RNA maturation transport localization RNA processing brief overview
RNA maturation transport & localization RNA processing: brief overview RNA export to the cytoplasm: model systems RNA degradation Links between RNA processing, transport, degradation RNA localization in the cytoplasm
All eukaryotic m. RNAs are processed Evidence for checkpoints? m. RNA transport
1. Capping Specific process for Pol II transcripts nuclear CBC binds to m 7 G-cap: role in splicing, transport, stability (exchanged in cytoplasm) Function in transport, stability, translation
2. Polyadenylation CPSF: cleavage and polyadenylation specificity factor CSt. F: cleavage stimulatory factor CF: cleavage factor PAP: poly. A polymerase PAB: poly. A binding protein Function in transport, stability, translation
3. Splicing
3. Splicing
Splicing is mediated by sn. RNPs
Compartments inside the nucleus 1. Nucleolus
2. Perichromatin granule clusters Carter et al. , Science (1993) 259: 1330
Balbiani Rings (Chironomus tentans)
BR RNP maturation
BR RNP translocation
BR RNP translocation
Retroviruses as m. RNA export mode
Gene location is influenced by gene ac Casolari et al. , 200
SUMMARY I. Multiple steps of m. RNA biosynthesis are tightly coupled II. Mex 67/TAP is one major m. RNA export factor Binding to m. RNA may already occur at the site of transcription III. Many questions remain - How are m. RNA substrates released in the cytoplasm? - Are there multiple m. RNA export pathways? - How is processing and transport mechanistically “coupled”? - Is m. RNA export regulated? etc.
RNA half-lives vary greatly but are highly coordinated
RNA half-lives vary greatly but are highly coordinated Wang et al. , PNAS 2002
AUUUA element regulates half-life
Examples of regulated m. RNA turnover A. + B. Casein m. RNA 30, 000 m. RNA/cell No change in transcription prolactin - 300 m. RNA/cell
m. RNA DEGRADATION m. RNA DECAY NMD ‘turnover’ ‘surveillance’
m. RNA DECAY m 7 Gppp poly A shortening m 7 Gppp Decapitation AAAAAAAA Deadenylase complex AAA Decapping enzyme (DCP 1 complex) AAA 5’-3’ exonucleolytic cleavage Xrn 1 complex
Decay factors localize to cytoplasmic processing bodies (P bodies) Sheth et al. Science 2003
m. RNA activity is regulated by multiple factors Active m. RNAs Inactive m. RNAs Storage Translation Decay Transport
Position of nonsense codon affects m. RNA amounts From Neu-Yilik et al. (2001) EMBO 20: 532 -540
Nonsense Mediated Decay Stop in penultimate exon/ 5’ of splicing mark m 7 Gppp Decapitation AAAAAAAA Decapping enzyme (DCP 1 complex) AAAAAAAA 5’-3’ exonucleolytic cleavage Xrn 1 complex
RNA localization m. RNA can be localized to subcellular compartments by actin or tubulin-dependent processes Examples: Xenopus: Vg 1 m. RNA (TGFb) to vegetal pole Drosophila: nanos, oskar m. RNA (posterior) and bicoid (anterior) (requires m. RNA binding protein staufen) prospero (into ganglion of mother cells; neuroblast TF) (requires staufen and miranda) Yeast: Ash 1 m. RNA to daughter cell
Examples of localized m. RNAs in various systems
3’ UTR determins localization of many m. RNAs lamellipodia staining perinuclear staining in myotubes
Ash 1 m. RNA specifically localizes to new daughter cells Bertrand et al. , Mol Cell (98) 2: 437 -445
Mechanism of Ash 1 m. RNA localization
SUMMARY I. m. RNA decay - regulated and non-regulated turn-over but apparently coordinated - ordered pathways (e. g. deadenylation, decapping, exonucleolytic degradation) - cross-talk between translation and turnover - important regulation via non-coding RNAs - turnover occurs in specific cytoplasmic compartments - NMD: recognition of premature stop codons II. Cytoplasmic m. RNA localization - ZIP code in 3’ UTR - both actin and tubulin-mediated - yeast mating type switch as a model: Ash 1 m. RNA localization (via 3’ UTR, She 2/3, Myo 4 and actin cables)
- Slides: 33