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PubMed1986

3B55: a repetitious sequence family which is transcribed and proportionately replicated in germ-line polyploid nuclei of Calliphora erythrocephala.

The chromosomes of dipteran polyploid nurse cell nuclei are functionally analogous to the oocyte lampbrush chromosomes of the Amphibia. In investigating the transcriptional and replicative activity of these nuclei in Calliphora erythrocephala we have identified a cloned highly repetitious DNA fragment which shows enhanced transcriptional activity in these cells and different replicative behavior in these germ-line polyploid nuclei as opposed to somatic polytene nuclei. The clone, 3B55, contains a 6.8-kb insert which consists primarily of tandemly repeated 200-bp sequences defined by RsaI sites. From Southern hybridizations to diploid (embryonic) genomic DNA, 3B55-related DNA was calculated to represent a significant fraction of the haploid genome (0.8% or 5000 kb). In situ hybridizations established that these sequences are present in the pericentric regions of four of the six chromosomes. Thus the 3B55 sequences have the properties of a satellite-type DNA family. Quantitation of the 3B55 200-bp monomer (which represents approximately 60% of the genomic 3B55 DNA sequences in all tissues examined) revealed that in somatic polytene salivary gland nuclei, 3B55 DNA is highly under-replicated to give a monomer representation of only 48 +/- 11 kb per haploid genome. However, in germ-line nurse cell nuclei, 3B55 DNA is proportionately replicated to give a monomer genomic representation (3560 +/- 344 kb) equivalent to that of diploid DNA (3011 +/- 202 kb). Transcripts complementary to 3B55 sequences are at least 25 times more abundant in total nurse cell nuclear RNA than in total embryonic nuclear RNA. These findings suggest an association of the 3B55 sequence family with some germ-line specific function.

Animals↗
PubMed1988

A ventrally localized inhibitor of melanization in Xenopus laevis skin.

Melanophores normally differentiate in dorsal but not in ventral skin of Xenopus laevis. We have sought factors which might regulate this differentiation pattern, and we have obtained a putative melanization inhibiting factor (MIF) from ventral but not from dorsal skin. Preliminary studies reveal that MIF is destroyed by heat or trypsin treatment, indicating its protein composition, and has a molecular weight in the range of 300 kDa. The effects of MIF on the differentiation of neural crest derivatives to melanophores were examined in vitro in the presence of tyrosine and fetal calf serum (FCS). Tyrosine enhances melanophore differentiation in vitro at concentrations equivalent to those estimated in adult Xenopus blood plasma (20 microM). FCS also stimulates melanization, by way of materials other than the tyrosine contained in FCS. MIF strongly inhibits outgrowth and melanization of neural crest cells from neural tube explants. MIF also inhibits the differentiation of melanoblasts contained in cultured explants of ventral skin. Inhibition of melanization or melanophore differentiation by MIF occurs even in the presence of L-tyrosine and/or FCS. We suggest that MIF plays an important role in the establishment of dorso-ventral pigment patterns in amphibia.

Animals↗
PubMed1989

Intracellular signals for developmental hemoglobin switching.

We have detected trans-acting factors that regulate developmental hemoglobin switching by fusing erythroid cells of different developmental programs. Adult erythroid cells of one anuran species, Xenopus laevis, were fused with tadpole erythroid cells of another frog, Rana catesbeiana. In a second set of experiments, dimethyl sulfoxide-induced murine erythroleukemia cells, which express only adult mouse globins, were fused with Rana tadpole erythroid cells, which express only embryonic and fetal-like globins. Adult Rana globin gene expression was detected in both sets of transient heterokaryons at 6 hr after fusion. Dot blots and Northern blots of total RNA from the heterokaryons contained material that reacted with an adult Rana alpha-globin probe; newly synthesized adult Rana hemoglobin tetramers were detected with native polyacrylamide gel electrophoresis. These results show that developmental stage-specific transacting factors for globin genes can function across vertebrate classes (mammalia to amphibia) and suggest that the mechanisms that regulate developmental hemoglobin switching are highly conserved.

Animals↗
PubMed1992

Expression of DNA ligases I and II during oogenesis and early development of Xenopus laevis.

