AP1G2 Affects Mitotic Cycles of Female and Male Gametophytes in Arabidopsis

Abstract

During sexual reproduction in flowering plants, haploid spores are formed from meiosis of spore mother cells. The spores then undergo mitosis, develop into female and male gametophytes, and give rise to seeds after fertilization. We identified a female sterile mutant <i>ap1g2-4</i> from EMS mutagenesis, and analyses of two T-DNA insertion mutants, <i>ap1g2-1</i> ^+/- and <i>ap1g2-3</i> ^-/-, and detected a partial female and male sterility. The <i>ap1g2</i> mutant gametophyte development was arrested at one nuclear stage. A complementation test using a genomic sequence of <i>AP1G2</i> with its native promoter restored the function in the three <i>ap1g2</i> mutant lines. Transcriptome profiling of <i>ap1g2</i> ovules revealed that four genes encoding clathrin assembly proteins PICALM5A/B and PICALM9A/B, which were involved in endocytosis, were downregulated, which were confirmed to interact with AP1G2 through yeast two-hybrid assays and BIFC analysis. Our result also demonstrated that <i>RALFL4-8-15-19-26 CML16</i> and several calcium-dependent protein kinases, including <i>CPK14-16-17</i>, were all downregulated in the ovules of <i>ap1g2-1</i> ^+/-. Moreover, Ca²⁺ concentration was low in impaired gametophytes. Therefore, we proposed that through interaction with PICALM5A/B and PICALM9A/B, AP1G2 may mediate gametogenesis accompanied by Ca²⁺ signaling in Arabidopsis. Our findings revealed a crucial role of AP1G2 in female and male gametogenesis in Arabidopsis and enhanced our understanding of the molecular mechanisms underpinning sexual reproduction in flowering plants.