Discovery of genes expressed in Hydra embryogenesis

Hydra's remarkable capacity to regenerate, to proliferate asexually by budding, and to form a pattern de novo from aggregates allows studying complex cellular and molecular processes typical for embryonic development. The underlying assumption is that patterning in adult hydra tissue relies on factors and genes which are active also during early embryogenesis. Previously, we reported that in Hydra the timing of expression of conserved regulatory genes, known to be involved in adult patterning, differs greatly in adults and embryos (Fr?bius, A.C., Genikhovich, G., Kürn, U., Anton-Erxleben, F. and Bosch, T.C.G., 2003. Expression of developmental genes during early embryogenesis of Hydra. Dev. Genes Evol. 213, 445-455). Here, we describe an unbiased screening strategy to identify genes that are relevant to Hydra vulgaris embryogenesis. The approach yielded two sets of differentially expressed genes: one set was expressed exclusively or nearly exclusively in the embryos, while the second set was upregulated in embryos in comparison to adult polyps. Many of the genes identified in hydra embryos had no matches in the database. Among the conserved genes upregulated in embryos is the Hydra orthologue of Embryonic Ectoderm Development (HyEED). The expression pattern of HyEED in developing embryos suggests that interstitial stem cells in Hydra originate in the endoderm. Importantly, the observations uncover previously unknown differences in genes expressed by embryos and polyps and indicate that not only the timing of expression of developmental genes but also the genetic context is different in Hydra embryos compared to adults.     

Geographic ranges, population structure, and ages of sexual and parthenogenetic snail lineages

Abstract A sexual reproduction is thought to doom organisms to extinction due to mutation accumulation and parasite exploitation. Theoretical models suggest that parthenogens may escape the negative effects of conspecific and biological enemiecs through escape in space. Through intensive sequencing of a mitochondrial DNA (mtDNA) and a nuclear intron locus in sexual and pathenogenetic freshwater snails (Campelom), I examine three questionss: (1) Are sexual mtDNA lineage more restricted geographically than parthenogenetic mtDNA lineages? (2) Are independent pathenogenetic lineages shorter lived than sexual lineages? (3) Do pathenogens have higher intraindividual nuclear sequence diversity and form well‐differentiated monophyletic groups as expected under the Meselson effect? Geographic ranges of parthenogenetic lineages are significantly larger than geographic ranges of sexual lineages. Based on coalescence times under different deographic assumptions, asexual lineages are short lived, but there is variation in clonal ages. Although alternative explanations exit, these results suggest that asexual lineages may persist in the short term through dispersal, and that various constraints may cause geographic restriction of sexual lineagess. Both allotriploid and diploid Campleloma parthenogens have significantly higher allelic divergence within individuals, but show limited nuclear sequence divergence from sexual ancestors. In contrast to previous allozyme evidence for nonhybrid origins of diploid Campeloma parthenogens, cryptic hybridization may account for elevated heterozygosity.     

The evolution of California's wild radish has resulted in the extinction of its progenitors

If two previously isolated taxa mutually assimilate through hybridization and subsequent biparental introgression, and if their introgressed descendants have the same or higher fitness than their parents, then gene flow should result in the local extinction of parental taxa via replacement by hybrid derivatives. These dramatic events may occur rapidly, even in a few generations. Given the speed at which such extinction by hybridization may occur, it may be difficult to identify that the process has occurred. Thus, documented instances of extinction by hybridization are rare, and especially so for cases in which both parents are replaced by the hybrid lineage. Here we report morphological and allozyme evidence for the local extinction of two Raphanus species in California via replacement by their hybrid-derived descendants. The results from a greenhouse experiment demonstrate that California wild radishes have a specific combination of traits from their progenitors, and comparison of our results to that of an earlier report indicate that pure parental types are no longer present in the wild. Our results also show the hybrid-derived lineage has transgressive fruit weight compared to its parents. Allozyme analysis demonstrates that California wild radishes are derived from hybridization between the putative parental species. However, that analysis also demonstrates that California wild radish has now become an evolutionary entity separate from both of its parents. We suggest that the aggressive colonizing behavior of the hybrid-derived lineage probably results from a novel combination of parental traits, rather than genetic variability of the population per se.     

Meiosis Aspects and Nucleolar Activity in Triatoma vitticeps (Triatominae, Heteroptera)

Some aspects of both the nucleolar organizer activity and meiosis were studied in the testes of Triatoma vitticeps (Heteroptera, Triatominae). The techniques used included squashing followed by lacto-acetic orcein staining, silver-ion impregnation, fluorescent banding (CMA₃, Quinacrine mustard and DAPI) and fluorescent in situ hybridization (FISH). A close relationship between heterochromatin and nucleolus in testicular cells was observed. During meiosis, the silver-ion impregnation pattern varied. At metaphase plate, a small body appeared apart from the chromosomes. In the spermatids this small body was seen in preparations stained with orcein and silver- ion impregnation but not with fluorochromes or FISH. These characteristics combined suggest that these corpuscles represent a source of ribonucleoproteins (RNP) – RNA and specific nucleolar proteins. Silver-ion impregnation and (FISH) revealed nucleolar organizer activity in two metaphase sex chromosomes (X). These results indicate that, in these species, nucleolar organizer regions (NORs) are located in the sex chromosomes, X chromosomes were CMA₃⁺ and Y chromosome was DAPI⁺.     

