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Making males from females: the effects of aromatase inhibitors on a parthenogenetic species of whiptail lizard.

The parthenogenetic whiptail lizard Cnemidophorus uniparens provides a good model for the study of sex determination and sexual differentiation because genetic variation is minimal and all unmanipulated embryos will develop as females. Thus any deviation from the established course of development can be identified as a treatment effect. Previous work has shown that early prenatal treatment with CGS16949A, a nonsteroidal aromatase inhibitor, causes hatchlings to develop as males. The present study explores more fully the effects of dosage and timing of application of CGS16949A and examines the sex-reversing potential of CGS20267, a new and reputedly more potent aromatase inhibitor. Eggs were treated with a range of dosages of the aromatase inhibitors. Hatchlings that received 1 microgram or more of either inhibitor were all male, while those that received 0.1 microgram or less were all female. No difference in potency between the two compounds was detected. Animals treated with 100 micrograms of CGS16949A on Day 20 of incubation or later were all female, while those treated on Day 5 were all male. Seven sex-reversed male parthenogens have been raised to sexual maturity. The animals appear similar morphologically and behaviorally to males of the sexually reproducing whiptail species. Spermatogenesis and spermiogenesis have been confirmed by histological examination of the testes and by postcopulatory cloacal swabs. Application of aromatase inhibitors has been shown to sex-reverse both avian and reptilian species. In mammals, the male-determining gene of the Y chromosome (SRY) may code for an intrinsic aromatase inhibitor. Studies show the gene's product has a binding domain which recognizes regulatory elements in the promoter of the aromatase gene.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Progesterone inhibits female-typical receptive behavior and decreases hypothalamic estrogen and progesterone receptor messenger ribonucleic acid levels in whiptail lizards (genus Cnemidophorus).

Female-typical sexual behavior in tetrapods is mediated primarily by estrogen and progesterone acting through intracellular receptors at specific sites in the mediobasal hypothalamus. Progesterone exerts both faciliatory and inhibitory actions on female sexual behavior and in well-studied rodent models, the inhibitory actions are exerted through downregulation of progesterone and estrogen receptors. This study examined progesterone effects on both female-typical sexual behavior and hypothalamic estrogen and progesterone receptor mRNA expression (ER- and PR-mRNA) in a sexual and parthenogenetic species of whiptail lizard. Progesterone capsules administered to ovariectomized female Cnemidophorus inornatus and Cnemidophorus uniparens following a receptivity-inducing dosage of estradiol benzoate (EB) strongly inhibited receptive behavior as compared to blank implanted controls. Progesterone capsules administered either before or after an EB injection also strongly downregulated ER- and PR-mRNA abundance in the ventromedial nucleus of the hypothalamus relative to blank implanted controls. The correlated decrease in both EB-induced receptive behavior and ER- and PR-mRNAs following progesterone administration are similar to findings in rats and guinea pigs, suggesting that this is an evolutionarily conserved mechanism in the regulation of female sexual behavior.

Animals↗

Evolutionary changes in dopaminergic modulation of courtship behavior in Cnemidophorus whiptail lizards.

Preoptic dopamine release is integral to the display of copulatory behaviors in male mammals and birds. However, while the anatomical distributions of the dopamine synthesizing enzyme tyrosine hydroxylase are similar among vertebrates, evolutionary changes in the functional role of dopamine are poorly understood. In this study, we tested whether a dopamine D1 receptor agonist would facilitate the display of courtship and copulatory behaviors in two related Cnemidophorine lizards (Cnemidophorus inornatus and Cnemidophorus uniparens). Cnemidophorus lizards offer a unique system to study evolutionary changes in functionality because ancestral (e.g., C. inornatus) and descendant (e.g., C. uniparens) species can be studied in parallel. Cnemidophorus uniparens is an all-female, parthenogenetic species and is the triploid descendant of the sexual and diploid species C. inornatus. Here we report that in castrated male C. inornatus and ovariectomized C. uniparens a dopamine D1 agonist increased the proportion of individuals mounting and decreased the latency to mount. Moreover, there was a species difference in sensitivity to the agonist: Mounting was elicited at a lower dose in C. uniparens than in C. inornatus. One possible explanation for this heightened sensitivity in the triploid parthenogen is that, by virtue of the increased ploidy, the parthenogen has elevated levels of D1 receptor in limbic brain areas modulating courtship behavior.

Animals↗

Koinophilia stabilizes bi-gender sexual reproduction against asex in an unchanging environment.

