CEEPC/IPM/CMSC - Abstrakt prezentace

(CEEPC/IPM/CMSC 2022 - FrP-22)
Comparison of SWATH-MS data analysis pipelines on proteomic data from human Huntington’s disease iPSC and NSC lines

Kateřina Vodičková Kepková 1, Jakub Červenka 1,2, Jiřina Tylečková 1, Jaromír Novák 1,2, Kateřina Budková 1,2, Petr Vodička 1 *

  1. Institute of Animal Physiology and Genetics of the Czech Academy of Sciences, Libechov, CZ
  2. Charles University, Faculty of Science, Department of Cell Biology, Prague, CZ

Abstrakt

Data-independent acquisition (DIA) is a robust, reproducible, high throughput quantitative MS approach that enables relative quantification of thousands of proteins in one sample. Several software tools and data analysis pipelines were developed over time to process DIA data. We used a specific variant of DIA known as SWATH-MS for characterization of induced pluripotent stem cells (iPSC) and neural stem cells (NSC) derived from Huntington’s disease (HD) patients and healthy controls (WT).
Using TTOF 5600, we analysed at quantifiable level over 3000 proteins. We then processed the MS data using two different data pipelines. Pipeline #1 consisted of protein identification for spectral library creation using Mascot, followed by quantification using sample specific spectral library in Skyline/MSstats. Pipeline #2 consisted of identification and quantification in MaxDIA, using in silico generated human spectral library. Both approaches identified more changes between WT and HD lines at NSC stage, compared to iPSC stage. Using Pipeline #1, we quantified 3142 proteins and identified 129 differently abundant proteins, while using Pipeline #2, we quantified 3397 proteins, of which 91 were identified as differently abundant. 2125 proteins were common in both approaches, but only 11 proteins were identified as significantly changed by both pipelines.
This shows that different decisions at various stages of data analysis, from peptide identification to quantification of protein families or using only proteotypic peptides for quantification, can to large extent influence overall results and care should be taken in data verification and interpretation.

* Korespondující autor: vodicka@iapg.cas.cz

Poděkování:

This study was supported by the Czech Science Foundation (22-24983S) and by the Czech Ministry of Education, Youth and Sports project InterCOST (LTC18079) under CellFit COST Action (CA16119).


Partneři společnosti

LabRules LCMS LabRules GCMS

Partneři

Bruker HPST Merck Pragolab Amedis EastPort Shimadzu Waters