Mre11 dimers coordinate DNA end bridging and nuclease processing in double-strand-break repair

Mre11 forms the core of the multifunctional Mre11-Rad50-Nbs1 (MRN) complex that detects DNA double-strand breaks (DSBs), activates the ATM checkpoint kinase, and initiates homologous recombination (HR) repair of DSBs. To define the roles of Mre11 in both DNA bridging and nucleolytic processing during initiation of DSB repair, we combined small-angle X-ray scattering (SAXS) and crystal structures of Pyrococcus furiosus Mre11 dimers bound to DNA with mutational analyses of fission yeast Mre11. The Mre11 dimer adopts a four-lobed U-shaped structure that is critical for proper MRN complex assembly and for binding and aligning DNA ends. Further, mutations blocking Mre11 endonuclease activity impair cell survival after DSB induction without compromising MRN complex assembly or Mre11-dependant recruitment of Ctp1, an HR factor, to DSBs. These results show how Mre11 dimerization and nuclease activities initiate repair of DSBs and collapsed replication forks, as well as provide a molecular foundation for understanding cancer-causing Mre11 mutations in ataxia telangiectasia-like disorder (ATLD).     

Micro Radiometric Method for Study of Acid-insoluble Materials in Monolayer Cell Cultures

A simple radiometric procedure for study of acid-insoluble products synthesized in monolayer cell cultures is described. Cell cultures were produced directly on the bottom surface of scintillation vials or on glass cover slips (8 X 30 mm). The cells were labeled and extracted; the radioactivity was determined while the cells remained affixed to the glass surface upon which they were grown. This procedure enabled rapid investigations of certain biosynthetic processes to be carried out by using many individual cell cultures. The method was applied to an investigation of (3)H-thymidine incorporation induced by vaccinia virus in a 5-bromodeoxyuridine-resistant cell line. (14)C-labeling was evaluated as an alternate procedure for cell quantitation.     

The western Mediterranean region provided the founder population of domesticated narrow-leafed lupin

Key message

This study revealed that the western Mediterranean provided the founder population for domesticated narrow-leafed lupin and that genetic diversity decreased significantly during narrow-leafed lupin domestication.

Abstract

The evolutionary history of plants during domestication profoundly shaped the genome structure and genetic diversity of today’s crops. Advances in next-generation sequencing technologies allow unprecedented opportunities to understand genome evolution in minor crops, which constitute the majority of plant domestications. A diverse set of 231 wild and domesticated narrow-leafed lupin (Lupinus angustifolius L.) accessions were subjected to genotyping-by-sequencing using diversity arrays technology. Phylogenetic, genome-wide divergence and linkage disequilibrium analyses were applied to identify the founder population of domesticated narrow-leafed lupin and the genome-wide effect of domestication on its genome. We found wild western Mediterranean population as the founder of domesticated narrow-leafed lupin. Domestication was associated with an almost threefold reduction in genome diversity in domesticated accessions compared to their wild relatives. Selective sweep analysis identified no significant footprints of selection around domestication loci. A genome-wide association study identified single nucleotide polymorphism markers associated with pod dehiscence. This new understanding of the genomic consequences of narrow-leafed lupin domestication along with molecular marker tools developed here will assist plant breeders more effectively access wild genetic diversity for crop improvement.

     

Continental scale analysis of bird migration timing: influences of climate and life history traits—a generalized mixture model clustering and discriminant approach

There is substantial evidence of climate-related shifts to the timing of avian migration. Although spring arrival has generally advanced, variable species responses and geographical biases in data collection make it difficult to generalise patterns. We advance previous studies by using novel multivariate statistical techniques to explore complex relationships between phenological trends, climate indices and species traits. Using 145 datasets for 52 bird species, we assess trends in first arrival date (FAD), last departure date (LDD) and timing of peak abundance at multiple Australian locations. Strong seasonal patterns were found, i.e. spring phenological events were more likely to significantly advance, while significant advances and delays occurred in other seasons. However, across all significant trends, the magnitude of delays exceeded that of advances, particularly for FAD (+22.3 and ?9.6 days/decade, respectively). Geographic variations were found, with greater advances in FAD and LDD, in south-eastern Australia than in the north and west. We identified four species clusters that differed with respect to species traits and climate drivers. Species within bird clusters responded in similar ways to local climate variables, particularly the number of raindays and rainfall. The strength of phenological trends was more strongly related to local climate variables than to broad-scale drivers (Southern Oscillation Index), highlighting the importance of precipitation as a driver of movement in Australian birds.     

