非洲爪蟾眼的形态发生研究

详细观察和描述了非洲爪蟾Xenopus laevis眼的发生和发育变化过程,并分别对各发育时期视网膜的厚度进行了定量分析.非洲爪蟾眼的发牛开始于眼原基的形成,进而形成视泡;晶状体的发生是在视杯外壁增厚的同时诱导覆盖其上的胚胎外胚层内层增厚,形成预定晶状体板;在视网膜和晶状体共同诱导下,预定角膜上皮变为透明的角膜.在视杯出现之前,预定RPE的厚度由厚变薄,NR层不断地增厚直至结构功能完善.     

Telomere Variation in Xenopus laevis

Eukaryotic telomeres are variable at several levels, from the length of the simple sequence telomeric repeat tract in different cell types to the presence or number of telomere-adjacent DNA sequence elements in different strains or individuals. We have investigated the sequence organization of Xenopus laevis telomeres by use of the vertebrate telomeric repeat (TTAGGG)_n and blot hybridization analysis. The (TTAGGG)_n-hybridizing fragments, which ranged from less than 10 to over 50 kb with frequently cutting enzymes, defined a pattern that was polymorphic between individuals. BAL 31 exonuclease treatment confirmed that these fragments were telomeric. The polymorphic fragments analyzed did not hybridize to 5S RNA sequences, which are telomeric according to in situ hybridization. When telomeric fragments from offspring (whole embryos) were compared to those from the spleens of the parents, the inheritance pattern of some bands was found to be unusual. Furthermore, in one cross, the telomeres of the embryo were shorter than the telomeres of the parents’ spleen, and in another, the male’s testis telomeres were shorter than those of the male’s spleen. Our data are consistent with a model for chromosome behavior that involves a significant amount of DNA rearrangement at telomeres and suggest that length regulation of Xenopus telomeres is different from that observed for Mus spretus and human telomeres.     

Gene-centromere mapping in Xenopus laevis

Gynogenesis was used to map eight loci to their centromeres in Xenopus laevis. Several loci remote from their centromeres were identified. This information may be useful in distinguishing gynogenetic diploid progeny produced by suppression of second polar bodies from gynogenetic diploid progeny homozygous at all loci produced by suppression of first cleavage of gynogenetic haploids.     

Engineered telomeres in transgenic Xenopus laevis

The expanding roles of telomeres in epigenetic gene regulation, nuclear organization, and human disease have necessitated the establishment of model organisms in which to study telomere function under normal developmental conditions. We present an efficient system for generating numerous vertebrate animals containing engineered telomeres using a Xenopus laevis transgenesis technique. Our results indicate Xenopus zygotes efficiently recognize telomeric repeats at chromosome break points and form telomeric complexes thus generating a new telomere. The resulting transgenic animals progress through normal development and successfully metamorphose into froglets despite the chromosome breakage. Overall, this presents an efficient mechanism for generating engineered telomeres in a vertebrate system and provides an opportunity to investigate epigenetic aspects of telomere function during normal vertebrate development.     

Evolution by recombination and transspecies polymorphism in the MHC class I gene of Xenopus laevis

The patterns of major histocompatibility complex (MHC) evolution involve duplications, deletions, and independent divergence of loci during episodes punctuated by natural selection. Major differences in MHC evolution among taxa have previously been attributed to variation in linkage patterns of class I and class II MHC genes. Here we characterize patterns of evolution in the MHC class Ia gene of Xenopus laevis in terms of polymorphism, recombination, and extent of transspecies polymorphism. We also compare these patterns to see if a correlation exists with linkage or separation of the MHC class I and class II regions as seen in amphibians and teleost fishes. In X. laevis, we find high levels of polymorphism. Also, genetic exchange is relatively frequent and occurs in intron II, reshuffling allelic forms of exons 2 and 3. Evolutionary relationships among class I alleles show an intermingling of alleles from divergent Xenopus species rather than a species-specific clustering. Results indicate that the patterns of evolution are similar to those found in salmonid fishes and are different from the mode of evolution seen in primates. Similar patterns of class Ia evolution in salmonid fishes and X. laevis suggest that nonlinkage of class I and class II regions alone is insufficient to explain some patterns of MHC evolution in salmonids.     

