Genomic innovations, transcriptional plasticity and gene loss underlying the evolution and divergence of two highly polyphagous and invasive Helicoverpa pest species.

BACKGROUND: Helicoverpa armigera and Helicoverpa zea are major caterpillar pests of Old and New World agriculture, respectively. Both, particularly H. armigera, are extremely polyphagous, and H. armigera has developed resistance to many insecticides. Here we use comparative genomics, transcriptomics and resequencing to elucidate the genetic basis for their properties as pests. RESULTS: We find that, prior to their divergence about 1.5 Mya, the H. armigera/H. zea lineage had accumulated up to more than 100 more members of specific detoxification and digestion gene families and more than 100 extra gustatory receptor genes, compared to other lepidopterans with narrower host ranges. The two genomes remain very similar in gene content and order, but H. armigera is more polymorphic overall, and H. zea has lost several detoxification genes, as well as about 50 gustatory receptor genes. It also lacks certain genes and alleles conferring insecticide resistance found in H. armigera. Non-synonymous sites in the expanded gene families above are rapidly diverging, both between paralogues and between orthologues in the two species. Whole genome transcriptomic analyses of H. armigera larvae show widely divergent responses to different host plants, including responses among many of the duplicated detoxification and digestion genes. CONCLUSIONS: The extreme polyphagy of the two heliothines is associated with extensive amplification and neofunctionalisation of genes involved in host finding and use, coupled with versatile transcriptional responses on different hosts. H. armigera's invasion of the Americas in recent years means that hybridisation could generate populations that are both locally adapted and insecticide resistant.

Obesity leads to declines in motor skills across childhood.

BACKGROUND: Poor motor skills have been consistently linked with a higher body weight in childhood, but the causal direction of this association is not fully understood. This study investigated the temporal ordering between children's motor skills and weight status at 5 and 10 years. METHODS: Participants were 668 children (54% male) who were studied from infancy as part of an iron deficiency anaemia preventive trial and follow-up study in Santiago, Chile. All were healthy, full-term and weighing 3 kg or more at birth. Cross-lagged panel modelling was conducted to understand the temporal precedence between children's weight status and motor proficiency. Analyses also examined differences in gross and fine motor skills among healthy weight, overweight, and obese children. RESULTS: A higher BMI at 5 years contributed to declines in motor proficiency from 5 to 10 years. There was no support for the reverse, that is, poor motor skills at 5 years did not predict increases in relative weight from 5 to 10 years. Obesity at 5 years also predicted declines in motor proficiency. When compared with normal weight children, obese children had significantly poorer total and gross motor skills at both 5 and 10 years. Overweight children had poorer total and gross motor skills at 10 years only. The differences in total and gross motor skills among normal weight, overweight and obese children appear to increase with age. There were small differences in fine motor skill between obese and non-obese children at 5 years only. CONCLUSIONS: Obesity preceded declines in motor skills and not the reverse. Study findings suggest that early childhood obesity intervention efforts might help prevent declines in motor proficiency that, in turn, may positively impact children's physical activity and overall fitness levels.

Tumour-suppression function of KLF12 through regulation of anoikis.

Suppression of detachment-induced cell death, known as anoikis, is an essential step for cancer metastasis to occur. We report here that expression of KLF12, a member of the Kruppel-like family of transcription factors, is downregulated in lung cancer cell lines that have been selected to grow in the absence of cell adhesion. Knockdown of KLF12 in parental cells results in decreased apoptosis following cell detachment from matrix. KLF12 regulates anoikis by promoting the cell cycle transition through S phase and therefore cell proliferation. Reduced expression levels of KLF12 results in increased ability of lung cancer cells to form tumours in vivo and is associated with poorer survival in lung cancer patients. We therefore identify KLF12 as a novel metastasis-suppressor gene whose loss of function is associated with anoikis resistance through control of the cell cycle.

SETD2 loss-of-function promotes renal cancer branched evolution through replication stress and impaired DNA repair.

Defining mechanisms that generate intratumour heterogeneity and branched evolution may inspire novel therapeutic approaches to limit tumour diversity and adaptation. SETD2 (Su(var), Enhancer of zeste, Trithorax-domain containing 2) trimethylates histone-3 lysine-36 (H3K36me3) at sites of active transcription and is mutated in diverse tumour types, including clear cell renal carcinomas (ccRCCs). Distinct SETD2 mutations have been identified in spatially separated regions in ccRCC, indicative of intratumour heterogeneity. In this study, we have addressed the consequences of SETD2 loss-of-function through an integrated bioinformatics and functional genomics approach. We find that bi-allelic SETD2 aberrations are not associated with microsatellite instability in ccRCC. SETD2 depletion in ccRCC cells revealed aberrant and reduced nucleosome compaction and chromatin association of the key replication proteins minichromosome maintenance complex component (MCM7) and DNA polymerase δ hindering replication fork progression, and failure to load lens epithelium-derived growth factor and the Rad51 homologous recombination repair factor at DNA breaks. Consistent with these data, we observe chromosomal breakpoint locations are biased away from H3K36me3 sites in SETD2 wild-type ccRCCs relative to tumours with bi-allelic SETD2 aberrations and that H3K36me3-negative ccRCCs display elevated DNA damage in vivo. These data suggest a role for SETD2 in maintaining genome integrity through nucleosome stabilization, suppression of replication stress and the coordination of DNA repair.

