Abstract
Gene expression is controlled at the transcriptional and post-transcriptional levels. The TACC2 gene was known to be associated with tumors but the control of its expression is unclear. We have reported that activity of the intronic promoter p10 of TACC2 in primary lesion of endometrial cancer is indicative of lymph node metastasis among a low-risk patient group. Here, we analyze the intronic promoter derived isoforms in JHUEM-1 endometrial cancer cells, and primary tissues of endometrial cancers and normal endometrium. Full-length cDNA amplicons are produced by long-range PCR and subjected to nanopore sequencing followed by computational error correction. We identify 16 stable, 4 variable, and 9 rare exons including 3 novel exons validated independently. All variable and rare exons reside N-terminally of the TACC domain and contribute to isoform variety. We found 240 isoforms as high-confidence, supported by more than 20 reads. The large number of isoforms produced from one minor promoter indicates the post-transcriptional complexity coupled with transcription at the TACC2 locus in cancer and normal cells.
Highlights
Gene expression is tightly regulated at the transcriptional and post-transcriptional levels
The intronic p10 promoter of transforming acidic coiled-coil-containing protein 2 (TACC2) is highly active in an endometrial cancer cell line, JHUEM‐1
We used nanopore sequencing to extensively profile TACC2 isoforms derived from the intronic promoter p10
Summary
Gene expression is tightly regulated at the transcriptional and post-transcriptional levels. SMRT sequencers measure fluorescence fluctuations that arise from the addition of a specific n ucleotide[14], whereas nanopore sequencers (MinION, GridION, and PromethION) measure ionic current fluctuations that occur when single-stranded nucleic acids pass through n anopores[15]. They make it possible to determine the entire transcript structure without cloning of individual cDNAs. Nanopore sequencing produces longer read lengths with lower sequencing costs, but generates more sequencing errors (about 10%19,20) than HiFi reads of SMRT sequencing that relies on consensus among multiple passages of sequencing through circularized template. The method to sequence a single strand, called 1D, is available currently, and there is a demand to obtain accurate sequences with that
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