Covarying patterns of macroinvertebrate and fish assemblages along natural and human activity gradients: implications for bioassessment

Bioassessment is based upon the premise that biological assemblages have predictable relationships with the surrounding natural and human environments. As the nature of these relationships can vary from region to region, it is important that environment–biota relationships be established prior to the initiation of any bioassessment program. In this study, multivariate analysis was used to establish how fish and benthic macroinvertebrate (BMI) assemblages in southwestern Ontario streams vary across natural and human activity gradients. The use of canonical correspondence analysis allowed us to determine that changes in community composition of both fish and BMI are strongly correlated with variation in the extent of human activity. The primary source of variation in community composition across activity gradients appeared to reflect a shift from intolerant to tolerant taxa as the extent of human activity increased. Habitat and feeding traits, for BMI and fish respectively, accounted for a secondary source of variation primarily attributable to differences in the extent of human activity at the reach scale. However, variation in human activity, especially at the basin scale, covaried with the dominant natural gradient of surface geology, making interpretation of the results difficult. Implications for bioassessment studies are discussed.     

Molecular Mechanisms of GD3-Induced Apoptosis in U-1242 MG Glioma Cells

An increasing amount of evidence indicates that the disialoganglioside GD3 is involved in apoptosis in many cell lines. Our previous studies demonstrated that endogenous GD3 expression induced apoptosis in U-1242 MG glioma cells transfected with the GD3 synthase gene (U1242MG-GD3 cells). In this paper, we present further investigations on the molecular mechanisms of GD3-induced apoptosis in this cell line. We found that endogenously synthesized GD3 localizes to the caveolae of this cell line, where it promotes the localization of death receptor 5 (DR5), tumor necrosis factor receptor-1 (TNF-R1), and Fas (Apo-1) to the caveolae. In addition, caspase-8 was translocated to the caveolar fraction and cleaved; the cleaved proteins were then re-located into the high density fractions. However, GD3 had no effect on the distribution of the adapter protein Fas-associated death domain (FADD). We conclude that GD3 functions as a regulatory molecule early in the extrinsic apoptosis pathway.O. M. Omran, H. E. Saqr––Both authors contributed equally to this work.     

Phosphorylation of Rad55 on serines 2, 8, and 14 is required for efficient homologous recombination in the recovery of stalled replication forks

DNA damage checkpoints coordinate the cellular response to genotoxic stress and arrest the cell cycle in response to DNA damage and replication fork stalling. Homologous recombination is a ubiquitous pathway for the repair of DNA double-stranded breaks and other checkpoint-inducing lesions. Moreover, homologous recombination is involved in postreplicative tolerance of DNA damage and the recovery of DNA replication after replication fork stalling. Here, we show that the phosphorylation on serines 2, 8, and 14 (S2,8,14) of the Rad55 protein is specifically required for survival as well as for normal growth under genome-wide genotoxic stress. Rad55 is a Rad51 paralog in Saccharomyces cerevisiae and functions in the assembly of the Rad51 filament, a central intermediate in recombinational DNA repair. Phosphorylation-defective rad55-S2,8,14A mutants display a very slow traversal of S phase under DNA-damaging conditions, which is likely due to the slower recovery of stalled replication forks or the slower repair of replication-associated DNA damage. These results suggest that Rad55-S2,8,14 phosphorylation activates recombinational repair, allowing for faster recovery after genotoxic stress.     

