A Proteomics Strategy for the Identification of FAT10-Modified Sites by Mass Spectrometry

The ubiquitin-like protein FAT10 (HLA-F adjacent transcript 10) is uniquely expressed in mammals. The fat10 gene is encoded in the MHC class I locus in the human genome and is related to some specific processes, such as apoptosis, immune response, and cancer. However, biological knowledge of FAT10 i...

Full description

Saved in:
Bibliographic Details
Published in:Journal of proteome research 2014-01, Vol.13 (1), p.268-276
Main Authors: Leng, Ling, Xu, Changming, Wei, Chao, Zhang, Jiyang, Liu, Boya, Ma, Jie, Li, Ning, Qin, Weijie, Zhang, Wanjun, Zhang, Chengpu, Xing, Xiaohua, Zhai, Linhui, Yang, Fan, Li, Mansheng, Jin, Chaozhi, Yuan, Yanzhi, Xu, Ping, Qin, Jun, Xie, Hongwei, He, Fuchu, Wang, Jian
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Chromatography, Affinity
HeLa Cells
Humans
Mass Spectrometry - methods
Molecular Sequence Data
Proteomics
Ubiquitins - chemistry
Ubiquitins - genetics
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:The ubiquitin-like protein FAT10 (HLA-F adjacent transcript 10) is uniquely expressed in mammals. The fat10 gene is encoded in the MHC class I locus in the human genome and is related to some specific processes, such as apoptosis, immune response, and cancer. However, biological knowledge of FAT10 is limited, owing to the lack of identification of its conjugates. FAT10 covalently modifies proteins in eukaryotes, but only a few substrates of FAT10 have been reported until now, and no FATylated sites have been identified. Here, we report the proteome-scale identification of FATylated proteins by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). We identified 175 proteins with high confidence as FATylated candidates. A total of 13 modified sites were identified for the first time by a modified search of the raw MS data. The modified sites were highly enriched with hydrophilic amino acids. Furthermore, the FATylation processes of hnRNP C2, PCNA, and PDIA3 were verified by a coimmunoprecipitation assay. We confirmed that most of the substrates were covalently attached to a FAT10 monomer. The functional distribution of the FAT10 targets suggests that FAT10 participates in various biological processes, such as translation, protein folding, RNA processing, and macromolecular complex assembly. These results should be very useful for investigating the biological functions of FAT10.
ISSN:1535-3893
1535-3907
DOI:10.1021/pr400395k