Structure and expression of spermidine synthase genes in apple: two cDNAs are spatially and developmentally regulated through alternative splicing.

Abstract

Three cDNAs (MdSPDS1, 2a and 2b) encoding spermidine synthase (SPDS), a key enzyme in the polyamine biosynthesis, have been cloned from apple [Malus sylvestris (L.) Mill. var. domestica (Borkh.) Mansf.]. The deduced amino acid sequences of their protein products share 76-83% identity with SPDSs of other higher plants. A comparison of the sequences of the three cDNAs and of the two corresponding genomic DNA fragments (SPDS1 and SPDS2) indicated that MdSPDS1 was transcribed from the SPDS1 sequence, whereas MdSPDS2a and MdSPDS2b were both derived from SPDS2 by alternative splicing. To learn more about the physiological roles of MdSPDS1, MdSPDS2a and MdSPDS2b, Northern analyses were carried out, together with measurements of polyamine content. Levels of both MdSPDS1 and MdSPD2a were higher in young leaves than in mature leaves and shoots. In fruits, mRNA levels were nearly as high as in young leaves and remained high during fruit development. By RT-PCR, MdSPDS2b transcripts were detected in mature leaves and shoots, but not in young leaves and fruits. These results indicate that MdSPDS2a and MdSPDS2b are differentially regulated in a tissue- and developmentally specific manner. The content of free polyamines in mesocarp tissues was measured at five stages of fruit development. At all stages, spermidine (Spd) was the predominant form of polyamine. The level of Spd was high at the early growth stage and declined to about 90% during later developmental stages. The possible regulation of SPDS expression during apple fruit development is discussed.