Abstract
In angiosperms, seed development is a tightly regulated process often involving different epigenetic mechanisms such as a phenomenon known as imprinting that regulates parental gene expression in a parent-of-origin-dependent manner. In Arabidopsis thaliana a significant site of imprinting is the endosperm, a product of double fertilization where two sperm cells fuse with the egg cell and the central cell of the female gametophyte. AGAMOUS-LIKE 36 (AGL36), a member of the MADS-box transcription factor type I Mγ family, has been identified as an imprinted gene that is controlled at the DNA level by DNA methyltransferase MET1 (METHYLTRANSFERASE1) and glycosylase DEMETER (DME), where MET1 silences the paternal allele and DME activates the maternal allele. A function has not been assigned to AGL36, but the protein has been shown to interact with AGL62. In this study, the biological role of AGL36 has been further investigated in relation to its interaction with AGL62 by the analysis of agl36;agl62 double mutants. In a series of phenotypic and genetic analysis, a 10% transmission reduction of the agl36;agl62 double mutant combination through the female gametophyte was observed, suggesting a role in the female gametophyte. Furthermore, in order to dissect AGL36 expression, transgenic AGL36 rescue lines were generated. AGL36 expression patterns in seeds, flowers and vegetative tissue and imprinting in seeds were studied in single and multiple locus transgenic lines. In general, the AGL36 transgenic lines were moderately to highly over-expressed, also in vegetative tissue, compared to endogenous AGL36. The elevated expression levels in seeds were, however not due to paternal expression, since the overall majority of the transgenes tested were imprinted as endogenous AGL36 and only expressed from the maternal genome. Contradictory to a current hypothesis postulating that imprinted genes expressed in vegetative tissues are deleterious, we find that high levels of AGL36 expression can be tolerated in leaves and flowers. In addition, a female gametophyte defect was observed in two independent lines with high levels of AGL36 expression, suggesting a role for AGL36 in female gametogenesis.