Putative role for RNA splicing in maize endosperm-embryo developmental interactions is revealed by the rough endosperm 3 (rgh3) seed mutant

Endosperm-embryo interactions are an important but poorly understood aspect of seed development. The Rough endosperm3 ( Rgh3) locus is involved in these interactions at a developmental level. The rgh3 seed mosaics marked with the pr1 anthocyanin gene indicate that Rgh3 is required in the endosperm for the normal development of the embryo. The Rgh3 locus also has an autonomous function in the embryo and is required for seedling viability. The Rgh3 locus maps to the long arm of chromosome 5. Complementation tests with other seed mutants mapped to the same region indicated that Rgh3 is a novel locus. We identified a tightly-linked transposon-tag from the rgh3-70 allele. This transposon insertion disrupts a predicted splicing factor with a U2 snRNP auxiliary factor homology motif (UHM) that is most similar to the U2AF35-Related Protein (URP). The UHMs are RNA recognition motif (RRM)-like domains that function in protein-protein interactions. Analysis of Rgh3/ZmURP cDNAs and genomic sequences suggests ZmURP produces alternatively spliced transcripts. Maize ESTs indicated that ZmURP is expressed in endosperm tissue. Consistent with this expression pattern, the rgh3 phenotype is characterized by an overproliferation of the aleurone cells and aberrant development of the basal endosperm transfer cell layer. An analysis with endosperm cell type specific markers in mutant rgh3 seeds suggests that the basal endosperm transfer cell defect occurs after cell type specification. Tissue culture of rgh3 endosperm cells showed robust growth and formed homozygous mutant callus. These data indicate that Rgh3 is not an essential gene for gene expression. Instead the data presented argue that the putative RNA splicing function of Rgh3 has an important role in cell differentiation, which then impacts endosperm-embryo developmental interactions.