We have analyzed the expression of DNA ligase I protein during oogenesis and early development of Xenopus laevis. The protein is already present in stage I oocytes and then accumulates throughout oogenesis to reach a steady state level by stage VI. It remains at this level at least until tadpole stage. In stage VI oocytes DNA ligase I protein is almost exclusively localized in the germinal vesicle. We have partially purified a DNA ligase II activity from stage VI oocytes, unfertilized eggs, and stage 8 embryos. An 80-kDa polypeptide can be specifically adenylated in all three purified extracts. It is not recognized by antibodies directed against DNA ligase I and is active on oligo(dT)-poly(rA) substrate. It could therefore represent DNA ligase II protein. The presence of both DNA ligases I and II in oocytes and embryos is inconsistent with the DNA ligase model that had been previously proposed for amphibia.

Animals↗
PubMed1984

A monoclonal antibody against nuclear lamina proteins reveals cell type-specificity in Xenopus laevis.

Immunofluorescence microscopy shows that the monoclonal murine antibody PKB8 stains the nuclear lamina of various somatic cells from vertebrates as diverse as mammals, birds and amphibia. It also decorates the nuclear periphery of oocytes from rat and chicken but does not react with spermatocytes, spermatids and spermatozoa. Immunoblotting experiments demonstrate reaction with lamina polypeptides A, B and C of rat, with lamina polypeptide A of chicken, and with lamina polypeptides LI and LII of erythrocytes of the frog, Xenopus laevis. Antibody PKB8 does, however, not bind, on blotted polypeptides and on sections through ovaries, to the pore complex-lamina polypeptide of Mr 68000 present in Xenopus oocytes. These results reveal the existence of a common antigenic determinant in all three lamina polypeptides of mammals, in one lamina polypeptide of chicken and in two amphibian lamina polypeptides. The immunological data also indicate that, in Xenopus laevis, pore complex-lamina polypeptides of somatic cells and oocytes are distinct. The Mr 68000 protein of Xenopus oocytes is also different from polypeptides LI and LII of somatic Xenopus cells by tryptic peptide mapping. The results suggest that nuclear pore complex-lamina polypeptides represent a family of related polypeptides containing regions highly conserved during evolution and that these polypeptides can be differentially expressed in cells of at least one species, Xenopus laevis.

Animals↗
PubMed1989

Cold-induced changes in amphibian oocytes.

Female Pleurodeles waltl newts (Amphibia, urodele), usually raised at 20 degrees C, were submitted to low temperatures; oocytes responded to this cold stress by drastic changes both in lampbrush chromosome structure and in protein pattern. Preexisting lateral loops of lampbrush chromosomes were reduced in size and number, while cold-induced loops which were tremendously developed, occurred on defined bivalents of the oocyte at constant, reproducible sites. A comparison of protein patterns in control and stressed oocytes showed two main differences: in stressed oocytes, overall protein synthesis was reduced, except for a set of polypeptides, the "cold-stress proteins"; second, there was a striking inversion of the relative amount of beta- and gamma-actin found in the oocyte nucleus before and after cold stress. Whereas beta-actin was the predominant form in control oocytes, gamma-actin became the major form in stressed oocytes.

Animals↗
PubMed1990

Paired arrangement of nonhomologous centromeres during vertebrate spermiogenesis.

Indirect immunofluorescence staining with human anti-kinetochore antibodies was used to study the position of centromeres during vertebrate spermiogenesis. Many species of Amphibia have a low chromosome number and very large spermatids and spermatozoa. The number of kinetochore dots correlates exactly with the haploid chromosome number. This implies that kinetochore duplication occurs in the interval between meiosis I and meiosis II. The nonhomologous centromeres are arranged in tandem during the entire course of spermiogenesis and in mature spermatozoa. A higher order centromere arrangement was found in spermiogenic cells of Anura and Urodela. In mammals, immunofluorescence analysis is complicated by the extreme condensation of chromatin during spermiogenesis and the high chromosome numbers. Nevertheless, centromere-centromere associations were observed in mammalian round spermatids and sporadically in testicular spermatozoa. This indicates that pair-wise association of centromeres is a universal principle of centromere arrangement at the postmeiotic stage.

Animals↗
PubMed1985

Projections of luteinizing hormone-releasing hormone and vasotocin fibers to the anterior part of the preoptic nucleus in the toad, Bufo japonicus.

The anterior part of the preoptic nucleus (APON) is a crucial locus for triggering male mate calling behavior in anuran amphibia. The projections to this locus of luteinizing hormone-releasing hormone and vasotocin fibers were immunohistochemically demonstrated in the brain of the toad (Bufo japonicus) using the avidin-biotin-peroxidase complex method. Immunoreactive (ir)-LH-RH perikarya are localized in the nucleus medialis septi and the nucleus of the diagonal band of Broca. A part of ir-LH-RH fibers arising from these nuclei project to the lateral part of the APON, where the APON neurons form their dendritic field, and often protrude into its medial neuronal cell mass. Meanwhile, a considerable number of vasotocin fibers arising from the ventral magnocellular part of the preoptic nucleus project anterior to these loci. These observations indicate that the LH-RH and vasotocin fibers may form ordinary and/or en passant synapses with the APON neurons to transmit peptidergic neuronal signals which are concerned with initiation of seasonal reproductive behavior.

