Mismatch Repair in Escherichia coli Cells Lacking Single-Strand Exonucleases ExoI, ExoVII, and RecJ

In vitro, the methyl-directed mismatch repair system of Escherichia coli requires the single-strand exonuclease activity of either ExoI, ExoVII, or RecJ and possibly a fourth, unknown single-strand exonuclease. We have created the first precise null mutations in genes encoding ExoI and ExoVII and find that cells lacking these nucleases and RecJ perform mismatch repair in vivo normally such that triple-null mutants display normal mutation rates. ExoI, ExoVII, and RecJ are either redundant with another function(s) or are unnecessary for mismatch repair in vivo.     

The unfolded protein response transducer Ire1p contains a nuclear localization sequence recognized by multiple beta importins

The Ire1p transmembrane receptor kinase/endonuclease transduces the unfolded protein response (UPR) from the endoplasmic reticulum (ER) to the nucleus in Saccharomyces cerevisiae. In this study, we analyzed the capacity of a highly basic sequence in the linker region of Ire1p to function as a nuclear localization sequence (NLS) both in vivo and in vitro. This 18-residue sequence is capable of targeting green fluorescent protein to the nucleus of yeast cells in a process requiring proteins involved in the Ran GTPase cycle that facilitates nuclear import. Mutagenic analysis and importin binding studies demonstrate that the Ire1p linker region contains overlapping potential NLSs: at least one classical NLS (within sequences 642KKKRKR647 and/or 653KKGR656) that is recognized by yeast importin alpha (Kap60p) and a novel betaNLS (646KRGSRGGKKGRK657) that is recognized by several yeast importin beta homologues. Kinetic binding data suggest that binding to importin beta proteins would predominate in vivo. The UPR, and in particular ER stress-induced HAC1 mRNA splicing, is inhibited by point mutations in the Ire1p NLS that inhibit nuclear localization and also requires functional RanGAP and Ran GEF proteins. The NLS-dependent nuclear localization of Ire1p would thus seem to be central to its role in UPR signaling.     

Molecular chaperones: Clasping the prize

The three-dimensional structure of the substrate-binding domain of DnaK, a bacterial Hsp70, shows how such molecular chaperones can be so promiscuous in recognizing different proteins, yet so accurate in discriminating between unfolded and folded forms of their polypeptide substrates.     

Responses of Plant Communities to Incremental Hydrologic Restoration of a Tide-Restricted Salt Marsh in Southern New England (Massachusetts, U.S.A.)

Hydrologic restoration of Hatches Harbor, a tide-restricted marsh on Cape Cod (Massachusetts), has resulted in significant plant community changes 7 years following the reintroduction of seawater. Since 1999, incremental increases in flow through a tide-restricting dike have facilitated the rapid decline of salt-intolerant vegetation, while encouraging the expansion of native salt marsh taxa. These changes show strong spatial gradients and are correlated with marsh surface elevation, distance from the point of seawater entry, and porewater salinity. Common reed ( Phragmites australis ) has not decreased in abundance but has migrated a considerable distance upslope. In the wake of this retreat native halophytes have proliferated. Now that maximum flow through the existing dike structure has been reached, continued recovery may be limited less by changing physicochemical conditions and more by rates of growth, seed dispersal, and seed germination of salt marsh taxa.     

Systematics of Catenulifera (anamorphic Hyaloscyphaceae) with an assessment of the phylogenetic position of Phialophora hyalina

Catenulifera, typified by C. rhodogena (=Scopulariopsis rhodogena), was established to accommodate the anamorphs of Hyphodiscus (Ascomycota, Helotiales) and to delimit these taxa from members of Phialophora section Catenulatae. Catenulifera rhodogena has been inferred as monophyletic, but its relationship to ascomycetes with poorly differentiated phialidic anamorphs remains enigmatic. To test the hypothesis that C. rhodogena is closely related to morphologically similar species of Phialophora and to further explore the systematics of Catenulifera, we analyzed nuclear rDNA and β-tubulin gene sequences of isolates identified as C. rhodogena, Hyphodiscus hymeniophilus, P. brachyconia, P. brevicollaris and P. hyalina. ITS-LSU and ITS-LSU-β-tubulin phylogenies positioned all isolates except P. hyalina in a single, well-supported clade that consisted of three subclades. Subclade 1 included fungicolous isolates of C. rhodogena and H. hymeniophilus that did not fluoresce when exposed to long-wave UV light. Subclade 2 contained fungicolous and lignicolous strains of C. rhodogena that produced fluorescent colonies and possessed a 366bp indel in the LSU rRNA gene. Neither lineage encompassed the ex-type strains of Cistella rubescens (=H. hymeniophilus) or S. rhodogena, but the former isolate was inferred as sister to Subclade 2. Subclade 3 included all isolates of P. brachyconia, a species recognized here as C. brachyconia comb. nov. A fourth isolate of P. brachyconia that was extralimital to Subclade 3 is described as C. luxurians sp. nov. The positions of C. brevicollaris comb. nov., a species based on P. brevicollaris, and C. luxurians were not resolved in the ITS-LSU phylogeny. P. hyalina is not closely related to Catenulifera.     

xni-deficient Escherichia coli are proficient for recombination and multiple pathways of repair

Single-strand-dependent DNA exonucleases play important roles in DNA repair and recombination in all organisms. In Escherichia coli the redundant functions provided by the RecJ, ExoI, ExoVII and ExoX exonucleases are required for mismatch repair, UV resistance and homologous recombination. We have examined whether the xni gene product, the single-strand exonuclease ExoIX, is also a member of this group. We find that deletion of xni has no effect on the above processes, or on resistance to oxidative damage, even in combination with other exonuclease mutations. We conclude that the xni gene product does not belong to this group of nucleases that play redundant roles in DNA recombination and repair.     

Hypermutation in stationary-phaseE. coli: tales from thelac operon

Escherichia coli cells are capable of complex regulatory responses to environmental conditions and stresses. In some circumstances, the response includes an increase in the mutation rate, effectively mutagenizing the genome. The classic example is the SOS response to DNA damage. Recent work indicates that other environmental stresses can also result in mutation of the genome. Modulation of mutation rate may be a more prevalent stress response than previously thought. In this review we focus on genome-wide mutation inE. coli cells subjected to a nonlethal genetic selection for reversion of alac frameshift allele. Reversion of thelac frameshift allele occurs via a novel mechanism that requires homologous recombination functions, and is enhanced by transiently diminished postsynthesis mismatch repair. A model for recombination-dependent stationary-phase mutation will be presented and its relevance for genome-wide mutation discussed.