Diverse Lifestyles and Strategies of Plant Pathogenesis Encoded in the Genomes of Eighteen Dothideomycetes Fungi

The class Dothideomycetes is one of the largest groups of fungi with a high level of ecological diversity including many plant pathogens infecting a broad range of hosts. Here, we compare genome features of 18 members of this class, including 6 necrotrophs, 9 (hemi)biotrophs and 3 saprotrophs, to analyze genome structure, evolution, and the diverse strategies of pathogenesis. The Dothideomycetes most likely evolved from a common ancestor more than 280 million years ago. The 18 genome sequences differ dramatically in size due to variation in repetitive content, but show much less variation in number of (core) genes. Gene order appears to have been rearranged mostly within chromosomal boundaries by multiple inversions, in extant genomes frequently demarcated by adjacent simple repeats. Several Dothideomycetes contain one or more gene-poor, transposable element (TE)-rich putatively dispensable chromosomes of unknown function. The 18 Dothideomycetes offer an extensive catalogue of genes involved in cellulose degradation, proteolysis, secondary metabolism, and cysteine-rich small secreted proteins. Ancestors of the two major orders of plant pathogens in the Dothideomycetes, the Capnodiales and Pleosporales, may have had different modes of pathogenesis, with the former having fewer of these genes than the latter. Many of these genes are enriched in proximity to transposable elements, suggesting faster evolution because of the effects of repeat induced point (RIP) mutations. A syntenic block of genes, including oxidoreductases, is conserved in most Dothideomycetes and upregulated during infection in L. maculans, suggesting a possible function in response to oxidative stress.     

First Nuclear DNA C-values for Another 25 Angiosperm Families

Nuclear DNA C-value is an important genomic biodiversity characterwith many uses. An international workshop sponsored by Annalsof Botany and held at the Royal Botanic Gardens, Kew, UK, in1997 identified major gaps in our knowledge of plant DNA C-valuesand recommended targets for new work. Improved taxonomic coveragewas highlighted as a key need for angiosperms, especially atthe familial level. In 1997 C-values were known for only approx.32% of angiosperm families; a goal of complete familial representationby 2002 was recommended. A review published in 2000 (Bennettet al.;Annals of Botany86: 859–909) noted poor progresstowards this aim: of the 691 first C-values for species only12 (1.7%) were for unrepresented families. We began new workto address this in 1999, reporting first DNA C-values for 25angiosperm families in 2001 (Hanson et al.;Annals of Botany87:251–258). Here we report first DNA C-values for a further25 angiosperm families, increasing familial coverage in angiospermsto approx. 45%. Such targeting remains essential to approachthe goal set by the 1997 workshop of familial coverage for angiospermswithin 5 years. The 4C DNA amounts presented here range from0.76 pg (similar toArabidopsis thaliana ) in Roridula gorgonias(Roridulaceae)to 29.74 pg in Gunnera manicata(Gunneraceae). 1C values were< 3.5 pg in 23 of the 25 families; these data provide furthersupport for the view that ancestral angiosperms almost certainlyhad small genomes (defined as 1C     

Effect of 5 alpha-dihydrotestosterone on sexual receptivity and neural progestin receptors in ovariectomized rats given pulsed estradiol

