<jats:title>Abstract</jats:title><jats:p>Serine metabolism is involved in various biological processes. Here we investigate primary functions of the phosphorylated pathway of serine biosynthesis in a non-vascular plant <jats:italic>Marchantia polymorpha</jats:italic> by analyzing knockout mutants of Mp<jats:italic>PGDH</jats:italic> encoding 3-phosphoglycerate dehydrogenase in this pathway. Growth phenotypes indicate that serine from the phosphorylated pathway in the dark is crucial for thallus growth. Sperm development requires serine from the phosphorylated pathway, while egg formation does not. Functional Mp<jats:italic>PGDH</jats:italic> in the maternal genome is necessary for embryo and sporophyte development. Under high CO<jats:sub>2</jats:sub> where the glycolate pathway of serine biosynthesis is inhibited, suppressed thallus growth of the mutants is not fully recovered by exogenously-supplemented serine, suggesting the importance of serine homeostasis involving the phosphorylated and glycolate pathways. Metabolomic phenotypes indicate that the phosphorylated pathway mainly influences the tricarboxylic acid cycle, the amino acid and nucleotide metabolism, and lipid metabolism. These results indicate the importance of the phosphorylated pathway of serine biosynthesis in the dark, in the development of sperm, embryo, and sporophyte, and metabolism in <jats:italic>M. polymorpha</jats:italic>.</jats:p>
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