A full-length rabbit oviductin cDNA(1909bp) was cloned. It consists of a 5’-UTR of 52bp, an open reading frame (ORF) of 1374bp and a 3’-UTR of 483bp and has more than 80% homology with that of other mammal oviductin...A full-length rabbit oviductin cDNA(1909bp) was cloned. It consists of a 5’-UTR of 52bp, an open reading frame (ORF) of 1374bp and a 3’-UTR of 483bp and has more than 80% homology with that of other mammal oviductins. N-terminal peptide (NTP) (384 residues) and C-terminal peptide (CTP) (73 residues) of deduced protein precursor has about 80% and 50% identity with that of other mammals respectively. Fusion proteins GST-NTP 368(1R-368N)and GST-CTP73 (369F-441A) were expressed and purified. NH2-terminal of CTP sequencing reveals that the purified protein is consistent with the deduced one. In order to study the function of NTP and CTP the mouse anti-NTP and rabbit anti-CTP antisera were prepared. Tissue-specific (skeleton muscle, oviduct, uterus, ovary, liver, heart and brain) analysis indicated that rabbit oviductin was only found in oviduct. The conditioned medium derived from the rabbit oviduct mucosa epithelial cells has a function of overcoming the early embryonic development block of Kunming mous e cultured in vitro. Anti-CTP antiserum could totally inhibit the early embryo development at 2-cell stage cultured in the conditioned culture medium, but anti-NTP antiserum couldn’t. There was a positive relationship between the ratio of early embryos at development block and the dosage of anti-CTP antiserum added in the conditioned culture medium. These results suggest that oviductin has a function not only on fertilization, but also on the release of early embryonic development block, and the later function domain of rabbit oviductin may be situate in its C-terminal.展开更多
Anti-keratin monoclonal antibody AF5 was introduced into fertilized eggs of Xenopus laevis., and its effects on embryonic development were studied. Survival rate of the antikeratin-injected embryos was much lower (onl...Anti-keratin monoclonal antibody AF5 was introduced into fertilized eggs of Xenopus laevis., and its effects on embryonic development were studied. Survival rate of the antikeratin-injected embryos was much lower (only 35.76% at gastrula) than that of the control (74.85% at gastrula), in which embryos were injected with mouse IgG. Most of survivors in the experimental series showed aberrant external appearance. On the other hand, in cleavage stage, ie 2-7 h after fertilization, immunohistochemi-cal staining of embryos showed that the experimental embryos were mostly keratin negative, while embryos of the control ones were keratin positive. When introducing this antikeratin into one cell of a 2-cell embryo, only the unin-jected half of the embryo continued its development while the other half could not develop at all. These results suggested that intact keratin cytoskeleton in early embryos is indispensable to the embryonic development of Xenopus laevis.展开更多
基金Supported by National Natural Science Foundation of China (39730460)National "973" Project (G1999055902)National Labora-
文摘A full-length rabbit oviductin cDNA(1909bp) was cloned. It consists of a 5’-UTR of 52bp, an open reading frame (ORF) of 1374bp and a 3’-UTR of 483bp and has more than 80% homology with that of other mammal oviductins. N-terminal peptide (NTP) (384 residues) and C-terminal peptide (CTP) (73 residues) of deduced protein precursor has about 80% and 50% identity with that of other mammals respectively. Fusion proteins GST-NTP 368(1R-368N)and GST-CTP73 (369F-441A) were expressed and purified. NH2-terminal of CTP sequencing reveals that the purified protein is consistent with the deduced one. In order to study the function of NTP and CTP the mouse anti-NTP and rabbit anti-CTP antisera were prepared. Tissue-specific (skeleton muscle, oviduct, uterus, ovary, liver, heart and brain) analysis indicated that rabbit oviductin was only found in oviduct. The conditioned medium derived from the rabbit oviduct mucosa epithelial cells has a function of overcoming the early embryonic development block of Kunming mous e cultured in vitro. Anti-CTP antiserum could totally inhibit the early embryo development at 2-cell stage cultured in the conditioned culture medium, but anti-NTP antiserum couldn’t. There was a positive relationship between the ratio of early embryos at development block and the dosage of anti-CTP antiserum added in the conditioned culture medium. These results suggest that oviductin has a function not only on fertilization, but also on the release of early embryonic development block, and the later function domain of rabbit oviductin may be situate in its C-terminal.
文摘Anti-keratin monoclonal antibody AF5 was introduced into fertilized eggs of Xenopus laevis., and its effects on embryonic development were studied. Survival rate of the antikeratin-injected embryos was much lower (only 35.76% at gastrula) than that of the control (74.85% at gastrula), in which embryos were injected with mouse IgG. Most of survivors in the experimental series showed aberrant external appearance. On the other hand, in cleavage stage, ie 2-7 h after fertilization, immunohistochemi-cal staining of embryos showed that the experimental embryos were mostly keratin negative, while embryos of the control ones were keratin positive. When introducing this antikeratin into one cell of a 2-cell embryo, only the unin-jected half of the embryo continued its development while the other half could not develop at all. These results suggested that intact keratin cytoskeleton in early embryos is indispensable to the embryonic development of Xenopus laevis.
