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Assessment of Selective mRNA Translation in Mammalian Cells by Polysome Profiling
Published on: October 28, 2014
Transcriptomic profiling of Ichthyophthirius multifiliis reveals polyadenylation of the large subunit ribosomal RNA
Jason W Abernathy1, De-Hai Xu, Ping Li
1The Fish Molecular Genetics and Biotechnology Laboratory, Department of Fisheries and Allied Aquacultures and Program of Cell and Molecular Biosciences, Aquatic Genomics Unit, 203 Swingle Hall, Auburn University, AL 36849, USA.
Abstract:
Polyadenylation of eukaryotic transcripts is usually restricted to mRNA, providing transcripts with stability from degradation by nucleases. Conversely, an RNA degradation pathway can be signaled through poly (A) tailing in prokaryotic, archeal, and organellar biology. Recently polyadenylated transcripts have also been discovered in rRNA in some eukaryotes including humans and yeast. Here we report the discovery of polyadenylated rRNAs in the ciliate teleost parasite Ichthyophthirius multifiliis, an important fish pathogen. Through large-scale analysis of ESTs, a large contig composed of the 28S rRNA with poly (A) tails was identified. Analysis using multiple sequence alignments revealed four potential polyadenylation sites including three internal regions and the 3? end of the rRNA. Further analysis using a polyadenylation test, re-sequencing, and gene-specific PCR using primers flanking the presumed poly (A) sites confirmed the presence of polyadenylated rRNA in this parasite. The functions of polyadenylation of rRNA in this organism are largely unknown at present, but the presence of internal polyadenylation sites, along with the presence of truncated segments of the rRNA, may suggest a role of the polyadenylation in the degradation pathway, a function typical of prokaryotes, archaea, and organelles. These results are in congruence with reports of a similar phenomenon in humans and yeast.
Insights
Polyadenylation, typically for mRNA stability, was found in rRNA of the fish parasite Ichthyophthirius multifiliis. This suggests a potential role in RNA degradation, similar to prokaryotes.
Area of Science:
- Molecular Biology
- Parasitology
- Eukaryotic Gene Expression
Background:
- Polyadenylation usually stabilizes eukaryotic mRNA against degradation.
- In prokaryotes and organelles, polyadenylation can signal RNA degradation.
- Polyadenylated rRNA has recently been observed in some eukaryotes like humans and yeast.
Purpose of the Study:
- To investigate the presence and characteristics of polyadenylated rRNA in the ciliate parasite Ichthyophthirius multifiliis.
- To explore potential polyadenylation sites and their implications for rRNA function in this fish pathogen.
Main Methods:
- Large-scale analysis of expressed sequence tags (ESTs) to identify polyadenylated rRNA contigs.
- Multiple sequence alignments to locate potential polyadenylation sites.
- Polyadenylation assays, re-sequencing, and gene-specific PCR to confirm findings.
Main Results:
- Discovery of polyadenylated 28S rRNA in Ichthyophthirius multifiliis.
- Identification of four potential polyadenylation sites, including internal regions and the 3' end.
- Confirmation of polyadenylated rRNA through experimental validation.
Conclusions:
- Polyadenylated rRNA exists in the fish parasite Ichthyophthirius multifiliis.
- The presence of internal polyadenylation sites and truncated rRNA segments suggests a role in RNA degradation.
- Findings align with similar observations in humans and yeast, indicating a conserved, albeit less understood, phenomenon.
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