Finding clues to the riddle of sex determination in zebrafish

How sex is determined has been one of the most intriguing puzzles in biology since antiquity. Although a fundamental process in most metazoans, there seems to be myriad of ways in which sex can be determined – from genetic to environmental sex determination. This variation is limited mainly to upstream triggers with the core of sex determination pathway being conserved. Zebrafish has gained prominence as a vertebrate model system to study development and disease. However, very little is known about its primary sex determination mechanism. Here we review our current understanding of the sex determination in zebrafish. Zebrafish lack identifiable heteromorphic sex chromosomes and sex is determined by multiple genes, with some influence from the environment. Recently, chromosome 4 has been identified as sex chromosome along with few sex-linked loci on chromosomes 5 and 16. The identities of candidate sex-linked genes, however, have remained elusive. Sex in zebrafish is also influenced by the number of meiotic oocytes in the juvenile ovary, which appear to instruct retention of the ovarian fate. The mechanism and identity of this instructive signal remain unknown. We hypothesize that sex in zebrafish is a culmination of combinatorial effects of the genome, germ cells and the environment with inputs from epigenetic factors translating the biological meaning of this interaction.     

Cystathionine-γ-lyase gene silencing with siRNA in monocytes/macrophages attenuates inflammation in cecal ligation and puncture-induced sepsis in the mouse

Hydrogen sulphide is an endogenous inflammatory mediator produced by cystathionine-γ-lyase (CSE) in macrophages. To determine the role of H₂S and macrophages in sepsis, we used small interference RNA (siRNA) to target the CSE gene and investigated its effect in a mouse model of sepsis. Cecal ligation puncture (CLP)-induced sepsis is characterized by increased levels of myeloperoxidase (MPO) activity, morphological changes in liver and pro-inflammatory cytokines and chemokines in the liver and lung. SiRNA treatment attenuated inflammation in the liver and lungs of mice following CLP-induced sepsis. Liver MPO activity increased in CLP-induced sepsis and treatment with siRNA significantly reduced this. Similarly, lung MPO activity increased following induction of sepsis with CLP while siRNA treatment significantly reduced MPO activity. Liver and lung cytokine and chemokine levels in CLP-induced sepsis reduced following treatment with siRNA. These findings show a crucial pro-inflammatory role for H₂S synthesized by CSE in macrophages in sepsis and suggest CSE gene silencing with siRNA as a potential therapeutic approach for this condition.     

Expressions of visual pigments and synaptic proteins in neonatal chick retina exposed to light of variable photoperiods

Light causes damage to the retina, which is one of the supposed factors for age-related macular degeneration in human. Some animal species show drastic retinal changes when exposed to intense light (e.g. albino rats). Although birds have a pigmented retina, few reports indicated its susceptibility to light damage. To know how light influences a cone-dominated retina (as is the case with human), we examined the effects of moderate light intensity on the retina of white Leghorn chicks (Gallus g. domesticus). The newly hatched chicks were initially acclimatized at 500 lux for 7 days in 12 h light: 12 h dark cycles (12L:12D). From posthatch day (PH) 8 until PH 30, they were exposed to 2000 lux at 12L:12D, 18L:6D (prolonged light) and 24L:0D (constant light) conditions. The retinas were processed for transmission electron microscopy and the level of expressions of rhodopsin, S- and L/M cone opsins, and synaptic proteins (Synaptophysin and PSD-95) were determined by immunohistochemistry and Western blotting. Rearing in 24L:0D condition caused disorganization of photoreceptor outer segments. Consequently, there were significantly decreased expressions of opsins and synaptic proteins, compared to those seen in 12L:12D and 18L:6D conditions. Also, there were ultrastructural changes in outer and inner plexiform layer (OPL, IPL) of the retinas exposed to 24L:0D condition. Our data indicate that the cone-dominated chick retina is affected in constant light condition, with changes (decreased) in opsin levels. Also, photoreceptor alterations lead to an overall decrease in synaptic protein expressions in OPL and IPL and death of degenerated axonal processes in IPL.     

Triphala,a formulation of traditional Ayurvedic medicine,shows protective effect against X-radiation in HeLa cells

Ayurveda is a holistic medical system of traditional medicine, and Triphala is one of the most popular formulations in Ayurveda. Triphala is composed of three kinds of herb, Terminalia chebula, Terminalia bellirica, and Emblica officinalis. Since Triphala is shown to exhibit a protective activity against ionizing radiation in mice, we investigated its activity in HeLa cells. We found that Triphala showed the protective effects against X-radiation and bleomycin, both of which generate DNA strand breaks, in HeLa cells. Further, Triphala efficiently eliminated reactive oxygen species (ROS) in HeLa cells. Thus, the antioxidant activity of Triphala would likely play a role in its protective actions against X-radiation and bleomycin because both agents damage DNA through the generation of ROS. These observations suggested that the radioprotective activity of Triphala can be, at least partly, studied with the cells cultured in vitro. The simple bioassay system with human cultured cells would facilitate the understanding of the molecular basis for the beneficial effects of Triphala.     