SREBP maintains lipid biosynthesis and viability of cancer cells under lipid- and oxygen-deprived conditions and defines a gene signature associated with poor survival in glioblastoma multiforme.

Oxygen and nutrient limitation are common features of the tumor microenvironment and are associated with cancer progression and induction of metastasis. The inefficient vascularization of tumor tissue also limits the penetration of other serum-derived factors, such as lipids and lipoproteins, which can be rate limiting for cell proliferation and survival. Here we have investigated the effect of hypoxia and serum deprivation on sterol regulatory element-binding protein (SREBP) activity and the expression of lipid metabolism genes in human glioblastoma multiforme (GBM) cancer cells. We found that SREBP transcriptional activity was induced by serum depletion both in normoxic and hypoxic cells and that activation of SREBP was required to maintain the expression of fatty acid and cholesterol metabolism genes under hypoxic conditions. Moreover, expression of stearoyl-CoA desaturase, the enzyme required for the generation of mono-unsaturated fatty acids, and fatty acid-binding protein 7, a regulator of glioma stem cell function, was strongly dependent on SREBP function. Inhibition of SREBP function blocked lipid biosynthesis in hypoxic cancer cells and impaired cell survival under hypoxia and in a three-dimensional spheroid model. Finally, gene expression analysis revealed that SREBP defines a gene signature that is associated with poor survival in glioblastoma.

FOXO3a regulates reactive oxygen metabolism by inhibiting mitochondrial gene expression.

Forkhead transcription factors of the O class (FOXOs) are important targets of the phosphatidylinositol 3-kinase/Akt pathway, and are key regulators of the cell cycle, apoptosis and response to oxidative stress. FOXOs have been shown to have tumour suppressor function and are important for stem cell maintenance. We have performed a detailed analysis of the transcriptional programme induced in response to Forkhead-box protein O3a (FOXO3a) activation. We observed that FOXO3a activation results in the repression of a large number of nuclear-encoded genes with mitochondrial function. Repression of these genes was mediated by FOXO3a-dependent inhibition of c-Myc. FOXO3a activation also caused a reduction in mitochondrial DNA copy number, expression of mitochondrial proteins, respiratory complexes and mitochondrial respiratory activity. FOXO3a has been previously implicated in the detoxification of reactive oxygen species (ROS) through induction of manganese-containing superoxide dismutase (SOD2). We observed that reduction in ROS levels following FOXO3a activation was independent of SOD2, but required c-Myc inhibition. Hypoxia increases ROS production from the mitochondria, which is required for stabilisation of the hypoxia-inducible factor-1α (HIF-1α). FOXO3a activation blocked the hypoxia-dependent increase in ROS and prevented HIF-1α stabilisation. Our data suggest that FOXO factors regulate mitochondrial activity through inhibition of c-Myc function and alter the hypoxia response.

Extensive synteny conservation of holocentric chromosomes in Lepidoptera despite high rates of local genome rearrangements.

The recent assembly of the silkworm Bombyx mori genome with 432 Mb on 28 holocentric chromosomes has become a reference in the genomic analysis of the very diverse Order of Lepidoptera. We sequenced BACs from two major pests, the noctuid moths Helicoverpa armigera and Spodoptera frugiperda, corresponding to 15 regions distributed on 11 B. mori chromosomes, each BAC/region being anchored by known orthologous gene(s) to analyze syntenic relationships and genome rearrangements among the three species. Nearly 300 genes and numerous transposable elements were identified, with long interspersed nuclear elements and terminal inverted repeats the most abundant transposable element classes. There was a high degree of synteny conservation between B. mori and the two noctuid species. Conserved syntenic blocks of identified genes were very small, however, approximately 1.3 genes per block between B. mori and the two noctuid species and 2.0 genes per block between S. frugiperda and H. armigera. This corresponds to approximately two chromosome breaks per Mb DNA per My. This is a much higher evolution rate than among species of the Drosophila genus and may be related to the holocentric nature of the lepidopteran genomes. We report a large cluster of eight members of the aminopeptidase N gene family that we estimate to have been present since the Jurassic. In contrast, several clusters of cytochrome P450 genes showed multiple lineage-specific duplication events, in particular in the lepidopteran CYP9A subfamily. Our study highlights the value of the silkworm genome as a reference in lepidopteran comparative genomics.

Mitochondrial dysfunction triggered by loss of HtrA2 results in the activation of a brain-specific transcriptional stress response.

