Expression and localization of two anther-specific genes in the tapetum and microspore of Lilium longiflorum

Two stage-specific genes have been isolated from a subtractive cDNA library constructed from developing anthers of Lilium longiflorum. Tandem 5ˊand 3ˊ-rapid amplifications of cDNA ends with polymerase chain reaction (5ˊand 3ˊ-RACE-PCR) were used to obtain the full length cDNA sequences. The LLA89 cDNA contained an open reading frame of 303 bp encoding an acidic polypeptide of 100 amino acids with a calculated molecular mass of 10.2 kDa. The LLA-142 cDNA contained an open reading frame of 171 bp encoding a basic polypeptide of 56 amino acids with a calculated molecular mass of 5.7 kDa. The protein encoded by LLA-89 had a strong hydrophobic region at the N-terminus, indicating the presence of a signal peptide (not found in the LLA-142 protein). Sequence alignment revealed that the protein encoded by LLA-142 is novel, while the protein encoded by LLA-89 is identical to a reported LIM4 protein with unknown function. Total RNA blot analysis indicated that the transcripts of LLA-89 and LLA-142 were anther-specific and differentially detected in the anther wall and in the microspore of developing anthers. The LLA-89 gene could be exogenously induced by gibberellin while the LLA-142 gene could not. In situ hybridization with antisense riboprobes for the two genes in the anther revealed strong signals localized to the tapetal layer of the anther wall. The function of LLA-89 and LLA-142 genes was further discussed.