Evolution of the LINE-like I element in the Drosophila melanogaster species subgroup

LINE-like retrotransposons, the so-called I elements, control the system of I-R (inducer-reactive) hybrid dysgenesis in Drosophila melanogaster. I elements are present in many Drosophila species. It has been suggested that active, complete I elements, located at different sites on the chromosomes, invaded natural populations of D. melanogaster recently (1920–1970). But old strains lacking active I elements have only defective I elements located in the chromocenter. We have cloned I elements from D. melanogaster and the melanogaster subgroup. In D. melanogaster, the nucleotide sequences of chromocentral I elements differed from those on chromosome arms by as much as 7%. All the I elements of D. mauritiana and D. sechellia are more closely related to the chromosomal I elements of D. melanogaster than to the chromocentral I elements in any species. No sequence difference was observed in the surveyed region between two chromosomal I elements isolated from D. melanogaster and one from D. simulans. These findings strongly support the idea that the defective chromocentral I elements of D. melanogaster originated before the species diverged and the chromosomal I elements were eliminated. The chromosomal I elements reinvaded natural populations of D. melanogaster recently, and were possibly introduced from D. simulans by horizontal transmission.     

ATP-sensitive K+ channel-mediated glucose uptake is independent of IRS-1/phosphatidylinositol 3-kinase signaling

We previously found that disruption of Kir6.2-containing ATP-sensitive K+ (KATP) channels increases glucose uptake in skeletal muscle, but the mechanism is not clear. In the present study, we generated knockout mice lacking both Kir6.2 and insulin receptor substrate-1 (IRS-1). Because IRS-1 is the major substrate of insulin receptor kinase, we expected disruption of the IRS-1 gene to reduce glucose uptake in Kir6.2 knockout mice. However, the double-knockout mice do not develop insulin resistance or glucose intolerance. An insulin tolerance test reveals the glucose-lowering effect of exogenous insulin in double-knockout mice and in Kir6.2 knockout mice to be similarly enhanced compared with wild-type mice. The basal 2-deoxyglucose uptake rate in skeletal muscle of double-knockout mice is increased similarly to the rate in Kir6.2 knockout mice. Accordingly, disruption of the IRS-1 gene affects neither systemic insulin sensitivity nor glucose uptake in skeletal muscles of Kir6.2-deficient mice. In addition, no significant changes were observed in phosphatidylinositol 3-kinase (PI3K) activity and its downstream signal in skeletal muscle due to lack of the Kir6.2 gene. Disruption of Kir6.2-containing Katp channels clearly protects against IRS-1-associated insulin resistance by increasing glucose uptake in skeletal muscles by a mechanism separate from the IRS-1/PI3K pathway.     

Drosophila big brain does not act as a water channel, but mediates cell adhesion

The neurogenic gene Drosophilabig brain (bib) has a high sequence homology to aquaporin-4. However, its cellular functions in Drosophila neurogenesis have remained elusive. Here we investigated cell adhesion, and the ion and water permeability of Bib. The adhesive function was examined by a cell aggregation assay using L cells. Bib-transfected L cells formed aggregated clusters, while control-L cells remained as a single cell suspension. Ion permeation was not confirmed in L cells stably expressing Bib. When expressed in COS7 cells, Bib exhibited limited water permeability. This newly found cell adhesive function of Bib may be important for Drosophila neurogenesis.     

EGFR signaling pathway negatively regulates PSA expression and secretion via the PI3K-Akt pathway in LNCaP prostate cancer cells

Epidermal growth factor (EGF) and its receptor (EGFR) are involved in hormone-refractory growth and poor prognosis of a subgroup of human prostate cancer. In this communication, we investigated the regulation of PSA by the EGFR signaling pathway using LNCaP C-81 prostate cancer cells. Administration of EGF stimulated the growth of LNCaP C-81 cells, however, PSA expression and secretion were suppressed. An EGFR inhibitor, AG1478, abrogated the PSA suppression effect by EGF, in concurrence with the suppression of tyro-phosphorylation levels of EGFR. Interestingly, the AR level was also decreased in EGF-treated LNCaP C-81 cells. Moreover, LY294002, but not PD98059, inhibited the PSA and AR suppression effect by EGF in concurrence with the suppression of phosphorylation levels of Akt. In conclusion, our results strongly suggest the existence of a novel androgen-independent PSA regulatory mechanism, i.e., the EGFR signaling pathway negatively regulates PSA expression which may be induced by the alteration of AR expression via the PI3K-Akt pathway in LNCaP C-81 cells.     

