Hybrid crosses and the genetic basis of interspecific divergence in lifespan in Pristionchus nematodes.

Characterizing the genetic basis of among-species variation in lifespan is a major goal of evolutionary gerontology research, but the very feature that defines separate species - the inability to interbreed - makes achieving this goal impractical, if not impossible, for most taxa. Pristionchus nematodes provide an intriguing system for tackling this problem, as female lifespan varies among species that can be crossed to form viable (although infertile) hybrids. By conducting reciprocal crosses among three species - two dioecious (long-lived Pristionchus exspectatus and short-lived Pristionchus arcanus) and one androdioecious (short-lived Pristionchus pacificus) - we found that female lifespan was long for all hybrids, consistent with the hypothesis that the relatively short lifespans seen for P. pacificus hermaphrodites and P. arcanus females are caused by independent, recessive alleles that are masked in hybrid genomes. Cross-direction had a small effect on survivorship for crosses involving P. exspectatus, indicating that nuclear-mitochondrial interactions may also influence Pristionchus longevity. Our findings suggest that long lifespan in P. exspectatus reflects the realization of an ancestral potential for extended longevity in the P. pacificus species complex. This work demonstrates the utility of interspecific hybrids for ageing research and provides a foundation for future work on the genetic architecture of interspecific lifespan variation.

An integrated physical and genetic map of the nematode Pristionchus pacificus.

The free-living nematode Pristionchus pacificus is one of several species that have recently been developed as a satellite system for comparative functional studies in evolutionary developmental biology. Comparisons of developmental processes between P. pacificus and the well established model organism Caenorhabditis elegans at the cellular and genetic levels provide detailed insight into the molecular changes that shape evolutionary transitions. To facilitate genetic analysis and cloning of mutations in P. pacificus, we previously generated a BAC-based genetic linkage map for this organism. Here, we describe the construction of a physical map of the P. pacificus genome based on AFLP fingerprint analysis of 7747 BAC clones. Most of the SSCP markers used to generate the genetic linkage map were derived from BAC ends, so that the physical genome map and the genetic map can be integrated. The contigs that make up the physical map are evenly distributed over the genetic linkage map and no clustering is observed, indicating that the physical map provides a valid representation of the P. pacificus genome. The integrated genome map thus provides a framework for positional cloning and the study of genome evolution in nematodes.

As good as they get: cells in nematode vulva development and evolution.

Cells are important modules of biological systems, and many evolutionary alterations involve changes in cell determination and cell proliferation. Genetic and molecular comparisons of nematode vulva development between Caenorhabditis, Pristionchus and Oscheius indicate that although the vulva is a stable organ, cell determination and proliferation change dramatically during nematode evolution.

Formation of the egg-laying system in Pristionchus pacificus requires complex interactions between gonadal, mesodermal and epidermal tissues and does not rely on single cell inductions.

The invariant cell lineage of nematodes allows the formation of organ systems, like the egg-laying system, to be studied at a single cell level. The Caenorhabditis elegans egg-laying system is made up of the vulva, the mesodermal gonad and muscles and several neurons. The gonad plays a central role in patterning the underlying ectoderm to form the vulva and guiding the migration of the sex myoblasts to their final position. In Pristionchus pacificus, the egg-laying system is homologous to C. elegans, but comparative studies revealed several differences at the cellular and molecular levels during vulval formation. For example, the mesoblast M participates in lateral inhibition, a process that influences the fate of two vulval precursor cells. Here, we describe the M lineage in Pristionchus and show that both the dorsal and ventral M sublineages are involved in lateral inhibition. Mutations in the homeotic gene Ppa-mab-5 cause severe misspecification of the M lineage, resembling more the C. elegans Twist than the mab-5 phenotype. Ectopic differentiation of P8.p in Ppa-mab-5 results from at least two separate interactions between M and P8.p. Thus, interactions among the Pristionchus egg-laying system are complex, involving multiple cells of different tissues occurring over a distance.

Functional comparison of the nematode Hox gene lin-39 in C. elegans and P. pacificus reveals evolutionary conservation of protein function despite divergence of primary sequences.

