Dataset Information

A mouse SWATH-MS reference spectral library enables deconvolution of species-specific proteomic alterations in human tumour xenografts

ABSTRACT: SWATH-MS enables accurate and reproducible label-free quantification of proteomes including in extensively studied model organisms such as the mouse. Here we present a comprehensive mouse reference spectral library (MouseRefSWATH) that allows quantification of up to 10,597 proteins (62.2% of mouse proteome) by SWATH-MS. We exploit MouseRefSWATH to develop an analytical pipeline for species-specific deconvolution of proteomic alterations in human tumour xenografts (XenoSWATH). This method overcomes the challenge of high protein sequence similarity between the mouse and human, facilitating the study of host microenvironment-tumour interactions from ‘bulk tumour’ measurements. We apply XenoSWATH to characterise a xenograft model of breast ductal carcinoma in-situ and uncover complex regulation of cell migration pathways that is not restricted to the tumour cells but also operates in the mouse stromal upon progression to invasive disease. MouseRefSWATH and XenoSWATH opens new possibilities for in-depth proteomic assessment to address wide-ranging biological questions involving this important model organism.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Mammary Gland Tumor Cell, Epithelial Cell, Mammary Gland

DISEASE(S): Breast Cancer

SUBMITTER: Lukas Krasny

LAB HEAD: Paul Huang

PROVIDER: PXD017209 | Pride | 2020-06-23

REPOSITORIES: Pride

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Publications

A mouse SWATH-mass spectrometry reference spectral library enables deconvolution of species-specific proteomic alterations in human tumour xenografts.

Krasny Lukas L Bland Philip P Burns Jessica J Lima Nadia Carvalho NC Harrison Peter T PT Pacini Laura L Elms Mark L ML Ning Jian J Martinez Victor Garcia VG Yu Yi-Ru YR Acton Sophie E SE Ho Ping-Chih PC Calvo Fernando F Swain Amanda A Howard Beatrice A BA Natrajan Rachael C RC Huang Paul H PH

Disease models & mechanisms 20200714 7

SWATH-mass spectrometry (MS) enables accurate and reproducible proteomic profiling in multiple model organisms including the mouse. Here, we present a comprehensive mouse reference spectral library (MouseRefSWATH) that permits quantification of up to 10,597 proteins (62.2% of the mouse proteome) by SWATH-MS. We exploit MouseRefSWATH to develop an analytical pipeline for species-specific deconvolution of proteomic alterations in human tumour xenografts (XenoSWATH). This method overcomes the chall ...[more]

PMID: 32493768

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Project description:ErbB-2 overexpression and amplification occurs in 15 - 30% of human invasive breast carcinomas associated with poor clinical prognosis. Previously, we have demonstrated that four ErbB-2/Neu tyrosine-autophosphorylation sites within the cytoplasmic tail of the receptor recruit distinct adaptor proteins and are sufficient to mediate transforming signals in vitro. Two of these sites representing the Grb2 (Neu-YB) and Shc (Neu-YD) binding sites can induce mammary tumourigenesis and metastasis. Here we show that Neu-YC and Neu-YE transgenic mice develop metastatic mammary tumours. A detailed comparison of pathological and transcriptional profiles among all Neu mutant mouse models revealed that Neu-YC, -YD and -YE mammary tumours shared similar pathological and transcriptional features correlating with their capacity to signal through a common adaptor like Shc. In contrast, the Neu-YB mouse model displayed a unique pathology with a high metastatic potential that correlates with a distinct transcriptional profile. We identified genes specifically expressed in YB-induced mammary tumours, including CXCL12/SDF-1Î± that promotes malignant tumour progression. Furthermore, Neu-YB tumour epithelial cells showed abundant intracellular CXCL12/SDF-1Î± protein, which may reflect the more aggressive phenotype among all Neu mutant mouse models. These findings indicate that activation of distinct Neu-coupled signalling pathways has a deep impact on the biological behaviour of Neu-induced tumours. Experiment Overall Design: Total RNA was isolated from 10 individual flash frozen mammary tumour samples derived from our MMTV/neundl-NYPD, -YB, -YC, -YD, -YE and the parental MMTV/neu NDL2-5 strains. Two RNA pools, containing equal amount of total RNA from five individual tumours, were generated from each strain and functioned as a biological repeat. Five hundred nanograms of total RNA from each pool was subjected to one round of T7 linear amplification using the Amino Allyl MessageAmpTM aRNA Kit (Ambion, Austin, Texas). Ten micrograms of the resulting aRNA was labelled with Cy3 and Cy5 dyes. Each of the 12 Samples represents a dyeswap pair.

Dataset Information

A mouse SWATH-MS reference spectral library enables deconvolution of species-specific proteomic alterations in human tumour xenografts

Publications

A mouse SWATH-mass spectrometry reference spectral library enables deconvolution of species-specific proteomic alterations in human tumour xenografts.

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