کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
5513561 | 1541213 | 2017 | 11 صفحه PDF | دانلود رایگان |
- Phosphorylation is a critical regulator of noncanonical activities of tRNA synthetases.
- Framework of approaches for identification of signaling pathways and phosphorylation events in tRNA synthetases.
- Isotope- and non-isotope-based approaches for detection of phosphorylation events.
- Biochemical and genetic approaches for identification of phospho-sites, signaling mechanisms, proximal kinases, and functional relevance.
Phosphorylation of many aminoacyl tRNA synthetases (AARSs) has been recognized for decades, but the contribution of post-translational modification to their primary role in tRNA charging and decryption of genetic code remains unclear. In contrast, phosphorylation is essential for performance of diverse noncanonical functions of AARSs unrelated to protein synthesis. Phosphorylation of glutamyl-prolyl tRNA synthetase (EPRS) has been investigated extensively in our laboratory for more than a decade, and has served as an archetype for studies of other AARSs. EPRS is a constituent of the IFN-γ-activated inhibitor of translation (GAIT) complex that directs transcript-selective translational control in myeloid cells. Stimulus-dependent phosphorylation of EPRS is essential for its release from the parental multi-aminoacyl tRNA synthetase complex (MSC), for binding to other GAIT complex proteins, and for regulating the binding to target mRNAs. Importantly, phosphorylation is the common driving force for the context- and stimulus-dependent release, and non-canonical activity, of other AARSs residing in the MSC, for example, lysyl tRNA synthetase (KARS). Here, we describe the concepts and experimental methodologies we have used to investigate the influence of phosphorylation on the structure and function of EPRS. We suggest that application of these approaches will help to identify new functional phosphorylation event(s) in other AARSs and elucidate their possible roles in noncanonical activities.
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Journal: Methods - Volume 113, 15 January 2017, Pages 72-82