Top-Down Proteomics Enables Comparative Analysis of Brain Proteoforms Between Mouse Strains

Over
the past decade, advances in mass spectrometry-based proteomics
have accelerated brain proteome research aimed at studying the expression,
dynamic modification, interaction and function of proteins in the
nervous system that are associated with physiological and behavioral
processes. With the latest hardware and software improvements in top-down
mass spectrometry, the technology has expanded from mere protein profiling
to high-throughput identification and quantification of intact proteoforms.
Murine systems are broadly used as models to study human diseases.
Neuroscientists specifically study the mouse brain from inbred strains
to help understand how strain-specific genotype and phenotype affect
development, functioning, and disease progression. This work describes
the first application of label-free quantitative top-down proteomics
to the analysis of the mouse brain proteome. Operating in discovery
mode, we determined physiochemical differences in brain tissue from
four healthy inbred strains, C57BL/6J, DBA/2J, FVB/NJ, and BALB/cByJ,
after probing their intact proteome in the 3.5–30 kDa mass
range. We also disseminate these findings using a new tool for top-down
proteomics, TDViewer and cataloged them in a newly established Mouse
Brain Proteoform Atlas. The analysis of brain tissues from the four
strains identified 131 gene products leading to the full characterization
of 343 of the 593 proteoforms identified. Within the results, singly
and doubly phosphorylated ARPP-21 proteoforms, known to inhibit calmodulin,
were differentially expressed across the four strains. Gene ontology
(GO) analysis for detected differentially expressed proteoforms also
helps to illuminate the similarities and dissimilarities in phenotypes
among these inbred strains.