Koinophilia, the tendency to choose mates with predominantly common phenotypic features, may be favourable because maladapted traits tend to have low frequencies. We used computer simulations to study competition between asexual and sexual reproduction, with and without koinophilia. The mutation process created new alleles, most of which were deleterious, but some small fraction was beneficial. With random mating, genderless sex (which does not suffer the two-fold cost of producing males) had a modest advantage over asex, while bi-gender sex was rapidly replaced by asex. In contrast, koinophilia allowed sex to persist, even when males represented half the population. The advantage of koinophilia was greatest when populations were large, the incidence of mildly disadvantageous mutations was high, and beneficial mutations were rare. Under these circumstances koinophilic bi-gender sexual reproduction resisted replacement by asexuals for over 10,000 generations, even when one sexual mutated into a parthenogen every generation. Thus, koinophilia may be an important factor maintaining sexual reproduction.

Animals↗

The survival of slow reproducers.

Multicellularity, and the attendant segregation of the germ line, entails the loss of reproductive capacity by the soma: in Volvox carteri, less than 1 cell in 100 contributes to the next generation. However, compensatory advantages are unlikely to be very large (Koufopanou & Bell, 1993. Proc. R. Soc. Lond. (B) 254,107-113). Somewhat similarly, sex implies the generation of males, hence a dramatic reproductive slowdown (Barton & Charlesworth, 1998. Science281, 1986-1990); yet, a compensating (two-fold) advantage of sex has not been found. Here, I try to evaluate the actual cost of maintaining slow reproductive cycles, namely cycles that necessitate the production of "dead end" units such as somatic cells or males. In a quantitative model for the competition of individuals with different, heritable reproductive rates, this cost turns out to be unexpectedly small, and may even sometimes become irrelevant. The bases for this are made fairly clear: thus, when all enjoy high fecundity (e.g. a long reproductive life) the handicap of a slower reproduction vanishes; alternatively, a slight separation of ecological niches may be sufficient for survival of slower but otherwise unchanged reproducers; and finally, inherent to slow reproduction is a low rate of destabilizing genetic change. These facts are largely independent of the formal model details, and are supported by direct computer simulations. They give a quantitative basis for analysing the evolution and prevalence of slow life cycles. The implications of these findings for the evolution of multicellularity are briefly discussed.

Animals↗

The effects of deleterious mutations in cyclically parthenogenetic organisms.

Cyclically parthenogenetic organisms experience benefits of both sexual and asexual reproductive modes in a constant environment. Sexual reproduction generates new genotypes and may facilitate the purging of deleterious mutations whereas asexuality has a two-fold advantage and enables maintenance of well-fitted genotypes. Asexual reproduction can have a drawback as increased linkage may lead to the accumulation of deleterious mutations. This study presents the results of Monte Carlo simulations of small and infinite diploid populations, with deleterious mutations occurring at multiple loci. The recombination rate and the length of the asexual period, interrupted by sexual reproduction, are allowed to vary. Here I show that the fitness of cyclical parthenogenetic population is dependent on the length of the asexual period. Increased length of the asexual period can lead both to increased segregational load following sexual reproduction and to a stronger effect of deleterious mutations on variation at a linked neutral marker, either by reducing or increasing the variation.

Animals↗

Analysis and identification of imprinted genes.

Genomic imprinting in mammals results in the unequal expression of the two parental alleles of specific genes. The existence of imprinting in the mouse emerged from nuclear transplantation studies and from the abnormal phenotypes associated with uniparental inheritance of particular chromosome segments. Over the past 5 years, 20 or so imprinted genes have been identified. This has emphasized the important roles played by some imprinted genes in development, permitted a description of the epigenetic properties associated with imprinting, and provided the first insights into the regulation of imprinting. In this article, we discuss the generation of experimental material in which imprinting effects can be analyzed, review the properties of imprinted genes, and discuss how to examine them using state-of-the-art techniques. Finally, we consider the means by which new imprinted genes can be identified.

Alleles↗

Rapid screening of DNA diversity using dot-blot technology and allele-specific oligonucleotides: maternity of hybrids and unisexual clones of hybrid origin (lizards, Cnemidophorus).

Allele-specific oligonucleotide probes, together with dot-blot methods, can provide rapid and inexpensive screening of DNA types in large samples of organisms. Here we demonstrate their use in: (1) determining types of mitochondrial DNA in hundreds of lizards from a dynamic hybrid zone; (2) discovering intraspecific geographic variation in genes; and (3) determining and verifying the maternal ancestry of unisexual, parthenogenetic lizards in clones of hybrid origin. These methods are broadly applicable in research involving rapid screening of DNAtypes in large samples of specimens for any gene with sequence data from which to design specific probes.

Alleles↗

Maintenance of pregnancy in pigs with limited viable embryos.