Hox transcription factors have been implicated in playing a central role in the evolution of animal morphology. Many studies indicate the evolutionary importance of regulatory changes in Hox genes, but little is known about the role of functional changes in Hox proteins. In the nematodes Pristionchus pacificus and Caenorhabditis elegans, developmental processes can be compared at the cellular, genetic, and molecular levels and differences in gene function can be identified. The Hox gene lin-39 is involved in the regulation of nematode vulva development. Comparison of known lin-39 mutations in P. pacificus and C. elegans revealed both conservation and changes of gene function. Here, we study evolutionary changes of lin-39 function using hybrid transgenes and site-directed mutagenesis in an in vivo assay using C. elegans lin-39 mutants. Our data show that despite the functional differences of LIN-39 between the two species, Ppa-LIN-39, when driven by Cel-lin-39 regulatory elements, can functionally replace Cel-lin-39. Furthermore, we show that the MAPK docking and phosphorylation motifs unique for Cel-LIN-39 are dispensable for Cel-lin-39 function. Therefore, the evolution of lin-39 function is driven by changes in regulatory elements rather than changes in the protein itself.

Microevolutionary analysis of the nematode genus Pristionchus suggests a recent evolution of redundant developmental mechanisms during vulva formation.

To identify the mechanisms by which molecular variation is introduced into developmental systems, microevolutionary approaches to evolutionary developmental biology have to be taken. Here, we describe the molecular and developmental characterization of laboratory strains of the nematode genus Pristionchus, which lays a foundation for a microevolutionary analysis of vulva development. We describe 13 laboratory strains of the Pristionchus genus that are derived from natural isolates from around the world. Mating experiments and ITS sequence analysis indicated that these 13 strains represent four different species: the gonochoristic species P. Iheritieriand three hermaphroditic species, P. pacificus, P. maupasi, and an as yet undescribed species Pristionchus sp., respectively. P. pacificus is represented by five different strains isolated from California, Washington, Hawaii, Ontario, and Poland. Developmental differences during vulva formation are observed between strains from different species but also between strains of P. pacificus, like the strains from California and Poland. In particular, redundant developmental mechanisms present during vulva formation in P. pacificus var. California are absent in other strains. Amplified restriction fragment length polymorphism (AFLP) analyses of the P. pacificus strains revealed that the American strains are highly polymorphic. In contrast, the developmentally distinct strain from Poland is identical to the Californian strain, suggesting that the developmental differences rely on a small number of changes in developmental control genes rather than the accumulation of changes at multiple loci.

The nematode even-skipped homolog vab-7 regulates gonad and vulva position in Pristionchus pacificus.

In free-living nematodes, developmental processes like the formation of the vulva, can be studied at a cellular level. Cell lineage and ablation studies have been carried out in various nematode species and multiple changes in vulval patterning have been identified. In Pristionchus pacificus, vulva formation differs from Caenorhabditis elegans with respect to several autonomous and conditional aspects of cell fate specification. To understand the molecular basis of these evolutionary changes, we have performed a genetic analysis of vulva formation in P. pacificus. Here, we describe two mutants where the vulva is shifted posteriorly, affecting which precursor cells will form vulval tissue in P. pacificus. Mutant animals show a concomitant posterior displacement of the gonadal anchor cell, indicating that the gonad and the vulva are affected in a similar way. We show that mutations in the even-skipped homolog of nematodes, vab-7, cause these posterior displacements. In addition, cell ablation studies in the vab-7 mutant indicate that the altered position of the gonad not only changes the cell fate pattern but also the developmental competence of vulval precursor cells. Investigation of Cel-vab-7 mutant animals showed a similar but weaker vulva defective phenotype to the one described for Ppa-vab-7.

Comparative genetics: a third model nematode species.

Recent studies have introduced Oscheius sp. CEW1 as a third nematode species accessible to genetic analysis, joining the better known Caenorhabditis elegans and Pristionchus pacificus. A group of vulva-defective mutants in Oscheius has been identified, with defects not seen in C. elegans.

Impact of low body weight on frequency of pediatric cardiac catheterization complications.