Mitogen-induced B-cell differentiation in Xenopus laevis

Abstract. Four genes are known to affect pigmentation in the Mexican axolotl. The purpose of this article is to review previous information pertinent to these genes and to reevaluate such information in light of new evidence that demonstrates (in a preliminary way) how pigments, and subsequently phenotypes, are affected by the various pigment genes. Each of the mutant phenotypes – m (melanoid), ax (axanthic), a (albino), and d (white) - is compared to the wild type (D). All of these genes are recessives, all of them affect phenotypic changes during development, and three of the four ( m, a , and d ) also affect specific biochemical (i.e., pigment) changes during development. In the axolotl, color patterns can be directly correlated to the presence(or absence) of a variety of pigments that are normally found in discrete pigment cells. Qualitative and quantitative analyses of the bright-colored pigments (pteridines and flavins in this case) present in axolotl skin demonstrate that these pigments vary significantly among the various phenotypes under consideration. These analyses raise some interesting questions with regard to how each of the pigment genes is believed to act, and numerous possibilities for continued experimentation are suggested.     

Facial Transplants in Xenopus laevis Embryos

Craniofacial birth defects occur in 1 out of every 700 live births, but etiology is rarely known due to limited understanding of craniofacial development. To identify where signaling pathways and tissues act during patterning of the developing face, a ''face transplant'' technique has been developed in embryos of the frog Xenopus laevis. A region of presumptive facial tissue (the "Extreme Anterior Domain" (EAD)) is removed from a donor embryo at tailbud stage, and transplanted to a host embryo of the same stage, from which the equivalent region has been removed. This can be used to generate a chimeric face where the host or donor tissue has a loss or gain of function in a gene, and/or includes a lineage label. After healing, the outcome of development is monitored, and indicates roles of the signaling pathway within the donor or surrounding host tissues. Xenopus is a valuable model for face development, as the facial region is large and readily accessible for micromanipulation. Many embryos can be assayed, over a short time period since development occurs rapidly. Findings in the frog are relevant to human development, since craniofacial processes appear conserved between Xenopus and mammals.     

Lipids of Xenopus laevis Spermatozoa

Xenopus laevis sperm lipid composition has been studied. The cholesterol content of Xenopus spermatozoa is 194 μ/mg DNA. Their content of glycolipids and phospholipids (measured as inorganic phosphorus) is respectively 40 and 27 μ/mg DNA. The phospholipid pattern is quite homogeneous and all the principal molecular species are present. In all the examined samples, a glycolipid with low mobility, not yet structurally identified, is present. Finally, using as a probe filipin, we have observed cholesterol distribution on the Xenopus sperm plasma membrane by freeze-fracture. In agreement with the chemical data here presented, Xenopus spermatozoa are heavily labelled by filipin. The filipin-cholesterol complexes seem to be distributed on the entire sperm plasma membrane and appear as protuberances on the P face, suggesting that most of the cholesterol reside in the inner leaflet of the membrane.     

Alternative housing for Xenopus laevis

At the authors' facility, housing arrangements for Xenopus laevis were cumbersome and labor-intensive, requiring technicians to wash frog tanks by hand several times a week. The authors describe an alternative housing solution they implemented by modifying a rack system that was originally used to maintain zebrafish. The rack's self-contained water circulation and filtration system saved technicians time and labor, and a commercial chiller attached to the mechanism efficiently controlled frogs' environmental temperature.     

