Results: 5

1.
FIG. 2

FIG. 2. From: Cytoplasmic Polyadenylation in Development and Beyond.

Model for CPE-mediated translational repression and activation. In immature oocytes, CPEB binds both the CPE and a hypothetical factor, factor X. Factor X, in turn, prevents translation either by interacting with the cap or by preventing eukaryotic initiation factors (i.e., eIF-4E) from recognizing the cap. A possible hexanucleotide binding factor (HNBF), which could be CPSF, is also indicated. Following oocyte maturation, CPEB induces cytoplasmic polyadenylation, which disrupts CPEB-factor X interaction and allows initiation factor binding to the cap and translation initiation.

Joel D. Richter. Microbiol Mol Biol Rev. 1999 June;63(2):446-456.
2.
FIG. 3

FIG. 3. From: Cytoplasmic Polyadenylation in Development and Beyond.

Essential features of poly(A) addition and removal during Xenopus development. In oocytes, CPEB binds the CPE and shifts an equilibrium between poly(A) tail growth or removal in the direction of growth. For mRNAs that do not contain a CPE, the equilibrium is shifted toward poly(A) tail removal, which is often referred to as default deadenylation. The enzyme responsible for deadenylation is PARN. HNBF refers to hexanucleotide binding factor. During embryogenesis, poly(A) tail elongation is directed by an embryonic-type CPE (eCPE), which is oligo(U)12–27. The eCPE is bond by the protein elrA, a member of the ELAV family of RNA binding proteins. In contrast to oocyte maturation, deadenylation in embryos is directed by the EDEN cis element. The EDEN sequence is bound by the protein EDEN-BP, which may interact, directly or indirectly, with PARN.

Joel D. Richter. Microbiol Mol Biol Rev. 1999 June;63(2):446-456.
3.
FIG. 1

FIG. 1. From: Cytoplasmic Polyadenylation in Development and Beyond.

Critical events during Xenopus oocyte maturation. Progesterone binds a putative cell surface receptor, which leads to a transient decrease in cyclic AMP (cAMP) levels and the activation of Eg2 kinase. Subsequently, dormant c-mos mRNA undergoes polyadenylation-induced translational activation. Newly synthesized Mos, a serine/threonine kinase, activates MAP kinase kinase (MAP kinase cascade), which culminates in the activation of MPF (a heterodimer composed of cyclin B and cdc2). Active MPF, which phosphorylates a number of substrates, is most directly responsible for the manifestations of oocyte maturation.

Joel D. Richter. Microbiol Mol Biol Rev. 1999 June;63(2):446-456.
4.
FIG. 5

FIG. 5. From: Cytoplasmic Polyadenylation in Development and Beyond.

Inhibition of cap ribose methylation abolishes progesterone-induced Mos synthesis and oocyte maturation in Xenopus. (A and B) Oocytes were incubated in the indicated concentrations of S-isobutylthioadenosine (SIBA), an analogue of S-adenosylmethionine, from which methyl groups are donated in methyltransferase reactions. SIBA is a stable competitive inhibitor of methyltransferase reactions. These oocytes were also incubated in the absence (lane 1) or presence (lanes 2 to 6) of progesterone and analyzed for the polyadenylation (A) and methylation (B) of injected c-mos mRNA. M refers to a marker for cap I formation, and N refers to noninjected mRNA processed in an identical manner. Note that while SIBA had little effect on polyadenylation, cap I and cap II formation was completely eliminated at 0.5 and 0.75 mM SIBA. (C) Western blot for Mos protein. Note that while immature oocytes contain no Mos (lane 1), this protein accumulates during oocyte maturation (lane 2). While SIBA has no effect on Mos synthesis at the lower concentrations, it completely prevents Mos accumulation at 0.5 and 0.75 mM. (D) General protein synthesis (i.e., [35S]methionine incorporation) in oocytes was unaffected by SIBA irrespective of concentration. (E) Oocyte maturation (as scored by GVBD) paralleled that of Mos synthesis. That is, the prevention of Mos synthesis by SIBA inhibited oocyte maturation. The bars in panels D and E represent the mean and standard deviation of three experiments. Reprinted from reference 47 with permission of the publisher.

Joel D. Richter. Microbiol Mol Biol Rev. 1999 June;63(2):446-456.
5.
FIG. 4

FIG. 4. From: Cytoplasmic Polyadenylation in Development and Beyond.

Poly(A) tail elongation in the central nervous system. (A) The method used to detect poly(A) tail length is the RT-PCR-based PAT [poly(A) test]. Here, oligo(dT) fused to a GC-rich anchor will anneal to multiple regions along the length of a poly(A) tail. When it is reverse transcribed, the resulting cDNAs will be heterogeneous in size. Following PCR with an mRNA-specific primer and the oligo(dT) anchor, the size heterogeneity will be maintained. Thus, mRNAs with long poly(A) tails will yield cDNAs of diverse sizes, the largest of which will approximate the longest poly(A) tail. On the other hand, mRNAs with short poly(A) tails will yield smaller cDNAs with discrete sizes. In addition, PCR with two mRNA-specific primers will result in products with discrete sizes (internal control). RT, reverse transcription. (B) Visual cortices were removed from rats born and raised in the dark (dark rearing) and then either not exposed to light or exposed to light for 30 to 360 min. The visual cortices were also removed from rats maintained on a standard 12-h light-dark cycle (std.). Following RNA extraction, PATs were performed for α-CaMKII mRNA, which contains a CPE, and neurofilament (NF) mRNA, which does not contain a CPE. The PATs used the same reverse transcription reaction. The products were resolved on an agarose gel and visualized by ethidium bromide staining. Note that the poly(A) tail of α-CaMKII mRNA was elongated in response to light, while the poly(A) tail of NF mRNA was unaffected. (C) The α-CaMKII PCR products from panel B were Southern blotted and probed with radiolabeled α-CaMKII 3′ UTR. This blot confirms that the ethidium bromide staining material in panel B corresponds to α-CaMKII sequences and also shows light-dependent polyadenylation of this mRNA. (D) An aliquot of visual cortex RNA annealed to excess oligo(dT) was incubated with RNase H, which removes the poly(A) tail. This was followed by a PAT for α-CaMKII mRNA, or reverse transcription-PCR with internal, mRNA-specific primers. This control confirms that the heterogeneously sized α-CaMKII sequences in panel B resulted from oligo(dT) priming of the poly(A) tail. (E) Reverse transcription-PCR with two α-CaMKII 3′ UTR-specific primers was performed on the same visual cortex RNA used in panel B. This control confirms that the 3′ UTR of α-CaMKII mRNA was intact, since the PCR product is discrete and has the predicted size. Reprinted from reference 109 with permission of the publisher.

Joel D. Richter. Microbiol Mol Biol Rev. 1999 June;63(2):446-456.

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