| Experiment Id | GSE294420 | Name | PAN2 Maintains mRNA PolyA Tail Homeostasis and Regulates Translation during Spermiogenesis in Mouse [PAIso-seq] |
| Experiment Type | RNA-Seq | Study Type | WT vs. Mutant |
| Source | GEO | Curation Date | 2026-07-31 |
| description | Elongation and shortening of the polyA tail are the most common type of post-transcriptional mRNA regulation in eukaryotic cells. Two protein complexes, PAN2-PAN3 and CCR4-NOT, are major cytoplasmic deadenylases that trim the mRNA polyA tails. While PAN2-PAN3 removes the distal portion of the long polyA tails, CCR4-NOT removes adenosine closer to the 3'-UTR. Some subunits in the CCR4-NOT complex have been shown to play synergistic biochemical functions during mouse germ cell development. However, the in vivo functions of the upstream deadenylase PAN2-PAN3 remain unclear, particularly in mammals. In this study, we found that germline-specific deletion of Pan2 leads to male infertility. Pan2 knockout did not affect the prophase of meiosis I, but led to abnormal spermiogenesis and apoptosis of male germ-cells. By Palso-seq and Ribo-lite, we found that the deletion of Pan2 increased the number of long-tail mRNAs in spermiogenesis and disrupted the translation efficiency of genes that should be highly translated at this stage. These findings highlight the crucial physiological functions of PAN2 in gametogenesis and expand our understanding of the posttranscriptional regulation of mRNAs in specific physiological processes. In brief, total RNA was extracted with TRIzol Reagent and Direct-zol RNA MicroPrep (Zymo Research, Cat. no. R2060), and the barcode containing adapter were ligated to the 3-end of mRNA. After being purified by the RNA Clean & Concentrator-5 kit (Zymo Research, Cat. no. R1016), the RNA were RT into cDNA by the UMI containing template-switching oligo (TSO) primer and the first PCR amplification were proceeded. Then the cDNA from rRNA will be removed by the CRISPR-Cas9 system and the second PCR amplification will be performed to get enough cDNA. Then the cDNA from each sample will be concatenated into long molecules by following the MAS-ISO-seq procedure. Finally, all the samples were mixed up according to the molar ratios of cDNA, and at least 500ng of cDNA was sent for SMRTbell library preparation and sequenced on PacBio Revio platform. |