Indomethacin treatment reduces microglia activation and increases numbers of neuroblasts in the subventricular zone and ischaemic striatum after focal ischaemia

Neuroblasts from the subventricular zone (SVZ) migrate to striatum following stroke, but most of them die in the ischaemic milieu and this can be related to exacerbated microglial activation. Here, we explored the effects of the non-steroidal anti-inflammatory indomethacin on microglial activation, neuronal preservation and neuroblast migration following experimental striatal stroke in adult rats. Animals were submitted to endothelin-1 (ET-1)-induced focal striatal ischaemia and were treated with indomethacin or sterile saline (i.p.) for 7 days, being perfused after 8 or 14 days. Immunohistochemistry was performed to assess neuronal loss (anti-NeuN), microglial activation (anti-Iba1, ED1) and migrating neuroblasts (anti-DCX) by counting NeuN, ED1 and DCX-positive cells in the ischaemic striatum or SVZ. Indomethacin treatment reduced microglia activation and the number of ED1⁺ cells in both 8 and 14 days post injury as compared with controls. There was an increase in the number of DCX⁺ cells in both SVZ and striatum at the same survival times. Moreover, there was a decrease in the number of NeuN⁺ cells in indomethacin-treated animals as compared with the control group at 8 days but not after 14 days post injury. Our results suggest that indomethacin treatment modulates microglia activation, contributing to increased neuroblast proliferation in the SVZ and migration to the ischaemic striatum following stroke.     

The rice <Emphasis Type="Italic">OsSAG12-2</Emphasis> gene codes for a functional protease that negatively regulates stress-induced cell death

Senescence is the final stage of plant development. Although expression of most of the genes is suppressed during senescence, a set of genes referred as senescence-associated genes (SAGs) is induced. Arabidopsis thaliana SAG12 (AtSAG12) is one such gene that has been mostly studied for its strict association with senescence. AtSAG12 encodes a papain-like cysteine protease, expressed predominantly in senescence-associated vacuoles. Rice genome contains multiple AtSAG12 homologues (OsSAGs). OsSAG12-1, the closest structural homologue of AtSAG12, is a negative regulator of developmental and stress-induced cell death. Proteolytic activity has not been established for any SAG12 homologues in vitro. Here, we report that OsSAG12-2, the second structural homologue of AtSAG12 from rice, codes for a functional proteolytic enzyme. The recombinant OsSAG12-2 protein produced in Escherichia coli undergoes autolysis to generate a functional protease. The matured OsSAG12-2 protein shows 27% trypsin-equivalent proteolytic activity on azocasein substrate. Dark-induced senescence activates OsSAG12-2 expression. Down-regulation of OsSAG12-2 in the transgenic artificial miRNA lines results in enhanced salt- and UV-induced cell death, even though it does not affect cell viability in the stress-free condition. Our results show that OsSAG12-2 codes for a functional protease that negatively regulates stress-induced cell death in rice.     

Desiccation stress induces developmental heterochrony in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Stressful environments are known to perturb developmental patterns in insects. In the purview of desiccation as a stressor, relatively little is known about the developmental consequences linked with desiccation tolerance. In this study, we have particularly focused on the exploration of the temporal profile of postembryonic development in response to desiccation exposure in Drosophila melanogaster and the associated trade-offs. We document a correlation between variations in 20-hydroxyecdysone levels and the altered timing of metamorphic events during the life cycle. Following desiccation, we observed an extension in the larval longevity whereas the duration of the pupal and adult stages was significantly shortened. Alternately, feeding of 20-hydroxyecdysone apparently led to the restoration of the normal temporal pattern of development in the desiccated group. In spite of the desiccation-responsive heterochronic shifts in development, the overall lifespan post recovery remained almost unaltered among the desiccated and undesiccated groups suggesting plasticity in developmental control. This observation reminisces ‘canalization-like’ phenomenon that buffers alterations in the overall lifespan. We thus identified a desiccation-responsive period in the lifespan of D. melanogaster during which variations in ecdysone levels are capable to alter the temporal course of development.     

Heterochromatin and the molecular mechanisms of ‘parent-of-origin’ effects in animals

Twenty five years ago it was proposed that conserved components of constitutive heterochromatin assemble heterochromatin-like complexes in euchromatin and this could provide a general mechanism for regulating heritable (cell-to-cell) changes in gene expressibility. As a special case, differences in the assembly of heterochromatin-like complexes on homologous chromosomes might also regulate the parent-of-origin-dependent gene expression observed in placental mammals. Here, the progress made in the intervening period with emphasis on the role of heterochromatin and heterochromatin-like complexes in parent-of-origin effects in animals is reviewed.