Evolutionarily conserved multisubunit RBL2/p130 and E2F4 protein complex represses human cell cycle-dependent genes in quiescence

The mammalian Retinoblastoma (RB) family including pRB, p107, and p130 represses E2F target genes through mechanisms that are not fully understood. In D. melanogaster, RB-dependent repression is mediated in part by the multisubunit protein complex Drosophila RBF, E2F, and Myb (dREAM) that contains homologs of the C. elegans synthetic multivulva class B (synMuvB) gene products. Using an integrated approach combining proteomics, genomics, and bioinformatic analyses, we identified a p130 complex termed DP, RB-like, E2F, and MuvB (DREAM) that contains mammalian homologs of synMuvB proteins LIN-9, LIN-37, LIN-52, LIN-54, and LIN-53/RBBP4. DREAM bound to more than 800 human promoters in G0 and was required for repression of E2F target genes. In S phase, MuvB proteins dissociated from p130 and formed a distinct submodule that bound MYB. This work reveals an evolutionarily conserved multisubunit protein complex that contains p130 and E2F4, but not pRB, and mediates the repression of cell cycle-dependent genes in quiescence.     

Electric field effects on membranes: gramicidin A as a test ground

Electric fields due to transmembrane potential differences or ionic gradients across the membrane are presumably crucial for many reactions across membranes or close to membranes like signal transduction, control of ion channels or the generation of neural impulses. Molecular dynamics simulations have been used to study the influence of external electric fields on a mixed gramicidin/phospholipid bilayer system. At high field strengths, formation of membrane electropores occurred both close and distal to the gramicidin. Gramicidin was found to stabilize the membrane adjacent to the protein but also at larger distances of up to 2-3 nm. As a result, membrane pore formation was found to be significantly suppressed for the mixed gramicidin/DMPC system. Moderate field strengths only weakly affected the structure and dynamics of the gramicidin. Spontaneous potassium passage events in external electric fields were observed for both the head-to-head helical conformation as well as for the double helical conformation of gramicidin A. The double-helical conformation was found to facilitate ion passage compared to the head-to-head helical dimer.     

Lead-induced alterations of apoptosis and neurotrophic factor mRNA in the developing rat cortex, hippocampus, and cerebellum

Previous reports have recently shown the prototypic neurotoxicant, lead, to induce apoptosis in the brains of developing organisms. In the current study, timed-pregnant rats were exposed to lead acetate (0.2% in the drinking water) 24 h following birth at postnatal day 1 (PND 1). Dams and pups were continuously exposed to lead through the drinking water of the dam until PND 20. Postnatal exposure in the pups resulted in altered mRNA levels of the following apoptotic and neurotrophic factors: caspase 2 and 3, bax, bcl-x, brain-derived neurotrophic factor (BDNF). Ribonuclease protection assays were conducted to measure the factors simultaneously at the following postnatal time points: 9, 12, 15, 20, 25, days. Our results suggest a brain region- and time-specific response following lead acetate exposure. The region most vulnerable to alterations occurs in the hippocampus with alterations beginning at PND 12, in which caspase 3, bcl-x, BDNF increase with lead exposure. Significant treatment effects were not observed for both the cortex and cerebellum.     

Generation of a germ cell-specific mouse transgenic Cre line, Vasa-Cre

Cell type-specific genetic modification using the Cre/loxP system is a powerful tool for genetic analysis of distinct cell lineages. Because of the exquisite specificity of Vasa expression (confined to the germ cell lineage in invertebrate and vertebrate species), we hypothesized that a Vasa promoter-driven transgenic Cre line would prove useful for the germ cell lineage-specific inactivation of genes. Here we describe a transgenic mouse line, Vasa-Cre, where Cre is efficiently and specifically expressed in germ cells. Northern analysis showed that transgene expression was confined to the gonads. Cre-mediated recombination with the Rosa26-lacZ reporter was observed beginning at approximately e15, and was >95% efficient in male and female germ cells by birth. Although there was a potent maternal effect with some animals showing more widespread recombination, there was no ectopic activity in most adults. This Vasa-Cre transgenic line should thus prove useful for genetic analysis of diverse aspects of gametogenesis and as a general deletor line.