Animals↗
PubMed1986

Comparative effects of corticotropin-releasing factor, arginine vasopressin, and related neuropeptides on the secretion of ACTH and alpha-MSH by frog anterior pituitary cells and neurointermediate lobes in vitro.

The ability of corticoliberin (CRF), urotensin I, sauvagine, arginine-vasopressin (AVP), and mesotocin to stimulate ACTH release by frog anterior pituitary cells and alpha-melanotropin (MSH) by frog neurointermediate lobe was studied in vitro using a perifusion technique. CRF and AVP were found to be potent stimulators of ACTH secretion, whereas urotensin I and sauvagine were totally inactive. In opposition to recent findings in the rat. CRF did not modify alpha-MSH secretion by the frog neurointermediate lobe. Mesotocin, which is present in the parenchymal cells of the frog pars intermedia, had no effect on alpha-MSH release in vitro. No potentiation of CRF-induced ACTH release was observed when anterior pituitary cells were incubated with a combination of AVP and CRF. Together with the recent elucidation of a CRF-like molecule in the frog diencephalon, these results suggest that, in Amphibia, CRF and AVP exert their stimulatory action specifically on distal lobe corticotrophs.

Adrenocorticotropic Hormone↗
PubMed1987

Effects of various doses of thyroxine and triiodothyronine on the metamorphosis of flounder (Paralichthys olivaceus).

Two experiments were carried out to clarify the effects of thyroxine (T4) and triiodothyronine (T3) on flounder metamorphosis. In the first experiment, three doses of T4 were administered to premetamorphic flounder larvae together with thiourea (TU). Thiourea treatment (30 ppm in ambient seawater) alone caused metamorphic stasis in the fish, although the beginning of the right eye migration was not completely inhibited. T4 at 100 and 10 ppb in ambient seawater induced metamorphosis in the thiourea-treated larvae. The fish of these groups completed translocation of the right eye to the left side and resorption of elongated dorsal fin rays. However, 1 ppb T4 failed to induce metamorphic climax. The elongated dorsal fin rays continued to grow until the end of the experiment in the group treated with TU alone and in combination with 1 ppb T4. In the second experiment, premetamorphic flounder larvae were treated with two doses of T4 and three doses of T3. The T3 treatment was several times more potent than T4 treatment in the induction of metamorphosis. The results of the present study suggest that the metamorphic events of flounder larvae are dependent on the thyroid hormone concentration, and the relative biological potency of T4 and T3 on flounder metamorphosis is similar to that observed in mammals and amphibia.

Animals↗
PubMed1992

Annual variations in the binding of insulin to hepatic membranes of the frog Rana esculenta.

Amphibia undergo regular annual cycles of metabolic activity that are influenced by both exogenous factors and hormones. Insulin binding to crude frog hepatic membranes was studied throughout the year. The general character of insulin binding was similar to that in other vertebrates; the maximum specific binding was achieved after 4 hr at 4 degrees, the optimum pH was 7.8, half-maximal displacement of bound insulin was from 9 x 10(-10) to 1 x 10(-9) M, and insulin analogs competed for the insulin receptor in line with their relative biological potencies. A biphasic Scatchard plot and negative cooperativity of the receptor were also observed in frog liver membranes. Affinity constants from Scatchard plots revealed high and low affinity binding sites which were unchanged during the year. The seasonal cycle, however, markedly affected the binding capacity for both sites. Maximum binding occurred in May-June and the minimum in November-December for both classes of receptors. Binding capacities ranged from 1.71 to 11.33 fmol/mg protein for the high affinity sites and from 432 to 3171 fmol/mg protein for the low affinity sites. It is concluded that annual cycles of insulin binding reflect modulation of receptor number rather than receptor affinity.

Animals↗
PubMed1990

Phospholipid composition of myelin and synaptosomal proteolipid from vertebrate brain.

