OsAP65, a rice aspartic protease, is essential for male fertility and plays a role in pollen germination and pollen tube growth.

Aspartic proteases (APs) comprise a large proteolytic enzyme family widely distributed in animals, microbes, viruses, and plants. The rice genome encodes 96 APs, of which only a few have been functionally characterized. Here, the identification and characterization of a novel AP gene, OsAP65, which plays an indispensable role in pollen tube growth in rice, is reported. The T-DNA insertion line of OsAP65 caused severe segregation distortion. In the progeny derived from an individual heterozygous for the T-DNA insertion, the wild type and T-DNA-carrying heterozygote segregated at a ratio close to 1:1, while homozygotes of disrupted OsAP65 (OsAP65-/-) were not recovered. Reciprocal crosses between heterozygotes and wild-type plants demonstrated that the mutant alleles could not be transmitted through the male gamete. Examination of the anthers from heterozygous plants revealed that the mutant pollen matured normally, but did not germinate or elongate. OsAP65 was expressed in various tissues and the transcript level in heterozygous plants was about half of the amount measured in the wild-type plants. The subcellular localization showed that OsAP65 is a pre-vacuolar compartment (PVC) protein. These results indicated that OsAP65 was essential for rice pollen germination and tube growth.