Interest is increasing in assisted reproductive technologies in the pig involving embryo cryopreservation, cloning, and genetic modification. Although inherently inefficient and variable in their outcome, the successful application of these techniques in this species is confounded by unique mechanisms for pregnancy recognition and maintenance that require a minimum number of viable embryos during a critical window of time early in gestation. These mechanisms require both local and systemic interactions between conceptuses and the maternal reproductive axis. Here, we describe a method wherein cotransfer of parthenogenetic embryos with the capacity for limited development can be used to sustain the pregnancy of fewer than four viable conceptuses to term.

Animals↗

A pollen allergen-encoding gene is expressed in wheat ovaries.

To isolate genes specifically expressed at the initiation of plant embryo development we have applied a sensitive subtractive hybridization technique for three isogenic wheat lines of the so-called 'Salmon system' with either zygotic or autonomous embryo development. Here we present a gene sequence showing a high homology to grass pollen allergens of type II/III thought to be expressed in pollen tissue only. Surprisingly, the pollen allergen-like sequence, designated Tri a III, is also expressed in gynoecia of the sexual, male fertile wheat line '(aestivum)-Salmon', whereas the two parthenogenetic and male sterile wheat lines '(caudata)-Salmon' and '(kotschyi)-Salmon' completely lack any Tri a III transcript. Our data suggest a positive correlation between the expression of this clone and the manifestation of male fertility. Northern and in situ hybridization analysis revealed that, in addition to its presence in pollen, Tri a III is expressed in the parenchymatous tissue of '(aestivum)-Salmon' ovaries exclusively at the day of anthesis. This precise temporal and spatial expression pattern suggests a more general function of the pollen allergen-like sequence Tri a III not limited to the exhibition of allergens in pollen grains.

Allergens↗

Evidence for a link between local and seasonal cycles in gene frequencies and latitudinal gene clines in a cyclic parthenogen.

In an earlier study (Rhombergh et al., Can. J. Genet. Cytol. 27: 224-232, 1985) of natural populations of the cyclic parthenogenetic Rose aphids, Macrosiphum rosae, 6 out of 31 loci were found to be polymorphic and one locus (Esterase-4) showed cyclic seasonal changes in gene and genotypic frequencies. Assuming that the Est-4 polymorphism was balanced and due to some climatic factor that varies seasonally, and realizing that most environmental factors that vary seasonally also vary latitudinally, we predicted existence of a latitudinal gene cline at this locus. In the present study we surveyed four polymorphic loci (chosen to be used as markers) in six geographic populations spanning over 1200 km between the United States and Canada and found all four loci to have latitudinal clines. We think that the gene clines are due to a latitudinal cline in the degree of advancement of local populations through the seasonal cycle, and have called such a pattern a 'seasonal phase cline'. The results are discussed in relation to the temporal instability of local patterns and persistence of genetic variability on the large scale in aphids. It is argued that population structure of aphids makes retention of selectively neutral or weakly selected polymorphisms difficult.

Animals↗

Speciation in the Artemia genus: mitochondrial DNA analysis of bisexual and parthenogenetic brine shrimps.

From the cloned mitochondrial DNAs (mtDNAs) isolated from two bisexual species, one Mediterranean, Artemia salina, and one American, Artemia franciscana, and two parthenogenetic (diploid and tetraploid) strains of Artemia parthenogenetica collected in Spain, physical maps have been constructed and compared. They are extremely different among themselves, much more than the differences between Drosophila melanogaster and D. yakuba and in the same range of different mammalian species such as mouse/rat or man/cow. The nucleotide sequences of two regions of mtDNA encoding parts of the cytochrome c oxidase subunit I (COI) and cytochrome b (Cytb) genes have been determined in the two bisexual species and the two parthenogenetic strains. Comparisons of these sequences have revealed a high degree of divergence at the nucleotide level, averaging more than 15%, in agreement with the differences found in the physical maps. The majority of the nucleotide changes are silent and there is a strong bias toward transitions, with the C<==>T substitutions being highly predominant. The evolutionary distance between the two Artemia parthenogenetica is high and there is no clear relationship with any of the bisexual species, including the one present nowadays in Spain. Using a combination of molecular (mtDNA) and morphological markers it is possible to conclude that all of these Artemia isolates should be actually considered as belonging to different species, even the two Artemia parthenogenetica diploidica and tetraploidica.

Amino Acid Sequence↗

The dependence of annulate lamellae formation on the nucleus in parthenogenetic rabbit eggs.

