Ikeda K, Posillico J T, Boyle M B, Webb A C, Brown E M, Broadus A E
Department of Internal Medicine, Yale University, New Haven, Connecticut 06510.
Mol Endocrinol. 1989 Jul;3(7):1084-9. doi: 10.1210/mend-3-7-1084.
Xenopus oocytes have been shown to faithfully translate, process, and secrete a number of secretory proteins after the injection of heterologous mRNAs. The oocyte has the capacity to perform a variety of posttranslational protein modifications but has been reported to be incapable of carrying out certain two-step cleavages which proceed via propeptide intermediates. We examined the ability of the oocyte to process preproPTH after the injection of parathyroid mRNA. Microinjected oocytes secreted material which could be detected in a sensitive cytochemical bioassay for PTH. This activity paralleled that of the PTH standard in the assay and was entirely eliminated by a competitive inhibitor of PTH binding, by preincubation with an anti-PTH antiserum, and by coinjecting oocytes with an oligonucleotide mixture complementary to PTH sequences. Immunoprecipitable proPTH and PTH were present in oocyte homogenates, but oocyte-conditioned medium contained only mature PTH(1-84). We conclude that the Xenopus oocyte is capable of accurately processing preproPTH to the mature secretory form of the peptide.
爪蟾卵母细胞已被证明在注射异源mRNA后能够忠实地翻译、加工和分泌多种分泌蛋白。卵母细胞有能力进行多种翻译后蛋白质修饰，但据报道它无法进行某些通过前肽中间体进行的两步切割。我们检测了注射甲状旁腺mRNA后卵母细胞加工前甲状旁腺素原（preproPTH）的能力。显微注射的卵母细胞分泌的物质可在一种针对甲状旁腺素（PTH）的灵敏细胞化学生物测定中检测到。这种活性与测定中PTH标准品的活性平行，并且通过PTH结合的竞争性抑制剂、与抗PTH抗血清预孵育以及与与PTH序列互补的寡核苷酸混合物共注射卵母细胞而被完全消除。免疫沉淀的前甲状旁腺素原和PTH存在于卵母细胞匀浆中，但卵母细胞条件培养基中仅含有成熟的PTH(1 - 84)。我们得出结论，爪蟾卵母细胞能够将前甲状旁腺素原准确加工成该肽的成熟分泌形式。
