Dissertation > Excellent graduate degree dissertation topics show

Dissection of the Regulatory Mechanism of the Ubiquitin-activating Enzyme Uba1in Autophagy Regulation

Author: ZhangShaoJin
Tutor: ZuoXiangHua; YuLi
School: Huazhong Agricultural University
Course: Animal Nutrition and Feed Science
Keywords: Autophagy The ubiquitin-activating enzyme Uba1 The target of rapamycincomplex1 Atg1 Atg13
CLC: S852.2
Type: Master's thesis
Year: 2012
Downloads: 144
Quote: 0
Read: Download Dissertation

Abstract


Autophagy is an evolutionarily conserved process by which portions of cytosol and organelles are sequestered into double-membrane vesicles and degraded upon fusion with lysosomal compartments. As autophagy is involved in cell growth, survival, development and death, the levels of autophagy must be properly regulated, as indicated by the fact that dysregulated autophagy has been linked to many human pathophysiologies, such as cancer, myopathies, neurodegeneration, heart and liver diseases, and gastrointestinal disorders. Therapeutic exploitation of this process requires a clear understanding of its regulation. Although substantial progress has recently been made in understanding the molecular mechanisms of the autophagy machinery, there is limited information on how cellular signaling pathways regulate this complex process and many unanswered questions remain in the regulation of autophagy.To figure out new regulators of autophagy, we screened a yeast knockout library (covering100%of the yeast nonessential genes) and a conditional temperature-sensitive mutants collection (covering~65%of the yeast essential genes) for strains that are deficient in autophagy using the GFP-Atg8assay, and identity the ubiquitin-activating enzyme Ubal temperature-sensitive mutant (ubal-ol) cells with a block in autophagy. Laser scanning confocal microscope (LSCM) and western blot analysis showed that the ubal-ol cells had severe defects in the nitrogen or glucose starvation induced autophagy, which can be rescued by a plasmid-borne wild type UBA1. The fact that the ubal-ol cells showed normal autophagy activity during rapamycin treatment indicated that Ubal probably act through the upstream of TORC1to regulate autophagy. Therefore we move on to detect the TORC1activity in the Ubal wild type (WT) and ubal-ol cells during starvation or rapamycin treatment, and found that ubal-ol delayed the nitrogen starvation induced TORC1inactivation but not the glucose starvation or rapamycin treatment induced TORC1inactivation. Furthermore, ubal-ol doesn’t affect TORCl’s reactivation after its inactivation, suggesting that Ubal probably act on some upstream negative regulator of TORC1or directly affect the TORC1activity to regulate the nitrogen starvation induced autophagy. Notably, TORC1was quickly inactivated during the glucose starvation in the uba1-o1cells but it still showed a severe block in autophagy, which means that Ubal might also act through the downstream of TORC1to regulate the glucose starvation induced autophagy. So we tried to check whether the Atg1complex could be recruited to the pre-autophagosomal structure (PAS) normally in the uba1-o1cells. The results showed that the Atg1complex in the uba1-o1cells could not be properly recruited to the PAS during the nitrogen or glucose starvation, but not that case during the rapamycin treatment. Together with the TORC1activity results, we hypothesized that the uba1-o1cells had defects in some step between TORC1inactivation and the Atg1complex being recruited to the PAS during the glucose starvation. Then we analyzed the phosphorylation states of Atg13and its interaction with Atg1in the uba1-o1cells during the nitrogen or glucose starvation, and found that Atg13were hyperphosphorylated and there were no interaction between Atg1and Atg13in both cases. Therefore, Ubal probably act on the phosphates responsible for Atg13dephosphorylation to regulate the glucose starvation induced autophagy.Overall, we successfully identified Uba1as a new regulator of autophagy and dissected the probable step at which Uba1regulated autophagy during the nitrogen or glucose starvation respectively. However, the direct targets through which Uba1regulates autophagy needs further study.

Related Dissertations

  1. The Effects of the Mesenchymal Stem Cells and Osteoblasts on the Biological Behavior of Multiple Myeloma Cells in Vitro and the Mechanisms Involved,R733.3
  2. Inhibition of NF-κB Contribute to Autophagy and Affect Cell Death in Intracerebral Hemorrhage Mice,R743.3
  3. Paeoniflorin Protects PC12 Cells from MPP~ + -induced Injury Via Autophagy-lysosome Pathway,R285.5
  4. Effect of Different Glucose Concentrations on Autophagy in Neonatal Rat Cardiomyocyte Culture,R542.2
  5. Apoptosis Mechanisms Research in Interfering TIGAR Gene and Epirubicin Treatmnet in Hepatic Carcinoma Cells,R735.7
  6. The Effect of Gefitinib in Radiosensitization Through Autophagy Regulation in Human Non-small-cell Lung Cancer A549 Cell Line,R734.2
  7. Effect of Chronic Intermittent Hypoxia on the Level of Autophagy in Mice Hippocampal CA1 Neurons,R766.43
  8. The Effect and Mechanism of Autophagy in Inhibiting the Proliferation of A549 Cells Induced by Rh-endostatin and Erlotinib,R96
  9. Role of Autophagy in Sevoflurane Preconditioning Mediated Delayed Myocardial Protection on Rat Cardiomyocytes Against Hypoxia/Reoxgenation Injury,R965
  10. Amiloride Is Protective in PC12 Cells Via Autophagy Pathway,R329
  11. The Effects of ER Stress on Cell Toxicity and Htt Levels in the Stable PC12 Cells Expressing Htt586,R363
  12. Intervention and Mechanisms of E-64d on Developmental Seizure-induced Cognitive Deficit,R720.597
  13. Effects of Lysosomal Enzyme Inhibitor on Recurrent Neonatal Seizure Induced Acute and Long-term Gene Expression and Behavioral Changes,R720.597
  14. Study on the Autophagy Related Gene after Focal Cerebral Ischemia/reperfusion and the Effects of Artificial Synthetic E-selectin in Rats,R651.1
  15. Experimental Study the Role of Autophagy in Early Brain Injury after Subarachnoid Hemorrhage in Rats,R743.35
  16. Study on the Autophagy Level of Organs in the Adult Exercising Mice,R87
  17. The Effects of Autophagy on Vascular Smooth Muscle Cells Proliferation,R363
  18. The Mechanisms of Autophagy Affecting Human Umbilical Vein Endothelial Cells Proliferation and Angiogenesis,R543.6
  19. Experimental Study of Proliferation of EPCs Effected by 3-MA in Rat’s Chronic Deep Vein Thrombosis,R543.6
  20. Effects of siRNA Inhibit Class I PI3K on Humangastric Cancer SGC7901 Cells,R735.2
  21. Effects of Combined Therapy of 5-FU and Inhibitor of ClassⅢ PI3K Human Gastric Cancer SGC7901 Cells,R735.2

CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Basic Veterinary Science > Animal physiology, biophysics,biochemistry
© 2012 www.DissertationTopic.Net  Mobile