Genetic Relationships Between Brazilian Species of Molossidae and Phyllostomidae (Chiroptera, Mammalia)

A comparative analysis of G-banded karyotypes was performed for seven species of Chiroptera, representing two families (Phyllostomidae and Molossidae). Despite the differences in diploid and fundamental numbers, extensive homologies between six karyotypes were identified: A . planirostris, P. lineatus, S. lilium, G. soricina, P. hastatus (Phyllostomidae) and M. rufus (Molossidae). Robertsonian rearrangements and pericentric inversions account for the differences between the karyotypes of phyllostomid and molossid species. The homologies and rearrangements observed reinforce the monophiletic origin of phyllostomids and the inclusion of species in different subfamilies. In situ hybridization with genomic DNA revealed considerable conservation of the karyotypes, including C. perspicillata, that did not show G-band homologies with the other species analyzed. For the first time, chromosomal evidence is presented of a common origin for Phyllostomidae and Molossidae.     

Tryptophan- and arginine-rich antimicrobial peptides: Structures and mechanisms of action

Antimicrobial peptides encompass a number of different classes, including those that are rich in a particular amino acid. An important subset are peptides rich in Arg and Trp residues, such as indolicidin and tritrpticin, that have broad and potent antimicrobial activity. The importance of these two amino acids for antimicrobial activity was highlighted through the screening of a complete combinatorial library of hexapeptides. These residues possess some crucial chemical properties that make them suitable components of antimicrobial peptides. Trp has a distinct preference for the interfacial region of lipid bilayers, while Arg residues endow the peptides with cationic charges and hydrogen bonding properties necessary for interaction with the abundant anionic components of bacterial membranes. In combination, these two residues are capable of participating in cation-π interactions, thereby facilitating enhanced peptide-membrane interactions. Trp sidechains are also implicated in peptide and protein folding in aqueous solution, where they contribute by maintaining native and nonnative hydrophobic contacts. This has been observed for the antimicrobial peptide from human lactoferrin, possibly restraining the peptide structure in a suitable conformation to interact with the bacterial membrane. These unique properties make the Arg- and Trp-rich antimicrobial peptides highly active even at very short peptide lengths. Moreover, they lead to structures for membrane-mimetic bound peptides that go far beyond regular α-helices and β-sheet structures. In this review, the structures of a number of different Trp- and Arg-rich antimicrobial peptides are examined and some of the major mechanistic studies are presented.     

Association of fragile site-associated (FSA) gene expression with epithelial differentiation and tumor development

A novel gene designated as fragile site-associated (FSA) gene was recently identified by positional cloning from the CHO 1q31 fragile site which plays an important role in regulating amplification of multidrug resistance (mdr1) gene in multidrug-resistant cells. FSA produces a message of approximately 16 kb which encodes an open-reading frame of 5005 amino acids. FSA shares sequence similarity with that in Caenorhabditis elegans lpd-3, a lipid storage gene. Using immunohistochemical staining and RNA in situ hybridization we report here that expression of FSA is associated with developmental programs of spermatogenesis and mammary gland in mice. Real-time RT-PCR results also support the upregulation of FSA expression in mammary gland development. Expression of FSA in many tissues including colon, skin, ovary, prostate, and bladder is mainly in the postmitotic, well-differentiated compartments. Moreover, levels of FSA expression are downregulated in tumors of these tissue origins. These results suggest that FSA also plays important roles in regulating mammalian epithelial growth and differentiation and tumor development.     

Steroid structural requirements for interaction of ostreolysin, a lipid-raft binding cytolysin, with lipid monolayers and bilayers

Ostreolysin, a cytolytic protein from the edible oyster mushroom (Pleurotus ostreatus), recognizes and binds specifically to membrane domains enriched in cholesterol and sphingomyelin (or saturated phosphatidylcholine). These events, leading to permeabilization of the membrane, suggest that a cholesterol-rich liquid-ordered membrane phase, which is characteristic of lipid rafts, could be its possible binding site. In this work, we present effects of ostreolysin on membranes containing various steroids. Binding and membrane permeabilizing activity of ostreolysin was studied using lipid mono- and bilayers composed of sphingomyelin combined, in a 1/1 molar ratio, with natural and synthetic steroids (cholesterol, ergosterol, β-sitosterol, stigmasterol, lanosterol, 7-dehydrocholesterol, cholesteryl acetate, and 5-cholesten-3-one). Binding to membranes and lytic activity of the protein are both shown to be dependent on the intact sterol 3β-OH group, and are decreased by introducing additional double bonds and methylation of the steroid skeleton or C17-isooctyl chain. The activity of ostreolysin mainly correlates with the ability of the steroids to promote formation of liquid-ordered membrane domains, and is the highest with cholesterol-containing membranes. Furthermore, increasing the cholesterol concentration enhanced ostreolysin binding in a highly cooperative manner, suggesting that the membrane lateral distribution and accessibility of the sterols are crucial for the activity of this new member of cholesterol-dependent cytolysins.