| Summary: | m<sup>6</sup>A is one of the most common and abundant modifications of RNA molecules present in eukaryotes. The methyltransferase complex, consisting of methyltransferase-like 3 (METTL3), METTL14, and WTAP, is responsible for the m<sup>6</sup>A modification of RNA. WTAP was identified as an mRNA splicing regulator. Its role as a regulatory subunit of the m<sup>6</sup>A methyltransferase complex in embryonic development remains largely unknown. To investigate the role of WTAP in porcine early embryonic development, si-WTAP was microinjected into porcine parthenogenetic zygotes. WTAP knockdown significantly reduced the blastocyst rate and global m<sup>6</sup>A levels, but did not affect the cleavage rate. Betaine was supplemented into the in vitro culture (IVC) to increase the m<sup>6</sup>A levels. Betaine significantly increased the global m<sup>6</sup>A levels but did not affect the blastocyst rate. Furthermore, the pluripotency genes, including <i>OCT4</i>, <i>SOX2</i>, and <i>NANOG</i>, were downregulated following WTAP knockdown. The apoptotic genes <i>BAX</i> and <i>CASPASE 3</i> were upregulated, while the anti-apoptotic gene <i>BCL2</i> was downregulated in WTAP knockdown blastocysts. TUNEL staining revealed that the number of apoptotic cells was significantly increased following WTAP knockdown. Our study indicated that WTAP has an indispensable role in porcine early embryonic development.
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