complexity makes the handling and analysis of DIA data more laborious, it has been demonstrated that the quanti cation by DIA is more robust compared to DDA. Recently, DIA has reached a protein coverage that is comparable to, or even exceeds, the one of DDA 2,3 .…”

Section: Introductionmentioning

confidence: 99%

In-Depth Benchmarking of DIA-type Proteomics Data Analysis Strategies Using a Large-Scale Benchmark Dataset Comprising Inter-Patient Heterogeneity

Fröhlich

Brombacher

Fahrner

et al. 2021

Preprint

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An overwhelming number of proteomics software tools and algorithms have been published for different steps of Data Independent Acquisition analysis of clinical samples. Nonetheless, there is still a lack of comprehensive benchmark studies evaluating which combinations of those isolated components perform best. Here, we used 92 lymph nodes from distinct patients to create a unique benchmark dataset representing real-world inter-individual heterogeneity. The publicly available dataset comprises 118 LC-MS/MS runs with > 12 million MS2 spectra and allowed us to objectively evaluate how well different combinations of spectral libraries, DIA software, sparsity reduction, normalization and statistical tests can detect differentially abundant proteins, while also taking sample size into account. Evaluation of 2 million data analysis workflows showed that a gas phase fractionation refined spectral library in combination with DIA-NN and Significance Analysis of Microarrays reliably detected differentially abundant proteins. Furthermore, DIA-NN and Spectronaut robustly avoided the false detection of truly absent proteins.

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A hybrid spectral library combining DIA-MS data and a targeted virtual library substantially deepens the proteome coverage

Cited by 8 publications

References 25 publications

Multiregional profiling of the brain transmembrane proteome uncovers novel regulators of depression

Multiregional profiling of the brain transmembrane proteome uncovers novel regulators of depression

KRT17 Facilitates Proliferation, Migration and Invasion in Esophageal Squamous Cell Carcinoma by Regulating mTOR/S6K1 pathway

In-Depth Benchmarking of DIA-type Proteomics Data Analysis Strategies Using a Large-Scale Benchmark Dataset Comprising Inter-Patient Heterogeneity

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