The overall risk of pediatric cardiac catheterization remains low despite the enormous new complexity and potential for complications brought on by the growth of interventional catheterization techniques. For all patients aged < 21 years, balloon interventions carry the highest risk, diagnostic procedures carry more risk than non-balloon interventions, and although weight < or = 5 kg is a significant risk factor for complications, irrespective of the type of procedure performed, weight < or = 2.5 kg did not alter that risk.

Novel cell-cell interactions during vulva development in Pristionchus pacificus.

Vulva development differs between Caenorhabditis elegans and Pristionchus pacificus in several ways. Seven of 12 ventral epidermal cells in P. pacificus die of apoptosis, whereas homologous cells in C. elegans fuse with the hypodermal syncytium. Vulva induction is a one-step process in C. elegans, but requires a continuous interaction between the gonad and the epidermis in P. pacificus. Here we describe several novel cell-cell interactions in P. pacificus, focusing on the vulva precursor cell P8.p and the mesoblast M. P8.p in P. pacificus, unlike its homologous cell in C. elegans, is incompetent to respond to gonadal signaling in the absence of other vulva precursor cells, but can respond to lateral signaling from a neighboring vulval precursor. P8.p provides an inhibitory signal that determines the developmental competence of P(5,7).p. This lateral inhibition acts via the mesoblast M and is regulated by the homeotic gene Ppa-mab-5. In Ppa-mab-5 mutants, M is misspecified and provides inductive signaling to the vulval precursor cells, including P8.p. Taken together, vulva development in P. pacificus displays novel cell-cell interactions involving the mesoblast M and P8.p. In particular, P8.p represents a new ventral epidermal cell type, which is characterized by novel interactions and a specific response to gonadal signaling.

Evolution of nematode development.

Multiple evolutionary variations occur in the cellular and genetic programming of nematode development. Many changes involve alterations of inductive interactions. Surprisingly, inductive processes vary during evolution, irrespective of changes in the final cell lineages and morphological structures. Genetic studies in some nematodes also shed light on the underlying mechanisms of evolutionary change.

Gestational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin induces developmental defects in the rat vagina.

At puberty, female rats exposed in utero to 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) exhibit a persistent thread of mesenchymal tissue surrounded by keratinized epithelium that partially occludes the vaginal opening. Our objective was to determine the earliest time during fetal development that morphological signs of this vaginal canal malformation could be detected and to obtain greater insight into mechanisms involved in this effect. Pregnant rats were administered a single dose of vehicle (control) or TCDD (1.0 microg/kg, po) on gestation day (GD) 15 and were sacrificed on GD 18, 19, 20, and 21 for histological evaluation of female. Gestational exposure to TCDD affected vaginal morphogenesis as early as GD 19, 4 days after exposure of pregnant dams. In exposed fetuses, the thickness of mesenchymal tissue between the caudal Mullerian ducts was increased, which resulted in a failure of the Mullerian ducts to fuse, a process normally completed prior to parturition. In addition, TCDD exposure appeared to inhibit the regression of Wolffian ducts. Thus, TCDD interferes with vaginal development by impairing regression of the Wolffian ducts, by increasing the size of interductal mesenchyme, and by preventing fusion of the Mullerian ducts. Taken together, these effects appear to cause the persistent vaginal thread defect observed in rats following in utero and lactational TCDD exposure.

Evolution of vulva development in the Cephalobina (Nematoda).

Ventral cord and vulva development are analyzed in a large sample of nematode species of the suborder Cephalobina. We find a specific range of evolutionary variations at distinct developmental steps. (1) Unlike Caenorhabditis elegans and relatives, the vulva is formed from the four precursor cells P(5-8).p or, exceptionally, from P(6, 7).p only. (2) The vulval competence group is restricted to these four cells or is larger. (3) The fates of more anterior and posterior Pn.p cells vary between closely related species (mostly cell death versus epidermal fate). (4) The mechanism of vulval cell fate patterning varies within a single genus, even between strains of the same species. (5) We describe the first example of a vulval cell lineage that is asymmetric between the anterior and the posterior sides of the vulva. For a selection of the investigated taxa, phylogenetic trees were constructed in order to map vulval characters and infer evolutionary polarities. We can conclude that in this group, death of the Pn.p cells probably constitutes a derived character state compared to a syncytial fate. Rhabditophanes sp. and Strongyloides ratti are placed as sister taxa, probably sharing an exclusive common ancestor in which the number of precursor cells forming the vulva was reduced from four to two.

