Bioinformatic Studies Of Protein Post-translational Modifications In The Regulation Of Mitosis And Autophagy

Post-translational modifications (PTMs) are covalent modifications of protein after its translation by attaching other biochemical functional groups to specific amino acid residues. Post-translational modifications can change conformation and activity of protein and extend protein's functions, and play an important role in the regulation of various biological processes, such as transcriptional regulation, signal transduction, cell cycle and cell death. As two important biological processes in eukaryotic cells, although with distinct biological functions, mitosis and autophagy share many similarities and relevance in both processes and functions. Mitosis and autophagy involve the participation of many proteins, and it has been demonstrated that post-translational modifications have critical roles in regulating mitosis and autophagy. Numerous experimental studies have identified a large number of proteins involved in these two processes; the rapid development of high-throughput proteomics promoted the studies of post-translational modifications and produced large amounts of data. How to collect, curate, analyze these data and extract useful information for further experimental research has become an urgent problem. In this work, the proteins and post-translational modifications in mitosis and autophagy were systematically studied.Mitosis is the basis of cell proliferation, organism normal growth and development and the transfer of genetic material. In eukaryotic cells, a large number of proteins spatially and temporally localize at distinct subcellular positions and organize various complexes including midbody, centrosome, kinetochore, spindle and telomere to orchestrate the chromosome segregation during mitosis. Firstly,1,872 experimentally identified proteins, which at least locate in one of the five positions in eight model species, were curated from the scientific literature manually. Furthermore, the ortholog detection was performed to identify potential proteins from 144 eukaryotic organisms. The primary references and post-translational modifications information were also integrated. Finally, a comprehensive database MiCroKiTS 4.0 was developed containing 87,983 unique proteins.Autophagy is a highly conserved process for degrading cytoplasmic contents, determines cell survival or death, and regulates the cellular homeostasis. Besides core ATG proteins, a large number of regulators together with various post-translational modifications are also involved in autophagy. Considering that autophagy is one of the major routes of cell death, proteins regulated in autophagy, apoptosis and necrosis were simultaneously collected from the literature in this work, and 4,241 experimentally identified proteins were obtained. The statistical enrichment analyses revealed that autophagy is most significantly associated with human diseases, whereas known cancer genes and drug targets were dramatically over-represented in human autophagy and cell death pathways. By re-constructing a human kinase-substrate phosphorylation network for autophagy, apoptosis and necrosis, mostly regulated kinases and substrates were identified, and the results are highly consistent with previous experiments. We further mapped 54,469 known sites of 11 types of post-translational modifications to curated proteins, and revealed that multiple types of post-translational modifications substrates were significantly enriched in autophagy and apoptosis. In addition, protein kinase, phosphatase and ubiquitin-protein ligases were enriched in autophagy and cell death pathway proteins. These data suggest that post-translational modifications are important in the regulation of autophagy and cell death pathway.Based on the above results, we constructed an integrative database of The Autophagy, Necrosis, ApopTosis OrchestratorS (THANATOS). The potential orthologs of known autophagy, apoptosis and necrosis proteins were computationally detected across 118 eukaryotes including 47 animals,23 plants and 48 fungi respectively, and three orthologs in zebrafish, Atg3, Keap1b and Skp2, were experimentally verified to be involved in regulating autophagy during the zebrafish embryonic development. Finally, there are 144,153 proteins associated with autophagy, apoptosis and necrosis in THANATOS 1.0 database. The known post-translational modifications, protein-protein interaction, primary references and other annotations were also integrated.Taken together, we systematically analyzed the proteins and post-translational modifications in mitosis and autophagy which are two important biological processes with similarities and relevance. Our study would serve as a useful resource for further analyzing the molecular mechanisms and the regulation of post-translational modifications during cell division and autophagy/cell death.