1. The phospholipid composition of the main proteolipid complexes of the nervous system was studied in myelin and synaptosomal membranes from brains of representatives of various vertebrate classes. 2. The relative content of acid phospholipids was much higher in proteolipid complexes from myelin and synaptosomal membranes of all vertebrates studied as compared to their content in the initial lipid extract (28-80% and 11-20% of total phospholipid content, respectively). 3. The relative content of acid phospholipids in proteolipid complexes of myelin membranes was much lower in brain of fishes and amphibia as compared to higher vertebrates. 4. The main acid phospholipids of proteolipid complexes was phosphatidylserine, phosphatidic acid being characteristic for myelin proteolipids and diphosphatidyl glycerol for synaptosomal proteolipids of all vertebrates studied.

Animals↗
PubMed1994

Origins of the parasitic habit in the nematoda.

Circumstances that probably attended and influenced the adoption and development of the parasitic habit amongst the Nematoda are examined. Features that allowed early terrestrial nematodes to exploit discontinuous habitats such as decomposing organic matter, are considered to have been advantageous to microbivorous Secernentea that became parasites of animals and plants. This development followed the appearance of a land flora and that the Amphibia were the first vertebrate hosts of nematodes. Life cycles involving intermediate hosts were essential in drier environments and in a aquatic ones where intermediate hosts preserve the infective stages; keeps them "in circulation", and makes them attractive to predators. It is concluded that the parasitic habit was adopted repeatedly in both Secernentea and Adenophorea, though the latter did not diversify as much. Convergence is a common feature of nematode evolution, and the typical life history pattern of 5 stadia separated by 4 moults is often greatly modified by suppression, extension and diversification of stages and their roles. There is a need to examine the nematodes, especially of invertebrates in the remaining rain forests of Gondwanaland before they disappear.

Adenophorea↗
PubMed1992

Identification of a nonapeptide motif in the vimentin head domain involved in intermediate filament assembly.

The assembly of soluble vimentin subunits into intermediate filaments (IFs) is dependent on information located in the amino-terminal domain. Using site-directed mutagenesis of a Xenopus laevis vimentin cDNA and an Escherichia coli production system to obtain pure mutated protein, we have identified, in the head domain, a nine amino acid motif (SSYRRIFGG), evolutionarily conserved from amphibia to man, which plays an important role in the orderly formation of IFs. Exchanges in the central di-arginine and in the two aromatic residues interfere with IF assembly of vimentin in vitro: on assembly under standard assembly conditions (160 mM-NaCl) most of the protein is included in dense aggregates, with a variable and minor proportion of IFs, whereas at lower ionic concentrations short and incomplete IF-like structures are formed. The deletion of the whole motif results in a protein that under standard assembly conditions (e.g. 160 mM-NaCl) predominantly and rapidly precipitates into large aggregates of non-IF material, whereas at lower ionic strength (e.g. 50 mM-NaCl) both IFs and dense aggregates are formed simultaneously. Our results show that the mutated protein can assume different forms at the same time and under the same conditions. This motif alone is insufficient for the formation of normal IFs as demonstrated by a mutant in which the motif has been brought closer to the alpha-helical rod domain by deletion of 55 internal amino acid residues. Corresponding observations have been made, by immunofluorescence microscopy, upon transfection of cultured epithelial cells lacking vimentin IFs. The importance of the head domain motif for the assembly and higher-order arrangement of IFs is discussed.

3T3 Cells↗
PubMed1985

Chemo- and baroreceptor innervation of the aortic trunk of the toad Bufo vulgaris.

In the toad Bufo vulgaris, afferent impulses were recorded from the fine branch of the vagus which terminates in the aortic trunk. Spontaneous single unit discharge was random. When the aorta was artificially perfused, units fired in response to flow change, hypoxia and hypoxia-hypercapnia of the perfusion fluid, and administration of NaCN or Ach. A few baroreceptor fibers fired synchronously with the blood pressure fluctuation. In anesthetized toads with other reflexogenic areas denervated, the intravenous administration of NaCN caused an increase in breathing and a slight elevation of blood pressure. When the nerve concerned was cut, these responses were abolished. Electrical stimulation of the nerve's central cut end brought about the same type of response as that to NaCN. Special cells closely resembling the glomus cells in the carotid labyrinth were found in the aortic wall. In amphibia, there is an aortic nerve corresponding to that of mammals, and the aorta is one of receptive areas predominantly contributing to respiratory control.

Animals↗
PubMed1987

A comparative study of Sertoli cell ectoplasmic specializations in selected non-mammalian vertebrates.

