The relationship between the length of anthers and the stage of development of microspores was examined in rice (Oryza sativa L. cv. Hayayuki). Anthers of < or = 2 mm and 2.1-2.2 mm in length and those ready to dehiscence were determined to be at the uninucleate, binucleate and trinucleate microspore stage, respectively. Two cDNAs (YY1 and YY2), representing genes that are specifically expressed in anthers at the uninucleate microspore stage, were isolated and characterized. YY1 cDNA encoded an open reading frame of 95 amino acids. Eight cysteine residues with the potential to form disulfide bridges were present in the amino acid sequence. There was a hydrophobic region at the N-terminus of the putative protein, suggesting that the YY1 protein might be secreted. This cysteine motif and the hydrophobic N-terminus are conserved among products of several anther-specific genes or cDNAs isolated from various plant species. These proteins are thought to form a superfamily of proteins that are confined to anthers. The YY1 transcript was localized in the tapetal cells and the peripheral cells of the vascular bundle. YY2 cDNA encoded an open reading frame of 389 amino acids and the deduced amino acid sequence exhibited substantial homology to that of chalcone synthase. Expression of YY2 mRNA was confined to the tapetal cells. The genes correspond to YY1 and YY2 cDNAs were shown to exist as single copies in the rice genome.