Previously it has been shown that, in the rabbit, although annulate lamellae (AL) are absent in the follicular oocytes, they appear in the fertilized eggs after the formation of the pronuclei. Furthermore, neither pronuclei nor AL appear when unfertilized eggs are aged in vivo or in vitro. This study was undertaken to determine whether AL formation requires presence of an intact nucleus, or whether the sperm alone contains the stimulatory factors essential to AL synthesis. Rabbit eggs were exposed to 10 degrees C, then incubated for 24 hours. Control eggs were incubated without cold-treatment. Electron microscopic observations indicated that two-thirds of the eggs formed one to two 'pronuclei', or subnuclei. The remainder one-third of the cold-treated eggs and the control eggs failed to form 'pronuclei'. AL were present in large amounts only in those activated eggs (parthenogenones) which formed 'pronuceli.' AL were absent in the control and the non-activated experimental eggs, both of which failed to form a 'pronucleus.' A few small AL were observed in eggs with subnuclei. Condensed fine textured nucleoli appeared precociously during cold-treatment in some eggs and they were present in the 'pronuclei' of activated eggs. It was concluded that the sperm is not necessary for AL formation, but the presence of an intact nucleus is mandatory.

Animals↗

Endoplasmic reticulum whorls as a source of membranes for early cytaster formation in parthenogenetically stimulated sea urchin eggs.

Sea urchin eggs exposed to a continuous hypertonic treatment rapidly form many concentric whorls of endoplasmic reticulum (ER) during the pre-activation period of the parthenogenetic development. These whorls, however, are only a temporary configurational alteration of ER which begin to break up just prior to egg activation. The conversion back to normal vesicles and lamellae occurs not only concurrently with the appearance of early cytastral areas, but also frequently in close association with the formation of these membranous areas. It is revealed here that membrane elements from disrupting whorls may become incorporated into adjacent, developing clear areas, early cytastral areas, and that this ER constitutes an initial major source of membranes for these early astral areas. Having previously suggested that the actual formation of ER whorls occurs in direct response to released intracellular calcium in hypertonic stressed eggs, the new findings, along with other related data and correlations, further suggest that whorl disruption and the formation of associated astral areas can be correlated with a corresponding decrease in the concentration of this released calcium in the cytoplasm.

Animals↗

Reproduction and sexual development in a freshwater gastrotrich. 2. Kinetics and fine structure of postparthenogenic sperm formation.

Spermatogenesis and spermiogenesis in Lepidodermella squammata are confined to the postparthenogenic phase of the life cycle and coincide with developmental changes in the bilateral female gonads. Male stages are bilateral but asynchronous, in the lateral abdomen anterior to the female gonads. Maximum observed sperm production is two packets per side, or 64 sperm. Sperm formation occurs more rapidly at 27 degrees C than at 20 degrees C (p less than 0.001), requiring as little as 1 day. Two spermatogonial mitotic divisions produce a clone of four primary spermatocytes connected by bridges (stage 1). Centrioles are absent. Development occurs within a cyst. Meiotic divisions produce 16 spermatids (stage 2), each containing a dense, elongate, tapered nucleus. Cytoplasmic membranes enclose one end of the nuclear rod, excluding all other organelles. Completion of this process results in stage 3, a packet of 16 sperm associated with one dense sphere, a modified 'residual body' containing cytoplasmic debris. The residual body then disappears, leaving the sperm packet of stage 4. Each mature sperm is a dense nuclear rod with surrounding membranes, lacking acrosome, mitochondrion, centrioles, and flagellum. Function of sperm has not been demonstrated. The spermatozoa are of a reduced type not previously described.

Animals↗

Reproduction and sexual development in a freshwater gastrotrich. 3. Postparthenogenic development of primary oocytes and the X-body.

Six small cells are present in each of the bilateral gonads of parthenogenically reproductive Lepidodermella squammata. Early in the extended postparthenogenic phase of the life history, these cells undergo limited proliferation followed by differentiation. Primary oocytes of three types are present 0.3 days after deposition of the final parthenogenic egg: small oocytes with presynaptic nuclei; intermediate oocytes with nuclei containing synaptonemal complexes; and larger oocytes with a germinal vesicle. Oocytes persist without further development at least until day four of the postparthenogenic phase. Older isolated animals may contain and even deposit an enlarged egg, but successful progeny does not result. Oocytes are located at the anterior pole of each of the bilateral gonads, adjacent to developing male tissues producing sperm. More posterior cells in the gonad are initially undifferentiated in the postparthenogenic phase. Dorsal and central cells first show specialization for secretory activity, and by day four contain peripheral layers of RER and central accumulations of polymorphic secretion droplets. The posterior and ventral cells produce secretion droplets that aggregate into an enlarging bilobed structure called the X-body. Two or three cells in each gonad contribute secretions to the X-body, which is intracellular in a secondary syncytium formed by the contributing cells. Functions for the postparthenogenic gametes and for the X-body are not yet demonstrated.

Animals↗