Rodent epididymal cDNAs identified by sequence homology to human and canine counterparts

<abstract>Aim: Identification of the rodent counterparts of human and canine epididymal cDNAs HE3, HE4 and Ce8/Ly6G5C by sequence homology and analysis of their expression patterns and regulation level in the rat. Methods: 'Electronic screening' of Expressed Sequence Tag (EST) and genomic databases, followed by RT-PCR and Northern blot analysis. Results: Rodent ESTs and genomic sequences homologous to HE3, HE4 and Ce8/Ly6G5C were identified in the public databases and the 'full-length' rat cDNAs cloned. To emphasise their homology to the human and canine genes, they were named Me3/Re3, Me4/Re4 and Re8 for mouse and rat counterparts, respectively. mRNA expression patterns were analysed in rats, including rat HEl and HE5/CD52 counterparts as controls. Re3 and Re8 mRNAs were only found in the rat epididymis, while Re4 showed a broader tissue distribution. Within the epididymis, Re3 and Re4 mRNAs were detected in all regions; Re8, on the other hand, was restricted to the caput. During postnatal development, Re3 and control mRNAs were found from the earliest stages investigated, while Re8 mRNA was observed only from day 24 postnatum, corresponding to the onset of spermatogenesis in the prepubertal testis. Castration and testosterone supplementation of adult male rats suggested that none of the cloned mRNAs was directly androgen-regulated. Efferent duct ligation, however, showed that Re8 mRNA levels depended on testicular factors other than androgens. Conclusion: The novel rodent cDNAs can now be used to monitor epididymal gene expression more closely and to set up various regulatory and functional studies.

Homologous cloning, characterization and expression of a new halophyte phytochelatin synthase gene in Suaeda salsa

The halophyte Suaeda salsa can grow in heavy metal-polluted areas along intertidal zones having high salinity.Since phytochelatins can effectively chelate heavy metals,it was hypothesized that S.salsa possessed a phytochelatin synthase(PCS) gene.In the present study,the cDNA of PCS was obtained from S.salsa(designated as SsPCS) using homologous cloning and the rapid amplification of cDNA ends(RACE).A sequence analysis revealed that SsPCS consisted of 1 916 bp nucleotides,encoding a polypeptide of 492 amino acids with one phytochelatin domain and one phytochelatin C domain.A similarity analysis suggested that SsPCS shared up to a 58.6%identity with other PCS proteins and clustered with PCS proteins from eudicots.There was a new kind of metal ion sensor motif in its C-terminal domain.The SsPCS transcript was more highly expressed in elongated and fibered roots and stems(P<0.05) than in leaves.Lead and mercury exposure significantly enhanced the mRNA expression of SsPCS(P<0.05).To the best of our knowledge,SsPCS is the second PCS gene cloned from a halophyte,and it might contain a different metal sensing capability than the first PCS from Thellungiella halophila.This study provided a new view of halophyte PCS genes in heavy metal tolerance.