Integrative analysis of transcriptome complexity in pig granulosa cells by long-read isoform sequencing

PeerJ. 2022 May 25:10:e13446. doi: 10.7717/peerj.13446. eCollection 2022.

Abstract

Background: In intensive and large-scale farms, abnormal estradiol levels in sows can cause reproductive disorders. The high incidence rate of reproductive disturbance will induce the elimination of productive sows in large quantities, and the poor management will bring great losses to the pig farms. The change in estradiol level has an important effect on follicular development and estrus of sows. To solve this practical problem and improve the productive capacity of sows, it is significant to further clarify the regulatory mechanism of estradiol synthesis in porcine granulosa cells (GCs). The most important function of granulosa cells is to synthesize estradiol. Thus, the studies about the complex transcriptome in porcine GCs are significant. As for precursor-messenger RNAs (pre-mRNAs), their post-transcriptional modification, such as alternative polyadenylation (APA) and alternative splicing (AS), together with long non-coding RNAs (lncRNAs), may regulate the functions of granulosa cells. However, the above modification events and their function are unclear within pig granulosa cells.

Methods: Combined PacBio long-read isoform sequencing (Iso-Seq) was conducted in this work for generating porcine granulosa cells' transcriptomic data. We discovered new transcripts and possible gene loci via comparison against reference genome. Later, combined Iso-Seq data were adopted to uncover those post-transcriptional modifications such as APA or AS, together with lncRNA within porcine granulosa cells. For confirming that the Iso-Seq data were reliable, we chose four AS genes and analyzed them through RT-PCR.

Results: The present article illustrated that pig GCs had a complex transcriptome, which gave rise to 8,793 APA, 3,465 AS events, 703 candidate new gene loci, as well as 92 lncRNAs. The results of this study revealed the complex transcriptome in pig GCs. It provided a basis for the interpretation of the molecular mechanism in GCs.

Keywords: Alternative splicing; Granulosa cells; Iso-Seq; LncRNAs; Pig.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Estradiol
  • Female
  • Granulosa Cells
  • Protein Isoforms / genetics
  • RNA, Long Noncoding* / genetics
  • Swine / genetics
  • Transcriptome* / genetics

Substances

  • RNA, Long Noncoding
  • Protein Isoforms
  • Estradiol

Grants and funding

This work was supported by the Natural Science Foundation of Inner Mongolia (2020BS03004), Research Program of Science and Technology at Universities of Inner Mongolia Autonomous Region of China (NJZZ20087), the Key Project of National Natural Science Foundation of China (31660340), Inner Mongolia University of Science Initiative Scientific Research Program (2019QDL-B43, 2019QDL-B44). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.