All snake species exhibit genetic sex determination with the ZZ͞ZW type of sex chromosomes. To investigate the origin and evolution of snake sex chromosomes, we constructed, by FISH, a cytogenetic map of the Japanese four-striped rat snake (Elaphe quadrivirgata) with 109 cDNA clones. Eleven of the 109 clones were localized to the Z chromosome. All human and chicken homologues of the snake Z-linked genes were located on autosomes, suggesting that the sex chromosomes of snakes, mammals, and birds were all derived from different autosomal pairs of the common ancestor. We mapped the 11 Z-linked genes of E. quadrivirgata to chromosomes of two other species, the Burmese python (Python molurus bivittatus) and the habu (Trimeresurus flavoviridis), to investigate the process of W chromosome differentiation. All and 3 of the 11 clones were localized to both the Z and W chromosomes in P. molurus and E. quadrivirgata, respectively, whereas no cDNA clones were mapped to the W chromosome in T. flavoviridis. Comparative mapping revealed that the sex chromosomes are only slightly differentiated in P. molurus, whereas they are fully differentiated in T. flavoviridis, and E. quadrivirgata is at a transitional stage of sex-chromosome differentiation. The differentiation of sex chromosomes was probably initiated from the distal region on the short arm of the protosex chromosome of the common ancestor, and then deletion and heterochromatization progressed on the sex-specific chromosome from the phylogenetically primitive boids to the more advanced viperids.comparative map ͉ chromosome homology ͉ FISH ͉ sex-determining gene ͉ reptile A ll snake species are subject to genetic sex determination with sex chromosomes, as are mammals and birds, and they have female heterogamety (ZZ males and ZW females). Comparative gene mapping between human and chicken revealed that human XX͞XY and chicken ZZ͞ZW sex chromosomes have no homologies (1, 2), suggesting that the sex chromosomes of mammals and birds were derived from different pairs of autosomes of the common ancestor. Beçak et al. (3) found that there is close karyological similarity between snakes and birds, such as distinct differentiation of macro-and microchromosomes and constant occurrence of ZW-type sex chromosomes. This finding leads us to predict the presence of homology between ophidian and avian sex chromosomes. However, no attempts have yet been made to investigate the conservation of the linkage homologies of snake chromosomes to human and chicken chromosomes by comparative gene mapping, although this approach would provide fundamental information on the genome evolution and the origin of sex-chromosome differentiation in amniotes. In another study (4), we constructed a preliminary cytogenetic map of the Japanese four-striped rat snake (Elaphe quadrivirgata) with 52 EST clones, which were isolated from the cDNA library of the brain tissue and were identified as snake homologues of human and chicken orthologous genes by a search of the DNA database. Of 52 EST clones, two genes, T...

Analysis of genomic resources available for cnidarians revealed that several key components of the vertebrate innate immune repertoire are present in representatives of the basal cnidarian class Anthozoa, but are missing in Hydra, a member of the class Hydrozoa, indicating ancient origins for many components of the innate immune system.

The remarkable capability of planarian regeneration is mediated by a group of adult stem cells referred to as neoblasts. Although these cells possess many unique cytological characteristics (e.g. they are X-ray sensitive and contain chromatoid bodies), it has been difficult to isolate them after cell dissociation. This is one of the major reasons why planarian regenerative mechanisms have remained elusive for a long time. Here, we describe a new method to isolate the planarian adult stem cells as X-ray-sensitive cell populations by fluorescence-activated cell sorting (FACS). Dissociated cells from whole planarians were labeled with fluorescent dyes prior to fractionation by FACS. We compared the FACS profiles from X-ray-irradiated and non-irradiated planarians, and thereby found two cell fractions which contained X-ray-sensitive cells. These fractions, designated X1 and X2, were subjected to electron microscopic morphological analysis. We concluded that X-ray-sensitive cells in both fractions possessed typical stem cell morphology: an ovoid shape with a large nucleus and scant cytoplasm, and chromatoid bodies in the cytoplasm. This method of isolating X-ray-sensitive cells using FACS may provide a key tool for advancing our understanding of the stem cell system in planarians.

Recent transcriptome analyses have revealed that a large body of noncoding regions of mammalian genomes are actually transcribed into RNAs. Our understanding of the molecular features of these noncoding RNAs is far from complete. We have identified a novel mRNA-like noncoding gene, named Gomafu, which is expressed in a distinct set of neurons in the mouse nervous system. Interestingly, spliced mature Gomafu RNA is localized to the nucleus despite its mRNA-like characteristics, which usually act as potent export signals to the cytoplasm. Within the nucleus, Gomafu RNA is detected as numerous spots that do not colocalize with known nuclear domain markers. Gomafu RNA is extremely insoluble and remains intact after nuclear matrix preparation. Furthermore, heterokaryon assays revealed that Gomafu RNA does not shuttle between the nucleus and cytoplasm, but is retained in the nucleus after its transcription. We propose that Gomafu RNA represents a novel family of mRNA-like noncoding RNA that constitutes a cell-type-specific component of the nuclear matrix.

Planarians are known for their strong regenerative ability. This ability has been considered to reside in the totipotent somatic stem cell called the "neoblast." Neoblasts contain a unique cytoplasmic structure called the "chromatoid body," which has similar characteristics to the germline granules of germline cells of other animals. The chromatoid bodies decrease in number and size during cytodifferentiation and disappear in completely differentiated cells during regeneration. However, germ cells maintain the chromatoid body during their differentiation from neoblasts. These observations suggest that the chromatoid body is concerned with the totipotency of cells. To understand the molecular nature of the chromatoid body in the neoblast, we focused on vasa (vas)-related genes, since VAS and VAS-related proteins are known to be components of the germline granules in Drosophila and Caenorhabditis elegans. By PCR, two vas-related genes (Dugesia japonica vasa-like gene, DjvlgA and DjvlgB) were isolated, and they were shown to be expressed in germ cells. Interestingly, DjvlgA was also expressed in a number of somatic cells in the mesenchymal space. In regenerating planarians, accumulation of DjvlgA-expressing cells was observed in both the blastema and the blastema-proximal region. In X-ray-irradiated planarians, which had lost regenerative capacity, the number of DjvlgA-expressing cells decreased drastically. These results suggest that the product of DjvlgA may be a component of the chromatoid body and may be involved in the totipotency of the neoblast.