Familial patent ductus arteriosus and bicuspid aortic valve with hand anomalies: a novel heart-hand syndrome.

The association between cardiac and limb defects, particularly those affecting the hand, has been well documented by the delineation of several heart-hand syndromes. Based on observations with a three-generation family with seven affected individuals, we describe a novel heart-hand syndrome comprising patent ductus arteriosus, bicuspid aortic valve, 5th metacarpal hypoplasia, and brachydactyly. The inheritance pattern was consistent with autosomal dominance, although X-linked dominance could not be excluded. Penetrance appeared to be complete, but there was variability of the cardiac and hand phenotypes. Because this new syndrome closely resembled Char syndrome (patent ductus arteriosus, 5th finger middle phalangeal hypoplasia, and minor facial anomalies), multipoint linkage analysis was performed using polymorphic DNA markers spanning the recently identified Char syndrome critical region at chromosomal bands 6p12-p21.1. This analysis formally excluded this 3-cM region, documenting that the two traits are not allelic. In sum, a novel heart-hand syndrome involving left ventricular outflow and aortic arch as well as an ulnar ray derivative has been identified. Because the hand anomalies can be subtle, thorough evaluation is suggested for families inheriting these cardiac defects as a mendelian trait.

Pristionchus pacificus, a nematode with only three juvenile stages, displays major heterochronic changes relative to Caenorhabditis elegans.

The nematode Pristionchus pacificus (Diplogastridae) has been described as a satellite organism for a functional comparative approach to the model organism Caenorhabditis elegans because genetic, molecular, and cell-biological tools can be used in a similar way in both species. Here we show that P. pacificus has three juvenile stages, instead of the usual four found in other nematodes. Embryogenesis is lengthened and many developmental events that take place during the first juvenile stage in C. elegans occur during late embryogenesis in P. pacificus. Video imaging and transmission electron microscopy revealed no embryonic moult. The timing of later developmental events relative to the moults differs between P. pacificus and C. elegans. In addition, the post-embryonic blast-cell divisions display a specific change in timing between the two species, resulting in heterochrony between different cell lineages, such as vulval and gonadal lineages. Developmental events appear to come into register during the last larval stage. Thus, differences in developmental timing between P. pacificus and C. elegans represent a deep heterochronic change. We designate the three juvenile stages of P. pacificus as J1 to J3. Comparison with other species of the family Diplogastridae indicates that this pattern represents an apomorphic character for the monophylum Diplogastridae.

Vulva formation in Pristionchus pacificus relies on continuous gonadal induction.

One of the best known features of vulva development in Caenorhabditis elegans is the induction of vulval precursor cells by the gonadal anchor cell. Induction is crucial for the initiation of pattern formation within the C. elegans vulva equivalence group, and it is therefore surprising to find that this aspect of vulva formation, in particular, varies greatly among nematodes. In some species which form vulvae in the posterior body region, no gonadal signal is necessary for vulva induction. In other nematodes, such as Panagrolaimus, Oscheius, and Rhabditella, vulva formation depends on two temporally distinct gonadal inductions which specify the different cell fates. Here we report our analysis of vulva induction in Pristionchus pacificus, a species which has recently been used as a genetic system to analyze the evolution of vulva development. Cell ablation studies in P. pacificus show that another mode of vulva induction exists. P. pacificus vulva formation depends on a continuous gonadal induction that starts several hours after hatching and continues until the birth of the anchor cell, some 20 h later. Mutations defective in gonadal induction result in the absence of vulva differentiation, suggesting that only one signaling system is involved in the gonadal-epidermal interaction. This new mode adds further to the great variety of gonadal inductions among nematode species.

The snare-assisted technique for transcatheter coil occlusion of moderate to large patent ductus arteriosus: immediate and intermediate results.

