Abstract
Alternative splicing (AS) may increase the number of proteoforms produced by a gene. Alzheimer’s disease (AD) is a neurodegenerative disease with well-characterized AS proteoforms. In this study, we used a proteogenomics strategy to build a customized protein sequence database and identify orthologous AS proteoforms between humans and mice on publicly available shotgun proteomics (MS/MS) data of the corpus callosum (CC) and olfactory bulb (OB). Identical proteotypic peptides of six orthologous AS proteoforms were found in both species: PKM1 (gene PKM/Pkm), STXBP1a (gene STXBP1/Stxbp1), Isoform 3 (gene HNRNPK/Hnrnpk), LCRMP-1 (gene CRMP1/Crmp1), SP3 (gene CADM1/Cadm1), and PKCβII (gene PRKCB/Prkcb). These AS variants were also detected at the transcript level by publicly available RNA-Seq data and experimentally validated by RT-qPCR. Additionally, PKM1 and STXBP1a were detected at higher abundances in a publicly available MS/MS dataset of the AD mouse model APP/PS1 than its wild type. These data corroborate other reports, which suggest that PKM1 and STXBP1a AS proteoforms might play a role in amyloid-like aggregate formation. To the best of our knowledge, this report is the first to describe PKM1 and STXBP1a overexpression in the OB of an AD mouse model. We hope that our strategy may be of use in future human neurodegenerative studies using mouse models.
Highlights
Proteogenomics is an emerging field that integrates proteomic, transcriptomic, and genomic data to assess the multiple regulatory levels of gene expression within the cell [1].The identification of peptide sequences through tandem mass spectrometry (MS/MS) is often achieved by matching experimental and theoretical mass spectra using a standard protein sequence database [2]
Our analysis identified a proteotypic peptide of the proteoform LCRMP-1 in humans (UniProt ID: Q14194-2) and mice (Ensemble ID: ENSMUST00000114158) in mice on publicly available shotgun proteomics (MS/MS) data of the olfactory bulb (OB)
To the best of our knowledge, this is the first report of PKM1 overexpression in the OB of an Alzheimer’s disease (AD)
Summary
Proteogenomics is an emerging field that integrates proteomic, transcriptomic, and genomic data to assess the multiple regulatory levels of gene expression within the cell [1].The identification of peptide sequences through tandem mass spectrometry (MS/MS) is often achieved by matching experimental and theoretical mass spectra using a standard protein sequence database [2]. Integrating data from multiple levels, such as the genome and transcriptome, permits the construction of customized protein sequence databases that can improve peptide identification in unassigned mass spectra [4]. Most of these unassigned mass spectra correspond to proteoforms derived from molecular events, such as post-translation modifications (PTMs) and alternative splicing (AS), which are usually absent in standard protein sequence databases [5]. 95% of human pre-mRNAs are alternatively spliced [8], with the resulting mRNA variants regulated in response to tissue- and developmental stage-specific factors [8] When translated, these AS variants can generate distinct proteoforms, increasing proteome diversity [9]. The human brain contains a large diversity of AS variants, which can be detected at the transcript [10] and protein [11]
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