ABSTRACT
Controlling translation initiation is an efficient way to regulate gene expression at the post-transcriptional level. However, current knowledge regarding regulatory proteins and their modes of controlling translation initiation is still limited. In this study, we employed tandem affinity purification and mass spectrometry to screen for unknown proteins associated with the translation initiation machinery. Ubiquitin specific peptidase 9, X-linked (USP9X), was identified as a novel binding partner, that interacts with the eukaryotic translation initiation factor 4B (eIF4B) in a mRNA-independent manner. USP9X-deficient cells presented significantly impaired nascent protein synthesis, cap-dependent translation initiation and cellular proliferation. USP9X can selectively alter the translation of pro-oncogenic mRNAs, such as c-Myc and XIAP. Moreover, we found that eIF4A1, which is primarily ubiquitinated at Lys-369, is the substrate of USP9X. USP9X dysfunction increases the ubiquitination of eIF4A1 and enhances its degradation. Our results provide evidence that USP9X is a novel regulator of the translation initiation process via deubiquitination of eIF4A1, which offers new insight in understanding the pivotal role of USP9X in human malignancies and neurodevelopmental disorders.
Subject(s)
Eukaryotic Initiation Factor-4A/metabolism , Protein Biosynthesis , Ubiquitin Thiolesterase/metabolism , Ubiquitination , Cell Proliferation/genetics , Eukaryotic Initiation Factor-4A/genetics , Eukaryotic Initiation Factors/genetics , Eukaryotic Initiation Factors/metabolism , HEK293 Cells , HeLa Cells , Humans , MCF-7 Cells , Mutation , Protein Binding , RNA Interference , RNA, Messenger/genetics , RNA, Messenger/metabolism , Substrate Specificity , Ubiquitin Thiolesterase/geneticsABSTRACT
Aging is a natural phenomenon that affects the entire physiology of an organism. Elucidating the molecular mechanisms underlying this complex process remains a major challenge today. Humans make poor models for research into aging because of their long life span. Thus, most of the current knowledge is through studies conducted in lower organisms. Large differences in life spans make it difficult to extrapolate the results of experiments carried out in model organisms to humans. Recent advances in genomic and proteomic technologies now permit generation of data pertaining to aging on a large-scale. In addition, several web-based community resources and databases are available that provide easy access to the available data. Use of bioinformatics and systems biology type of approaches provide a framework to start dissecting this complex biological phenomenon. Here, we discuss various genomic, transcriptomic and proteomic approaches that have the potential to provide a comprehensive mechanistic insight into the aging process.
Subject(s)
Aging , Computational Biology , Proteomics , Research , Animals , Humans , RNA, Messenger/genetics , Systems BiologySubject(s)
Chromosomes, Human, X/genetics , Alternative Splicing , Amino Acid Sequence , Base Sequence , Chromosome Mapping , Computational Biology , DNA, Complementary/genetics , Databases, Genetic , Databases, Protein , Exons , Humans , Molecular Sequence Data , Open Reading Frames/physiology , Sequence AlignmentABSTRACT
Mass spectrometry is already established as the method of choice for protein identification. Hitherto, a major limitation of mass spectrometry was the inability to quantitate protein levels. With the development of novel methods, such as the AQUA method by Gerber et al., this hurdle has finally been overcome, unleashing the power of mass spectrometry to study protein dynamics in cells.
