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Identification and characterization of novel mouse PDE4D isoforms: molecular cloning, subcellular distribution and
Ananth Chandrasekaran1, Kai Yee Toh, Sean Haoquan Low
1Singapore Institute of Clinical Sciences, Singapore.
Abstract:
We report here the cloning and characterization of short and supershort mouse PDE4D isoforms. PDE4D is one of the phosphodiesterase enzyme families with multiple promoters and splice variants. PDE4 isoforms present in humans, rats and mice share considerable homology in their catalytic and regulatory domains. In this study, we have identified the novel PDE4D2 variant3 (PDE4D2v3) and PDE4D10 isoforms and the mouse orthologs of PDE4D1, PDE4D2 variant1 (PDE4D2v1), PDE4D2 variant2 (PDE4D2v2) and PDE4D6 isoforms. These isoforms have many different lengths of 5'UTR, signifying the use of different transcription start sites. Our data indicate that many novel PDE4D isoforms exist as a result of alternative mRNA splicing, each isoform having unique N-terminal regions and multiple transcription start sites. Subcellular distribution study showed that the PDE4D1 short isoforms are localized to the nucleus while the supershort isoforms (PDE4D2v1, PDE4D2v2, PDE4D2v3, PDE4D6 and PDE4D10) are restricted to the cytoplasm. Deletion study confirmed that the N-terminus of PDE4D1 is necessary for nuclear targeting. In addition, we showed that the unique N-terminus contains nuclear localization signal sequence. Identifying novel tissue-specific PDE4D isoforms with unique N-terminal regions may aid in the development of selective phosphodiesterase inhibitors.
Insights
Researchers identified novel short and supershort phosphodiesterase 4D (PDE4D) isoforms in mice. These variants, generated by alternative splicing, exhibit distinct subcellular localizations, potentially aiding selective inhibitor development.
Area of Science:
- Molecular Biology
- Enzymology
Background:
- Phosphodiesterase 4D (PDE4D) is a key enzyme family with numerous splice variants.
- PDE4 isoforms in humans, rats, and mice share significant homology in critical domains.
Purpose of the Study:
- To clone and characterize short and supershort mouse PDE4D isoforms.
- To investigate the role of unique N-terminal regions and transcription start sites in PDE4D isoform diversity.
Main Methods:
- Cloning and characterization of novel PDE4D isoforms.
- Analysis of 5' untranslated regions (5'UTRs) to identify transcription start sites.
- Subcellular localization studies using deletion analysis.
Main Results:
- Identification of novel mouse PDE4D isoforms: PDE4D2v3, PDE4D10, and orthologs of PDE4D1, PDE4D2v1, PDE4D2v2, PDE4D6.
- Demonstration of diverse 5'UTR lengths, indicating multiple transcription start sites.
- Short PDE4D1 isoforms localize to the nucleus, while supershort isoforms are cytoplasmic.
- N-terminal regions are crucial for nuclear targeting and contain nuclear localization signals.
Conclusions:
- Alternative mRNA splicing and multiple transcription start sites generate diverse PDE4D isoforms.
- Unique N-terminal regions dictate subcellular localization (nucleus vs. cytoplasm).
- Discovery of novel, tissue-specific PDE4D isoforms may facilitate the development of targeted phosphodiesterase inhibitors.
