Molecular cloning of junctin from human and developing rabbit heart

Canine junctin is a 26-kDa transmembrane protein found in the sarcoplasmic reticulum (SR) membrane in cardiac and skeletal muscle. Junctin has recentl y been shown to bind directly to calsequestrin, the ryanodine receptor, and triadin, Junctin is thought to play a role in facilitating (and perhaps re gulating) Ca2+ release from the SR. Immature heart exhibits decreased utili zation of SR Ca2+ stores for cell contraction. We have cloned human and rab bit cardiac junctin and investigated the expression of junctin in developin g rabbit heart. Human junctin was cloned from an adult cardiac cDNA library , Rabbit junctin was cloned by RT-PCR. Northern blot analysis demonstrates a single primary mRNA transcript of approximately 2.8 kb in hearts from bot h species. Sequence analysis demonstrates greater than 97% homology between the predicted amino acid sequences of human, rabbit, and canine junctin in the putative transmembrane domain and subsequent initial 61 amino acid por tion of the putative luminal domain. These domains also exhibit sequence ho mology with triadin, The C-terminal region shows much lower (72 to 75%) seq uence homology among the three species. In addition, Northern blot analysis demonstrates that the expression of junctin increases markedly in postnata l rabbit myocardium, These findings suggest that the putative transmembrane domain and subsequent initial portion of the putative luminal domain of ju nctin play an important role in the binding of junctin to calsequestrin, th e ryanodine receptor, and triadin in the postnatal heart. Furthermore, the previously described increase in SR Ca2+ release with development is associ ated with the increased expression of junctin, (C) 2000 Academic Press.