Rad54: the Swiss Army knife of homologous recombination?

Homologous recombination (HR) is a ubiquitous cellular pathway that mediates transfer of genetic information between homologous or near homologous (homeologous) DNA sequences. During meiosis it ensures proper chromosome segregation in the first division. Moreover, HR is critical for the tolerance and repair of DNA damage, as well as in the recovery of stalled and broken replication forks. Together these functions preserve genomic stability and assure high fidelity transmission of the genetic material in the mitotic and meiotic cell divisions. This review will focus on the Rad54 protein, a member of the Snf2-family of SF2 helicases, which translocates on dsDNA but does not display strand displacement activity typical for a helicase. A wealth of genetic, cytological, biochemical and structural data suggests that Rad54 is a core factor of HR, possibly acting at multiple stages during HR in concert with the central homologous pairing protein Rad51

Range Maps for the Terrestrial Natural Communities of Nebraska

This document includes range maps for the 83 terrestrial natural (plant) communities described in Terrestrial Ecological Systems and Natural Communities of Nebraska (Version IV - March 9, 2010) by Rolfsmeier and Steinauer 2010. Each map includes the known range and potential range for individual community types. The known range includes areas where a community has been documented and areas where it has not been documented but thought very likely to occur. The potential range includes areas where the community has not been documented but is thought to have some potential to occur. The maps should be considered approximations of the community ranges and are meant to be used at a coarse scale

The Distribution and Reproductive Phenology of the Milkweeds (Asclepiadaceae: \u3ci\u3eAsclepias\u3c/i\u3e and \u3ci\u3eCynanchum\u3c/i\u3e) in Nebraska

The Asclepiadaceae are represented in Nebraska by seventeen native species of Asclepias and one of Cynanchum. Two species are here added to the State\u27s flora: Asclepias asperula and A. purpurascens. Detailed county distribution maps are presented for all species based upon our extensive field and herbarium work, and numerous additions and corrections are made to the maps published in the Atlas of the Flora of the Great Plains in 1977. A map is presented showing the zone of hybridization of A. speciosa with A. syriaca in central Nebraska. Data are given on flowering and fruiting phenology for each species, and floral odors are described

New and Corrected Floristic Records for Nebraska

Nineteen species (including eight Eurasian ones) are newly recorded for Nebraska: Alopecurus arundinaceus, Amaranthus californicus, Asclepias asperula, A. purpurascens, Cardamine {lexuosa*, Centaurea diffusa, Dipsacus laciniatus, Eriochloa villosa, Euclidium syriacum, Gentiana alba, Geranium viscosissimum, Geum vernum, Goodyera oblongifolia, Haplopappus multicaulis, Heterotheca latifolia, Lathyrus tuberosus, Polygonum douglasii, Scirpus saximontanus, Veronica biloba. Twenty-one others are shown to be more widespread in Nebraska than previously known, one has a more restricted range than previously reported, two (Scirpus smithii, S. torreyi) are deleted from the flora based upon corrected identifications, and the status of some rarely-collected species is updated. Thirty-two additions, two deletions, and two corrections to the recently published flora of Seward County are also presented along with thirteen additions to the flora of Keith County

New and Corrected Floristic Records for Nebraska

Nineteen species (including eight Eurasian ones) are newly recorded for Nebraska: Alopecurus arundinaceus, Amaranthus californicus, Asclepias asperula, A purpurascens, Cardamine {lexuosa*, Centaurea diffusa, Dipsacus laciniatus, Eriochloa villosa, Euclidium syriacum, Gentiana alba, Geranium viscosissimum, Geum vernum, Goodyera oblongifolia, Haplopappus multicaulis, Heterotheca latifolia, Lathyrus tuberosus, Polygonum douglasii, Scirpus saximontanus, Veronica biloba. Twenty-one others are shown to be more widespread in Nebraska than previously known, one has a more restricted range than previously reported, two (Scirpus smithii, S. torreyi) are deleted from the flora based upon corrected identifications, and the status of some rarely-collected species is updated. Thirty-two additions, two deletions, and two corrections to the recentlypublished flora of Seward County are also presented along with thirteen additions to the flora of Keith County

Molecular Characterization of the α-Glucosidase Gene (\u3ci\u3emalA\u3c/i\u3e) from the Hyperthermophilic Archaeon \u3ci\u3eSulfolobus solfataricus\u3c/i\u3e

Acidic hot springs are colonized by a diversity of hyperthermophilic organisms requiring extremes of temperature and pH for growth. To clarify how carbohydrates are consumed in such locations, the structural gene (malA) encoding the major soluble α-glucosidase (maltase) and flanking sequences from Sulfolobus solfataricus were cloned and characterized. This is the first report of an α-glucosidase gene from the archaeal domain. malA is 2,083 bp and encodes a protein of 693 amino acids with a calculated mass of 80.5 kDa. It is flanked on the 5’ side by an unusual 1-kb intergenic region. Northern blot analysis of the malA region identified transcripts for malA and an upstream open reading frame located 5’ to the 1-kb intergenic region. The malA transcription start site was located by primer extension analysis to a guanine residue 8 bp 5’ of the malA start codon. Gel mobility shift analysis of the malA promoter region suggests that sequences 3’ to position 233, including a consensus archaeal TATA box, play an essential role in malA expression. malA homologs were detected by Southern blot analysis in other S. solfataricus strains and in Sulfolobus shibatae, while no homologs were evident in Sulfolobus acidocaldarius, lending further support to the proposed revision of the genus Sulfolobus. Phylogenetic analyses indicate that the closest S. solfataricus α-glucosidase homologs are of mammalian origin. Characterization of the recombinant enzyme purified from Escherichia coli revealed differences from the natural enzyme in thermostability and electrophoretic behavior. Glycogen is a substrate for the recombinant enzyme. Unlike maltose hydrolysis, glycogen hydrolysis is optimal at the intracellular pH of the organism. These results indicate a unique role for the S. solfataricus α-glucosidase in carbohydrate metabolism

