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Antisense RNA directed against the 3' noncoding region prevents dormant mRNA activation in mouse oocytes
S Strickland1, J Huarte, D Belin
1Institute of Histology and Embryology, University of Geneva Medical School, Switzerland.
Abstract:
Primary mouse oocytes contain untranslated stable messenger RNA for tissue plasminogen activator (t-PA). During meiotic maturation, this maternal mRNA undergoes a 3'-polyadenylation, is translated, and is degraded. Injections of maturing oocytes with different antisense RNA's complementary to both coding and noncoding portions of t-PA mRNA all selectively blocked t-PA synthesis. RNA blot analysis of t-PA mRNA in injected, matured oocytes suggested a cleavage of the RNA.RNA hybrid region, yielding a stable 5' portion, and an unstable 3' portion. In primary oocytes, the 3' noncoding region was susceptible to cleavage, while the other portions of the mRNA were blocked from hybrid formation until maturation occurred. Injection of antisense RNA complementary to 103 nucleotides of its extreme 3' untranslated region was sufficient to prevent the polyadenylation, translational activation, and destabilization of t-PA mRNA. These results demonstrate a critical role for the 3' noncoding region of a dormant mRNA in its translational recruitment during meiotic maturation of mouse oocytes.
Insights
The 3' untranslated region of dormant tissue plasminogen activator (t-PA) mRNA is crucial for its activation during mouse oocyte maturation. This region controls polyadenylation, translation, and degradation, enabling t-PA synthesis.
Area of Science:
- Molecular Biology
- Developmental Biology
- Reproductive Biology
Background:
- Maternal messenger RNAs (mRNAs) are stored in primary oocytes and activated during meiotic maturation.
- Tissue plasminogen activator (t-PA) mRNA is present in an untranslated form in primary mouse oocytes.
Purpose of the Study:
- To investigate the role of specific regions of t-PA mRNA in its translational control during mouse oocyte meiotic maturation.
- To identify the regulatory elements responsible for translational recruitment and subsequent mRNA decay.
Main Methods:
- Oocyte maturation was induced in vitro.
- Antisense RNA injections were used to target specific regions of t-PA mRNA.
- RNA blot analysis was performed to assess mRNA cleavage and stability.
- Polyadenylation status and t-PA synthesis were evaluated.
Main Results:
- Antisense RNAs targeting both coding and noncoding regions of t-PA mRNA selectively inhibited t-PA synthesis.
- RNA blot analysis indicated cleavage of t-PA mRNA, generating a stable 5' portion and an unstable 3' portion.
- The 3' noncoding region was susceptible to cleavage in primary oocytes, but protected until maturation.
- Targeting the extreme 3' untranslated region (103 nucleotides) prevented polyadenylation, translation, and destabilization.
Conclusions:
- The 3' untranslated region of dormant t-PA mRNA plays a critical role in its translational activation during mouse oocyte meiotic maturation.
- Specific sequences within the 3' untranslated region regulate polyadenylation, translation, and mRNA decay.
- These findings highlight the importance of post-transcriptional regulation in oocyte development.