Studies on Maturation in Xenopus laevis Oocytes

The following results were obtained: 1) The oxygen comsumption of progesterone-stimulated X. laevis oocytes increases at the time of germinal vesicle (GV) breakdown. 2) Continous treatment with 1 mM KCN, 1 mM and 0.1 mM DNP completely inhibits GV breakdown. 3) Pretreatment experiments with KCN, DNP and cycloheximide show that binding of progesterone to the plasma membrane and the specific hormone receptor requires neither energy, nor protein synthesis. 4) 1 h pulses of DNP (1mM) or cycloheximide (50 mug/ml) were applied, at various time intervals after progesterone treatment: early pulses strongly delay, but do not prevent GV breakdown; late pulses just before GV breakdown induce a number of cytological abnormalities, which ultimately lead to cytolysis. The significance of these results is discussed and the hypothesis that synthesis of the maturation producing factor (MPF) requires both energy and protein synthesis is proposed.     

Hypothalamic neurosecretion and metamorphosis in Xenopus laevis

Summary Normal and propylthiouracil (PTU) treated Xenopus laevis tadpoles were fixed during all stages of metamorphosis and sagittal sections were stained with aldehyde fuchsin (AF) or pseudoisocyanine (PIC). Whereas AF positive neurosecretory material could only be demonstrated in the preoptic nucleus from late prometamorphosis, increasing amounts of PIC positive material were found in cells of the dorsal part of the preoptic region from early premetamorphosis. The development of these cells correlated with that of the thyrotropic cells and the thyroids. Likewise, signs of hyperactivity in thyroids and thyrotropic cells of PTU treated larvae were accompanied by a depletion of the dorsal PIC positive cells. In the ventral preoptic region PIC positive cells developed from late prometamorphosis in control larvae, but failed to do so in PTU treated animals. It is argued that the differentiation of the PIC positive cells is largely dependent on thyroid hormones; that the dorsal PIC positive cells may produce a thyrotropin releasing factor; and that the function of these dorsal cells is inhibited by thyroid hormones.The authors thank Dr. P. G. W. J. van Oordt for his active interest and helpful advices, Miss Tineke Aafjes for technical assistance and Mr. H. van Kooten for making the photographs.     

Oogenesis in Xenopus laevis (Daudin)

Summary This study was designed to explore the relationship of estrogen, human chorionic gonadotropin (HCG), and food availability to endocytosis in developing oocytes. When estrogen alone is administered to an animal, large amounts of vitellogenin are synthesized by the liver and secreted into the circulatory system, where it accumulates. Under these conditions there is no evidence of endocytosis at the surface of the oocytes. Other studies have shown that following HCG injection into estrogen-treated animals, vitellogenin is removed from the circulation and the oocyte surface is highly contoured and displays endocytotic activity. Food deprivation has much the same effect on oocyte endocytosis as does estrogen. When animals are given HCG and subsequently starved for 20 days, developing oocytes show little endocytotic activity. We conclude that HCG acts to promote or stimulate endocytosis in developing oocytes while estrogen and/or starvation inhibits this process.Research sponsored by the Energy Research and Development Adiministration under contract with Union Carbide Corporation.Predoctoral fellow supported by Grant GM 1974 from the National Institute of General Medical Sciences, National Institutes of Health.     

The cortical response in Xenopus laevis ova

A dependence on extracellular calcium has been demonstrated for fertilization and the cortical response to pricking in Xenopus ova. Neither event occurred in calcium-free solutions or in the presence of divalent cation chelating agents. The calcium-sensitive phase of the cortical response to pricking in dejellied eggs was restricted to the 5–10 sec immediately following the activation stimulus; the initial phase of activation was not calcium dependent. In contrast, the cortical response in dejellied Xenopus ova exposed to the chemical activating agents, urethan or methyl urethan, was independent of extracellular calcium. Experimental evidence was presented for the involvement of a direct, nonpropagated cortical reaction in response to urethan stimulation as opposed to a propagated reaction in response to pricking. A cortical response in dejellied eggs was not induced spontaneously by high concentrations of potassium, and the prick response was unaffected by inhibitors of energy transfer processes. Molecular mechanisms operative in the initiation and propagation of the cortical response in animal eggs have been discussed.