pr5b00480_si_002.pdf (4.88 MB)
Finding Missing Proteins from the Epigenetically Manipulated Human Cell with Stringent Quality Criteria
journal contribution
posted on 2015-09-04, 00:00 authored by Lijuan Yang, Xinlei Lian, Wanling Zhang, Jie Guo, Qing Wang, Yaxing Li, Yang Chen, Xingfeng Yin, Pengyuan Yang, Fei Lan, Qing-Yu He, Gong Zhang, Tong WangThe chromosome-centric human proteome
project (C-HPP) has made
great progress of finding protein evidence (PE) for missing proteins
(PE2–4 proteins defined by the neXtProt), which now becomes
an increasingly challenging field. As a majority of samples tested
in this field were from adult tissues/cells, the developmental stage
specific or relevant proteins could be missed due to biological source
availability. We posit that epigenetic interventions may help to partially
bypass such a limitation by stimulating the expression of the “silenced”
genes in adult cells, leading to the increased chance of finding missing
proteins. In this study, we established in vitro human
cell models to modify the histone acetylation, demethylation, and
methylation with near physiological conditions. With mRNA-seq analysis,
we found that histone modifications resulted in overall increases
of expressed genes in an even distribution manner across different
chromosomes. We identified 64 PE2–4 and six PE5 proteins by
MaxQuant (FDR < 1% at both protein and peptide levels) and 44 PE2–4
and 7 PE5 proteins by Mascot (FDR < 1% at peptide level) searches,
respectively. However, only 24 PE2–4 and five PE5 proteins
in Mascot, and 12 PE2–4 and one PE5 proteins in MaxQuant searches
could, respectively, pass our stringently manual spectrum inspections.
Collectively, 27 PE2–4 and five PE5 proteins were identified
from the epigenetically modified cells; among them, 19 PE2–4
and three PE5 proteins passed FDR < 1% at both peptide and protein
levels. Gene ontology analyses revealed that the PE2–4 proteins
were significantly involved in development and spermatogenesis, although
their chemical–physical features had no statistical difference
from the background. In addition, we presented an example of suspicious
PE5 peptide spectrum matched with unusual AA substitutions related
to post-translational modification. In conclusion, the epigenetically
manipulated cell models should be a useful tool for finding missing
proteins in C-HPP. The mass spectrometry data have been deposited
to the iProx database (accession number: IPX00020200).