Suppression of Vps13 adaptor protein mutants reveals a central role for PI4P in regulating prospore membrane extension

Vps13 family proteins are proposed to function in bulk lipid transfer between membranes, but little is known about their regulation. During sporulation of Saccharomyces cerevisiae, Vps13 localizes to the prospore membrane (PSM) via the Spo71–Spo73 adaptor complex. We previously reported that loss of any of these proteins causes PSM extension and subsequent sporulation defects, yet their precise function remains unclear. Here, we performed a genetic screen and identified genes coding for a fragment of phosphatidylinositol (PI) 4-kinase catalytic subunit and PI 4-kinase noncatalytic subunit as multicopy suppressors of spo73Δ. Further genetic and cytological analyses revealed that lowering PI4P levels in the PSM rescues the spo73Δ defects. Furthermore, overexpression of VPS13 and lowering PI4P levels synergistically rescued the defect of a spo71Δ spo73Δ double mutant, suggesting that PI4P might regulate Vps13 function. In addition, we show that an N-terminal fragment of Vps13 has affinity for the endoplasmic reticulum (ER), and ER-plasma membrane (PM) tethers localize along the PSM in a manner dependent on Vps13 and the adaptor complex. These observations suggest that Vps13 and the adaptor complex recruit ER-PM tethers to ER-PSM contact sites. Our analysis revealed that involvement of a phosphoinositide, PI4P, in regulation of Vps13, and also suggest that distinct contact site proteins function cooperatively to promote de novo membrane formation.     

Behavioral phenotypes of Disc1 missense mutations in mice

To support the role of DISC1 in human psychiatric disorders, we identified and analyzed two independently derived ENU-induced mutations in Exon 2 of mouse Disc1. Mice with mutation Q31L showed depressive-like behavior with deficits in the forced swim test and other measures that were reversed by the antidepressant bupropion, but not by rolipram, a phosphodiesterase-4 (PDE4) inhibitor. In contrast, L100P mutant mice exhibited schizophrenic-like behavior, with profound deficits in prepulse inhibition and latent inhibition that were reversed by antipsychotic treatment. Both mutant DISC1 proteins exhibited reduced binding to the known DISC1 binding partner PDE4B. Q31L mutants had lower PDE4B activity, consistent with their resistance to rolipram, suggesting decreased PDE4 activity as a contributory factor in depression. This study demonstrates that Disc1 missense mutations in mice give rise to phenotypes related to depression and schizophrenia, thus supporting the role of DISC1 in major mental illness.     

Mitochondrial alterations related to programmed cell death in tobacco cells under aluminium stress

The present investigation was undertaken to verify whether mitochondria play a significant role in aluminium (Al) toxicity, using the mitochondria isolated from tobacco cells (Nicotiana tabacum, non-chlorophyllic cell line SL) under Al stress. An inhibition of respiration was observed in terms of state-III, state-IV, succinate-dependent, alternative oxidase (AOX)-pathway capacity and cytochrome (CYT)-pathway capacity, respectively, in the mitochondria isolated from tobacco cells subjected to Al stress for 18 h. In accordance with the respiratory inhibition, the mitochondrial ATP content showed a significant decrease under Al treatment. An enhancement of reactive oxygen species (ROS) production under state-III respiration was observed in the mitochondria isolated from Al-treated cells, which would create an oxidative stress situation. The opening of mitochondrial permeability transition pore (MPTP) was seen more extensively in mitochondria isolated from Al-treated cells than in those isolated from control cells. This was Ca(2+) dependent and well modulated by dithioerythritol (DTE) and Pi, but insensitive to cyclosporine A (CsA). The collapse of inner mitochondrial membrane potential (DeltaPsi(m)) was also observed with a release of cytochrome c from mitochondria. A great decrease in the ATP content was also seen under Al stress. Transmission electron microscopy analysis of Al-treated cells also corroborated our biochemical data with distortion in membrane architecture in mitochondria. TUNEL-positive nuclei in Al-treated cells strongly indicated the occurrence of nuclear fragmentation. From the above study, it was concluded that Al toxicity affects severely the mitochondrial respiratory functions and alters the redox status studied in vitro and also the internal structure, which seems to cause finally cell death in tobacco cells.     

Insulin/IGF signaling regulates the change in commitment in imaginal discs and primordia by overriding the effect of juvenile hormone

At the beginning of the final larval (fifth) instar of Manduca sexta, imaginal precursors including wing discs and eye primordia initiate metamorphic changes, such as pupal commitment, patterning and cell proliferation. Juvenile hormone (JH) prevents these changes in earlier instars and in starved final instar larvae, but nutrient intake overcomes this effect of JH in the latter. In this study, we show that a molecular marker of pupal commitment, broad, is up-regulated in the wing discs by feeding on sucrose or by bovine insulin or Manduca bombyxin in starved final instar larvae. This effect of insulin could not be prevented by JH. In vitro insulin had no effect on broad expression but relieved the suppression of broad expression by JH. This effect of insulin was directly on the disc as shown by its reduction in the presence of insulin receptor dsRNA. In starved penultimate fourth instar larvae, broad expression in the wing disc was not up-regulated by insulin. The discs became responsive to this action of insulin during the molt to the fifth instar together with the ability to become pupally committed in response to 20-hydroxyecdysone. Thus, the Manduca bombyxin acts as a metamorphosis-initiating factor in the imaginal precursors.