OBJECTIVES: The purpose of this study was to evaluate the feasibility, safety and efficacy of using a snare-assisted technique to coil occlude the moderate to large size patent ductus arteriosus (PDA). BACKGROUND: Transcatheter occlusion of small PDAs using Gianturco coils is safe and effective. However, in larger size PDAs and/or those with short PDA length, the procedure still carries risks of coil embolization, incomplete occlusion and failure to implant the coil. METHODS: From January 1994 to June 1997, the records of 104 consecutive snare-assisted coil occlusions of moderate to large PDAs (minimum diameter >2.0 mm) were reviewed. Immediate and intermediate outcomes including complete and partial occlusion, failure to implant and complications were analyzed with respect to ductal type and size. RESULTS: Patient age ranged from 0.1 to 70.1 years (median 3.3 years). Minimum PDA diameter ranged from 2.1 to 6.8 mm (mean 3.0 +/- 0.9 mm). Angiographic types were A-62, B-13, C-6, D-14 and E-9. Using the snare-assisted technique, coil placement was successful in 104/104 patients (100%), irrespective of size or angiographic type. Immediate complete closure was observed in 73/104 (70.2%) and was related to smaller PDA size, but not to angiographic type. Complete closure was documented in 102/104 (98.1%) at 2- to 16-month follow-up. Successful closure was unrelated to PDA size or type. Coil embolization to the pulmonary artery occurred in 3/104 (2.9%) patients and was not related to PDA size or type. The need for multiple coils was found in 28/104 patients (26.9%), and was related to larger PDA size, but not to angiographic type. CONCLUSIONS: The snare-assisted delivery technique allows successful occlusion of moderate to large PDAs up to 6.8 mm, irrespective of angiographic type. This technique permits improved control and accuracy of coil placement, and facilitates delivery of multiple coils.

Evolutionary change in the functional specificity of genes.

Species throughout the animal kingdom share not only housekeeping but also many key regulatory genes. Nonetheless, species differ from one another developmentally and thus, also morphologically. One of the general aims of comparative developmental genetics is to understand how similar molecules can generate the known diversity of biological form. Here, we argue that gene function can change in different ways during the evolution of developmental processes. Genes can be recruited to serve completely new functions in a new regulatory linkage (co-option), they can change their molecular specificity while remaining in the original (homologous) developmental program and can, at the same time, retain other functions. We describe evidence for such evolutionary patterns based on the comparison of loss-of-function mutations of homologous genes of the two free-living nematodes Caenorhabditis elegans and Pristionchus pacificus. Ultimately, it is the interplay of conservation and change of the specificity of genes and genetic networks that generates developmental novelty over evolutionary time.

Ah receptor and ARNT protein and mRNA concentrations in rat prostate: effects of stage of development and 2,3,7, 8-tetrachlorodibenzo-p-dioxin treatment.

Effects of stage of development and 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD) exposure on aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (ARNT) protein concentrations in reproductive organs of male rats were determined. AhR protein levels in developing rat ventral and dorsolateral prostate decreased with age, declining approximately 70% between Postnatal Days (PND) 1 and 21. ARNT protein levels also decreased with age in dorsolateral, but not ventral prostate. The developmental decreases in prostatic AhR and ARNT protein were associated with decreases in AhR and ARNT mRNA. AhR and ARNT protein concentrations in fetal urogenital sinus on Gestation Days (GD) 16, 18, and 20 were similar to levels in ventral prostate on PND 7. TCDD exposure of adult male rats (0.2, 1, 5, or 25 micrograms/kg po, 24 h) decreased AhR but not ARNT protein in ventral and dorsolateral prostate, vas deferens, and epididymis. In utero and lactational TCDD exposure (1.0 micrograms/kg dam po, GD 15) did not alter ARNT levels but reduced prostatic AhR protein levels on PND 7 and delayed the developmental decrease in AhR protein in ventral and dorsolateral prostate. Finally, pretreatment of rat pups for 24 h with TCDD (5 micrograms/kg ip) down-regulated prostatic AhR protein on PND 7, but not on PND 1. Thus, prostatic AhR and ARNT protein and mRNA levels are regulated with age, whereas only AhR protein concentration is altered by TCDD exposure. Because in utero and lactational TCDD exposure only decreased prostatic AhR on PND 7, it is unlikely that down-regulation of AhR is the mechanism by which perinatal TCDD exposure impairs prostate development.