Genomic imprinting at a porcine ZNF locus via a canonical imprinting mechanism.

Genomic imprinting causes parent-of-origin dependent gene expression, primarily driven by a subset of germline differentially methylated regions that function as imprinting control regions at promoter-associated CpG islands. While these mechanisms have been investigated in depth in mice and humans, our understanding of the molecular basis of genomic imprinting in pigs remains limited, particularly at a non-orthologous porcine locus. This study aimed to investigate a pig locus displaying a canonical DNA methylation-dependent imprinting pattern and explore its potential involvement in transcription-coupled imprinting mechanisms. By comparing parthenogenetic and control porcine day-21 embryos, we identified a maternally methylated differentially methylated region in a previously uncharacterized pig locus, LOC100520903 (ZNF300-like), which may serve as an imprinting control region. This was accompanied by a significantly higher paternal mRNA expression of LOC100520903 in control embryos compared to parthenogenetic embryos (p < 0.05), as detected by RNA-seq. At this locus, a previously unannotated transcript with an alternative first exon located far upstream was predominantly expressed in oocytes (supported by >10 RNA-seq junction reads), alongside a promoter marked by H3K4me3 and an adjacent long terminal repeat element (E-value = 5.8e-62). This transcript was no longer detected from embryogenesis onward (0 reads), at which point the annotated LOC100520903 transcripts became expressed. Concurrently, oocyte-specific DNA methylation was observed at the CpG island of the LOC100520903 gene promoter, indicative of maternal methylation. Moreover, in the pig liver and brain, paternal monoallelic expression was consistently observed based on haplotype-tagged RNA-seq reads. Our findings provide mechanistic insights into genomic imprinting at the porcine LOC100520903 locus.