The Nse5-Nse6 dimer mediates DNA repair roles of the Smc5-Smc6 complex

Stabilization and processing of stalled replication forks is critical for cell survival and genomic integrity. We characterize a novel DNA repair heterodimer of Nse5 and Nse6, which are nonessential nuclear proteins critical for chromosome segregation in fission yeast. The Nse5/6 dimer facilitates DNA repair as part of the Smc5-Smc6 holocomplex (Smc5/6), the basic architecture of which we define. Nse5-Nse6 [corrected] (Nse5 and Nse6) [corrected] mutants display a high level of spontaneous DNA damage and mitotic catastrophe in the absence of the master checkpoint regulator Rad3 (hATR). Nse5/6 mutants are required for the response to genotoxic agents that block the progression of replication forks, acting in a pathway that allows the tolerance of irreparable UV lesions. Interestingly, the UV sensitivity of Nse5/6 [corrected] is suppressed by concomitant deletion of the homologous recombination repair factor, Rhp51 (Rad51). Further, the viability of Nse5/6 mutants depends on Mus81 and Rqh1, factors that resolve or prevent the formation of Holliday junctions. Consistently, the UV sensitivity of cells lacking Nse5/6 can be partially suppressed by overexpressing the bacterial resolvase RusA. We propose a role for Nse5/6 mutants in suppressing recombination that results in Holliday junction formation or in Holliday junction resolution.     

The expression of nifB gene from Herbaspirillum seropedicae is dependent upon the NifA and RpoN proteins

The putative nifB promoter region of Herbaspirillum seropedicae contained two sequences homologous to NifA-binding site and a -24/-12 type promoter. A nifB::lacZ fusion was assayed in the backgrounds of both Escherichia coli and H. seropedicae. In E. coli, the expression of nifB::lacZ occurred only in the presence of functional rpoN and Klebsiella pneumoniae nifA genes. In addition, the integration host factor (IHF) stimulated the expression of the nifB::lacZ fusion in this background. In H. seropedicae, nifB expression occurred only in the absence of ammonium and under low levels of oxygen, and it was shown to be strictly dependent on NifA. DNA band shift experiments showed that purified K. pneumoniae RpoN and E. coli IHF proteins were capable of binding to the nifB promoter region, and in vivo dimethylsulfate footprinting showed that NifA binds to both NifA-binding sites. These results strongly suggest that the expression of the nifB promoter of H. seropedicae is dependent on the NifA and RpoN proteins and that the IHF protein stimulates NifA activation of nifB promoter.     

Ppc89 links multiple proteins, including the septation initiation network, to the core of the fission yeast spindle-pole body

The spindle-pole body (SPB), the yeast analog of the centrosome, serves as the major microtubule (MT) organizing center in the yeast cell. In addition to this central function, the SPB organizes and concentrates proteins required for proper coordination between the nuclear-division cycle and cytokinesis. For example, the Schizosaccharomyces pombe septation-initiation network (SIN), which is responsible for initiating actomyosin ring constriction and septation, is assembled at the SPB through its two scaffolding components, Sid4 and Cdc11. In an effort to identify novel SIN interactors, we purified Cdc11 and identified by mass spectrometry a previously uncharacterized protein associated with it, Ppc89. Ppc89 localizes constitutively to the SPB and interacts directly with Sid4. A fusion between the N-terminal 300 amino acids of Sid4 and a SPB targeting domain of Ppc89 supplies the essential function of Sid4 in anchoring the SIN. ppc89Delta cells are inviable and exhibit defects in SPB integrity, and hence in spindle formation, chromosome segregation, and SIN localization. Ppc89 overproduction is lethal, resulting primarily in a G2 arrest accompanied by massive enlargement of the SPB and increased SPB MT nucleation. These results suggest a fundamental role for Ppc89 in organization of the S. pombe SPB.     

Double-stranded DNA translocation: structure and mechanism of hexameric FtsK

FtsK is a DNA translocase that coordinates chromosome segregation and cell division in bacteria. In addition to its role as activator of XerCD site-specific recombination, FtsK can translocate double-stranded DNA (dsDNA) rapidly and directionally and reverse direction. We present crystal structures of the FtsK motor domain monomer, showing that it has a RecA-like core, the FtsK hexamer, and also showing that it is a ring with a large central annulus and a dodecamer consisting of two hexamers, head to head. Electron microscopy (EM) demonstrates the DNA-dependent existence of hexamers in solution and shows that duplex DNA passes through the middle of each ring. Comparison of FtsK monomer structures from two different crystal forms highlights a conformational change that we propose is the structural basis for a rotary inchworm mechanism of DNA translocation.