Identification of a system required for the functional surface localization of sugar binding proteins with class III signal peptides in Sulfolobus solfataricus

The hyperthermophilic archaeon Sulfolobus solfataricus contains an unusual large number of sugar binding proteins that are synthesized as precursors with a class III signal peptide. Such signal peptides are commonly used to direct archaeal flagellin subunits or bacterial (pseudo)pilins into extracellular macromolecular surface appendages. Likewise, S. solfataricus binding proteins have been suggested to assemble in higher ordered surface structures as well, tentatively termed the bindosome. Here we show that S. solfataricus contains a specific system that is needed for the functional surface localization of sugar binding proteins. This system, encoded by the bas (bindosome assembly system) operon, is composed of five proteins: basABC, three homologues of so-called bacterial (pseudo)pilins; BasE, a cytoplasmic ATPase; and BasF, an integral membrane protein. Deletion of either the three (pseudo)pilin genes or the basEF genes resulted in a severe defect of the cells to grow on substrates which are transported by sugar binding proteins containing class III signal peptides, while growth on glucose and maltose was restored when the corresponding genes were reintroduced in these cells. Concomitantly, ΔbasABC and ΔbasEF cells were severely impaired in glucose uptake even though the sugar binding proteins were normally secreted across the cytoplasmic membrane. These data underline the hypothesis that the bas operon is involved in the functional localization of sugar binding proteins at the cell surface of S. solfataricus. In contrast to surface structure assembly systems of Gram-negative bacteria, the bas operon seems to resemble an ancestral simplified form of these machineries.

Template-dependent polymerization across discontinuous templates by the heterodimeric primase from the hyperthermophilic archaeon Sulfolobus solfataricus

The eukaryotic-like primase from the hyperthermophilic archaeon Sulfolobus solfataricus (SsoPriSL) exhibits a range of activities including template-dependent de novo primer synthesis, primer extension and template-independent terminal nucleotidyl transfer using either rNTPs or dNTPs. Remarkably, the enzyme is able to synthesize products far longer than templates in vitro. Here we show that the long products resulted from template-dependent polymerization across discontinuous templates (PADT) by SsoPriSL. PADT was initiated through either primer synthesis or terminal transfer, and occurred efficiently on templates containing contiguous dCs. Template switching took place when the 3′-end of a growing strand synthesized on one template annealed to another template directly or following the terminal addition of nucleotides, and was subsequently extended on the new template. The key to PADT was the ability of SsoPriSL to promote strand annealing. SsoPriSL catalyzed PADT with either dNTPs or rNTPs as the substrates but preferred the latter. The enzyme remained active in PADT but became inefficient in primer synthesis in vitro when temperature was raised from 55°C to 70°C. Our results suggest that SsoPriSL is capable of bridging noncomplementary DNA ends and, therefore, may serve a role in double-strand DNA break repair in Archaea

A truncated DNA-damage-signaling response is activated after DSB formation in the G1 phase of Saccharomyces cerevisiae

In Saccharomyces cerevisiae, the DNA damage response (DDR) is activated by the spatio-temporal colocalization of Mec1-Ddc2 kinase and the 9-1-1 clamp. In the absence of direct means to monitor Mec1 kinase activation in vivo, activation of the checkpoint kinase Rad53 has been taken as a proxy for DDR activation. Here, we identify serine 378 of the Rad55 recombination protein as a direct target site of Mec1. Rad55-S378 phosphorylation leads to an electrophoretic mobility shift of the protein and acts as a sentinel for Mec1 activation in vivo. A single double-stranded break (DSB) in G1-arrested cells causes phosphorylation of Rad55-S378, indicating activation of Mec1 kinase. However, Rad53 kinase is not detectably activated under these conditions. This response required Mec1-Ddc2 and loading of the 9-1-1 clamp by Rad24-RFC, but not Rad9 or Mrc1. In addition to Rad55–S378, two additional direct Mec1 kinase targets are phosphorylated, the middle subunit of the ssDNA-binding protein RPA, RPA2 and histone H2A (H2AX). These data suggest the existence of a truncated signaling pathway in response to a single DSB in G1-arrested cells that activates Mec1 without eliciting a full DDR involving the entire signaling pathway including the effector kinases

Histone Deacetylase Complexes Promote Trinucleotide Repeat Expansions

Genetic analysis in budding yeast and in cultured human astrocytes reveals that specific histone deacetylase complexes accelerate expansion mutations in DNA triplet repeats