Ectoplasmic specializations (ES) facing spermatids were studied in species representative of four classes of non-mammalian vertebrates (Pisces--bluegill; Amphibia--bullfrog; Reptilia--red eared turtle; Aves--domestic chicken). ES was not seen in the bluegill but was present in all other species studied. In the frog, turtle, and chicken, ES did not resemble its mammalian counterpart and could only be characterized by the presence of 6 nm filaments (presumedly actin) within the somatic cell facing the head region of elongating spermatids. ES filaments were sparse in the frog and were sometimes associated with more deeply situated endoplasmic reticulum. Turtle ES filaments were abundant and encircled the acrosomal region of the spermatid head and were delimited by fenestrated saccules of endoplasmic reticulum. In the chicken, ES filaments were prominent but less abundant than in the turtle. Six nanometer filaments of the chicken ES appeared in a tangled mass and were not associated with clearly defined endoplasmic reticulum. In the three species where ES was found, it first developed as spermatids became entrenched within the surrounding somatic cell. Neither cell elongation, nuclear elongation, or movement of the nucleus to the cell surface was synchronized with the onset of ES development. That ES development was seen concomitant with spermatid entrenchment and spermatid orientation suggested a role for ES in these processes. This hypothesis was further strengthened by observations in the fish where ES was lacking and where spermatid entrenchment within the somatic cell, did not occur. The study also supported the hypothesis that ES acts as a cytoskeletal mantle to which other cytoskeletal elements within the cell interact to affect the position of elongate spermatids within the epithelium. The dissolution of ES prior to spermiation and concomitant loss of a close relationship between cells suggests that ES is also related to somatic cell-germ cell adhesion and therefore plays an important role in the spermiation process.

Animals↗
PubMed1991

Plasma prostaglandin F2 alpha in the male Triturus carnifex (Laur.) during the reproductive annual cycle and effects of exogenous prostaglandin on sex hormones.

Plasma patterns of prostaglandin F2 alpha (PGF2 alpha) and sex hormones (progesterone, androgens and 17 beta-estradiol) have been studied in the male crested newt, Triturus carnifex (Laur.), during the sexual cycle. The effects of exogenous PGF2 alpha on sex steroids have also been observed. In addition, effects of one week's captivity are reported. The patterns of plasma sex hormones, during the annual cycle, are consistent with the results previously reported for the same newt species. PGF2 alpha plasma level peaks in April, is low in summer, and progressively increases during autumn to peak again in December. The April PGF2 alpha peak coincides with a plasma estradiol increase and with an androgens drop. In April-collected newts, moreover, PGF2 alpha treatment induces a significant estradiol increase. These findings lead us to suppose that at the end of the breeding season (April) a PGF2 alpha-dependent estradiol synthesis occurs which could be implied in reproductive period termination. In several vertebrates, including some amphibian species, in fact, chronic administration of estradiol results in a strong inhibition of testicular endocrine tissue activity. The putative role of PGF2 alpha-dependent estradiol production in the gonadal regulation in amphibia living in temperate zones is discussed. The autumn PGF2 alpha increase has been tentatively related to the recovery gonadal processes and secondary sexual character development.

Androgens↗
PubMed1995

Distinct hydro-osmotic receptors for the neurohypophysial peptides vasotocin and hydrins in the frog Rana esculenta.

The biological properties of vasotocin, hydrin 1 (vasotocinyl-Gly-Lys-Arg) and hydrin 2 (vasotocinyl-Gly), in particular the hydro-osmotic activities on the frog skin, the frog urinary bladder and the frog kidney, have been compared. Hydrins are as active or more active than vasotocin on the first two organs but they are virtually devoid of antidiuretic activity in the rat and the frog, in contrast to vasotocin. It appears that where the oxytocin ring (residues 1-6), present in the three peptides, is necessary for the action on the three organs, the C-terminal amidated group of vasotocin is necessary for the renal receptor but not for the skin and bladder receptors. It is known that amphibians have two types of vasotocin receptors, V1 and V2, homologous to the vascular/hepatic V1 and the renal V2 vasopressin receptors of mammals, respectively. We suggest that adaptation has led to specialization of (at least) two subtypes of hydro-osmotic V2 receptors, the renal subtype on which vasotocin is mainly active for the reabsorption of tubular water, and the skin/bladder subtype on which hydrin 2 is specifically involved in ensuring the rehydration of the animal. Cooperative evolution might have created in anuran Amphibia, on the one hand, two hydro-osmotic peptides, vasotocin and hydrin 2, derived from a single precursor through differential processing; and on the other hand, two corresponding receptors in kidney and skin for internal and external water recovery.

Amino Acid Sequence↗