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Table of contents
INTRODUCTION
1.0: mRNPs
2.0: EJC
2.1: The EJC core components
2.1.1: eIF4A3
2.1.2: MAGOH‐Y14
2.1.3: MLN51
2.2: The tetrameric EJC core
2.3: EJC assembly
2.3.1: The pre‐EJC core
2.3.2: The recruitment of eIF4A3 by CWC22
2.4: EJC peripheral factors
2.4.1: Splicing‐related peripheral factors
2.4.2: Export factors
2.4.3: NMD related factors
2.4.4: Other EJC related factors
3.0: EJC life cycle
3.1: The localization of EJC core components
3.2: EJC remodeling and variability
3.2.1: EJC disassembly
3.2.2: Re‐cycling of EJCs to the nucleus
4.0: EJC functions
4.1: EJC modulates splicing
4.1.1: EJC dependent regulation of splicing in human cells
4.2: EJC enhances translation
4.2.1: EJC enhances translation in mTOR pathway
4.2.2: Translation enhancement by MLN51
4.3: EJC in quality control of mRNAs
4.3.1: The importance of NMD
4.3.2: NMD components
4.3.3: NMD Mechanism
4.3.4: Role of EJC in NMD
4.3.5: Role of EJC in translation control of natural NMD targets
4.4: EJC participates to mRNA export
4.4.1: General mRNA export adaptors and receptors
4.4.2: Role of EJC in mRNA transport
5.0: Global view of EJC deposition on mRNAs
5.1: Differential EJC loading on human transcriptome
5.2: The canonical and non‐canonical EJC
6.0: mRNA localization and local translation
6.1: mRNA localization
6.2: Importance of mRNA localization
6.2.1: Energy efficiency for the cell
6.2.2: Spatio‐temporal translation
6.2.3: Storage
6.2.4: One transcript, multi‐functional protein
6.3: Examples of localized mRNAs
6.4: Mechanism of mRNA localization
6.4.1: Directed transport of mRNA in cytoplasm: Cis‐regulatory elements and trans‐acting factors
6.4.2: Directed transport of mRNA in cytoplasm: Recruitment of molecular motors
6.4.3: Role of EJC in sub‐cellular localization
7.0: EJC in physiological contexts
7.1: EJC in development
7.2: EJC in mammalian brain development
7.3: EJC in human diseases
8.0: Development of mammalian Brain
8.1: Primary progenitors
8.2: Intermediate progenitors
8.3: Role of NSC in brain development
8.4: Ependymal cells
8.4.1: Physiological Functions of ependymal cells in brain
8.4.1.1: Regulation of neuronal niche
8.4.1.2: CSF maintenance
8.4.1.3: Metabolic protection of brain
8.4.1.4: Protection of brain from infections
8.4.1.5: Repair of brain after stroke
8.4.2: Ependymal differentiation in mammalian brain
8.4.3: Centriole amplification in ependymal differentiation
9.0: Cell‐cycle and cell‐quiescence
10.0: Centrosome
10.1: Centrosome Composition
10.2: Centrosome duplication in cycling cells
10.3: Centrosome in post‐mitotic cells
10.3.1: Formation of primary cilia
10.3.1.1: Cilia structure and functions
10.3.1.2: Primary cilia in neurodevelopmental disorders
10.3.2: Centrosome amplification in post‐mitotic cells
10.4: Centrosome functions
10.4.1: Centrosome functions in cell fate
10.4.2: Centrosomes in human diseases
11.0: Questions Asked
RESULTS (Part 1): Article
RESULTS (Part 2): Additional results
1.0: Identification of EJC‐bound mRNAs in NSC
1.1: EJC RIP‐seq strategy
1.2: Cytoplasmic fractionation of NSC
1.3: Tests of immuno‐depletion
1.4: EJC‐RIP and RNA purification
1.5: Analysis of sequencing results
1.6: Statistical validation of RIPseq
1.7: Identity of EJC bound transcripts in NSC
2.0: Visualization of expression of mRNAs by smFISH
2.1: smFISH model
2.2: Design and Synthesis of Fluorescent Oligonucleotide Probe Sets
2.2.1. Design
2.3: Results obtained
2.3.1: smFISH in cycling MEF
2.3.2: smFISH in quiescent NSC
2.3.3: Distribution of total mRNAs in quiescent NSC
2.3.4: smFISH in quiescent MEF
DISCUSSION AND PERSPECTIVES
1.0: Resume of our results
2.0: Presence of mRNAs at centrosome in variety of cell types
3.0: Accumulation of untranslated mRNAs at centrosome
3.1: Hypothesis 1: Translation dependent disassembly of EJCs
3.2: Hypothesis 2: Degradation or diffusion of EJC bound mRNAs
4.0: Functions of untranslated mRNAs at centrosome
MATERIALS AND METHODS
1.0: CELL CULTURE
1.1: Neural Stem Cell culture
1.2: Mouse Embryonic Fibroblast culture
2.0: Transfection of plasmids
3.0: Immunofluorescence and Microscopy
3.1: Image analysis.
4.0: PROTEIN ANALYSIS
4.1: Protein extraction
4.2: Immunoprecipitation
4.3: SDS‐PAGE
4.4: Western Blot Analysis
5: RNA analysis
5.1: Cellular fractionation
5.2: RNA Immuno Precipitation
5.3: RNA isolation
5.3.1: Isolation of immunoprecipitated RNA
5.3.1.1: RNA precipitation
5.3.2: Total RNA isolation
5.4: Quantitative RT‐PCR
6.0: smFISH analysis
6.1: Probe synthesis
6.2: Hybridization of probes with FlapY‐Cy3
6.3: Cell fixation
6.4: In situ hybridization
7.0: List of buffers
REFERENCES
