Anti-angiogenic VEGFAxxxb transcripts are not expressed in the medio-basal hypothalamus of the seasonal sheep

PLoS One. 2018 May 10;13(5):e0197123. doi: 10.1371/journal.pone.0197123. eCollection 2018.

Abstract

This study investigated Vegfa expression in the pars tuberalis (PT) of the pituitary and medio-basal hypothalamus (MBH) of sheep, across seasons and reproductive states. It has recently been proposed that season impacts alternative splicing of Vegfa mRNA in the PT, which shifts the balance between angiogenic VEGFAxxx and anti-angiogenic VEGFAxxxb isoforms (with xxx the number of amino acids of the mature VEGFA proteins) to modulate seasonal breeding. Here, we used various RT-PCR methodologies and analysis of RNAseq datasets to investigate seasonal variation in expression and splicing of the ovine Vegfa gene. Collectively, we identify 5 different transcripts for Vegfa within the ewe PT/MBH, which correspond to splicing events previously described in mouse and human. All identified transcripts encode angiogenic VEGFAxxx isoforms, with no evidence for alternative splicing within exon 8. These findings led us to investigate in detail how "Vegfaxxxb-like" PCR products could be generated by RT-PCR and misidentified as endogenous transcripts, in sheep and human HEK293 cells. In conclusion, our findings do not support the existence of anti-angiogenic VEGFAxxxb isoforms in the ovine PT/MBH and shed new light on the interpretation of prior studies, which claimed to identify Vegfaxxxb isoforms by RT-PCR.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Female
  • HEK293 Cells
  • Humans
  • Hypothalamus / metabolism*
  • Protein Isoforms / biosynthesis
  • Protein Isoforms / genetics
  • RNA, Messenger / biosynthesis*
  • Seasons*
  • Sheep / genetics
  • Sheep / metabolism*
  • Vascular Endothelial Growth Factor A / biosynthesis*
  • Vascular Endothelial Growth Factor A / genetics

Substances

  • Protein Isoforms
  • RNA, Messenger
  • Vascular Endothelial Growth Factor A

Grants and funding

This work was supported by a Career Integration Grant (N°320553) from the FP7-Marie-Curie actions.