Experiments were carried out to examine the mechanism whereby 5 alpha-dihydrotestosterone (DHT) antagonizes the stimulatory effects of estrogen plus progesterone (P) on sexual receptivity (lordosis) in the ovariectomized rat. Estradiol (E2; 1 microgram s.c. in 10% ethanol) was administered in a discontinuous (pulsed) treatment regimen thought to mimic phase requirements of estrogen action; two injections of E2 were given either 6 or 12 h apart (first injection, Hour 0). Progesterone (0.5 mg in oil) was injected at Hour 20, and behavioral testing occurred at Hour 24. Dihydrotestosterone (2.5 mg s.c. in 10% ethanol/propylene glycol) inhibited lordosis when it was given before (-12 or -3 h), between (+3, or -3 and +3 h), or after (+8 h) the two E2 injections, but was not effective when given at +20 h. Significant inhibition of E2 + P-induced lordosis was achieved by 2.5 but not 1.0 or 0.2 mg DHT at -3 h, while uterine weights in the same animals were reduced significantly by 2.5 and 1.0 mg DHT. Serum E2 and DHT concentrations peaked rapidly after injection, declining to near baseline by 3 and 12 h, respectively. Induction of cytosolic progestin receptors (cPR) in the preoptic area and medial basal hypothalamus by estrogen was not prevented by DHT when animals were given the two pulsed E2 injections or daily injections of estradiol benzoate, although P was able to override the inhibitory behavioral effects of DHT in the latter but not the former group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ovarian and haemolymph titres of ecdysteroid during the gonadotrophic cycle in Diploptera punctata

The free (non-conjugated) ecdysteroid in the ovaries during the first gonadotrophic cycle of Diploptera punctata was identified as 20-hydroxyecdysone. The hormone, quantified by radioimmunoassay and by ultraviolet absorbance, was detectable in the ovary toward the end of vitellogenesis; the quantity increased rapidly during chorion formation. Ovaries with chorionated eggs contained 67 μg of 20-hydroxyecdysone per g fresh weight. The haemolymph free-ecdysteroid, not identified physicochemically, was quantified by radioimmunoassays. The highest concentration was observed at adult emergence; the titre declined between days 1–3 and then remained at a relatively constant level through oviposition (which occurs between day 7 and 8); titres in pregnant females were higher. Ovariectomized females exhibited the same pattern of ecdysteroid titres in the haemolymph as the sham operated controls throughout the period corresponding to the first gonadotrophic cycle. Thus the ovary may not be the only source of haemolymph ecdysteroid related to reproduction in adult females.     

Effect of a conjugated linoleic acid and omega-3 fatty acid mixture on body composition and adiponectin

This study aimed to determine the effect of supplementation with conjugated linoleic acids (CLAs) plus n-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFAs) on body composition, adiposity, and hormone levels in young and older, lean and obese men. Young (31.4+/-3.9 years) lean (BMI, 23.6+/-1.5 kg/m2; n=13) and obese (BMI, 32.4+/-1.9 kg/m2; n=12) and older (56.5+/-4.6 years) lean (BMI, 23.6+/-1.5 kg/m2; n=20) and obese (BMI, 32.0+/-1.6 kg/m2; n=14) men participated in a double-blind placebo-controlled, randomized crossover study. Subjects received either 6 g/day control fat or 3 g/day CLA (50:50 cis-9, trans-11:trans-10, cis-12) and 3 g/day n-3 LC-PUFA for 12 weeks with a 12-week wash-out period between crossovers. Body composition was assessed by dual-energy X-ray absorptiometry. Fasting adiponectin, leptin, glucose, and insulin concentrations were measured and insulin resistance estimated by homeostasis model assessment for insulin resistance (HOMA-IR). In the younger obese subjects, CLA plus n-3 LC-PUFA supplementation compared with control fat did not result in increased abdominal fat and raised both fat-free mass (2.4%) and adiponectin levels (12%). CLA plus n-3 LC-PUFA showed no significant effects on HOMA-IR in any group but did increase fasting glucose in older obese subjects. In summary, supplementation with CLA plus n-3 LC-PUFA prevents increased abdominal fat mass and raises fat-free mass and adiponectin levels in younger obese individuals without deleteriously affecting insulin sensitivity, whereas these parameters in young and older lean and older obese individuals were unaffected, apart from increased fasting glucose in older obese men.