Cellular stress responses can be activated following functional defects in organelles such as mitochondria and the endoplasmic reticulum. Mitochondrial dysfunction caused by loss of the serine protease HtrA2 leads to a progressive movement disorder in mice and has been linked to parkinsonian neurodegeneration in humans. Here, we demonstrate that loss of HtrA2 results in transcriptional upregulation of nuclear genes characteristic of the integrated stress response, including the transcription factor CHOP, selectively in the brain. We also show that loss of HtrA2 results in the accumulation of unfolded proteins in the mitochondria, defective mitochondrial respiration and enhanced production of reactive oxygen species that contribute to the induction of CHOP expression and to neuronal cell death. CHOP expression is also significantly increased in Parkinson's disease patients' brain tissue. We therefore propose that this brain-specific transcriptional response to stress may be important in the advance of neurodegenerative diseases.

Txp40, a ubiquitous insecticidal toxin protein from Xenorhabdus and Photorhabdus bacteria.

Xenorhabdus and Photorhabdus are gram-negative bacteria that produce a range of proteins that are toxic to insects. We recently identified a novel 42-kDa protein from Xenorhabdus nematophila that was lethal to the larvae of insects such as Galleria mellonella and Helicoverpa armigera when it was injected at doses of 30 to 40 ng/g larvae. In the present work, the toxin gene txp40 was identified in another 59 strains of Xenorhabdus and Photorhabdus, indicating that it is both highly conserved and widespread among these bacteria. Recombinant toxin protein was shown to be active against a variety of insect species by direct injection into the larvae of the lepidopteran species G. mellonella, H. armigera, and Plodia interpunctella and the dipteran species Lucilia cuprina. The protein exhibited significant cytotoxicity against two dipteran cell lines and two lepidopteran cell lines but not against a mammalian cell line. Histological data from H. armigera larvae into which the toxin was injected suggested that the primary site of action of the toxin is the midgut, although some damage to the fat body was also observed.

cDNA cloning of a salivary chymotrypsin-like protease and the identification of six additional cDNAs encoding putative digestive proteases from the green mirid, Creontiades dilutus (Hemiptera: Miridae).

RT-PCR with degenerate primers was used to amplify partial cDNA fragments for one serine protease gene and three cysteine protease genes from poly(A) RNA isolated from the midgut of the green mirid, Creontiades dilutus. The serine protease amplicon showed homology to insect trypsin-like protease genes, and all three cysteine protease amplicons showed homology to cathepsin L-like protease genes.RT-PCR was also used to amplify fragments of three serine protease genes from salivary gland poly(A) RNA. One of these salivary gland serine protease amplicons was used to screen a whole organism cDNA library to isolate a full length cDNA clone, designated CdSp1 (Accession AY055753), which encodes a putative chymotrypsin-like protease. CdSp1 codes for a 293 amino acid protein that contains a signal peptide and activation peptide, as well as the catalytic triad present in all serine proteases and several of the binding pocket residues characteristic of chymotrypsins. In situ hybridisation showed that the transcript is expressed in the posterior lobe of the principal salivary gland, but not in the anterior lobe of the principal salivary gland, the accessory salivary gland or the midgut.

Risks among youths who have multiple sisters who were adolescent parents.

CONTEXT: Past research has revealed that having a sister who gave birth as a teenager is associated with increases in young people's likelihood of engaging in risky sexual behavior. To date, however, no study has determined if having several sisters who were adolescent mothers further raises youths' chances of engaging in risky activities. METHODS: Data were collected from 1,510 predominantly Hispanic and black 11-17-year-olds in a California program for youths who have at least one pregnant or parenting sister. Correlational analyses, analyses of variance and regression analyses were conducted to assess the effects of having multiple teenage parenting sisters on a variety of outcomes that are known risk factors for teenage pregnancy RESULTS: Twenty-four percent of participants had two or more sisters who had given birth as teenagers. The likelihood of having multiple adolescent parenting sisters was greatest in large families, but was unrelated to youths' other background characteristics. In analyses controlling for background factors, females with many parenting sisters had increased levels of behavioral problems (school problems, drug or alcohol use, and delinquent behavior) and an elevated likelihood of being sexually experienced. Having lived with two or more parenting sisters (as opposed to having lived with only one) was related to more permissive sexual and childbearing attitudes among young women and to earlier first intercourse among young men. Males with a sister who gave birth at a young age had elevated levels of delinquent behavior and promiscuous sexual behavior. CONCLUSIONS: As the number of teenage parenting sisters rises, youths'--particularly females'--risk of pregnancy involvement increases beyond the level associated with having only one teenage parenting sister. Screening for the number, living situation and age at first birth of parenting sisters is likely to be useful for programs seeking to identify youths at high risk of an early pregnancy.

The younger siblings of teenage mothers: a follow-up of their pregnancy risk.

This study followed 243 younger brothers and younger sisters of parenting teens and nonparenting teens across a 1.5-year period. The average age of siblings was 13.6 years at Time 1 and 15 years at Time 2. Relative to other youths, the sisters of parenting teens exhibited a sharp increase in drug and alcohol use and partying behavior across time and had the highest pregnancy rate at Time 2 (15%). The siblings of parenting teens spent 10 hr a week caring for their sisters' children, and, for girls, many hours of child care was associated with negative outcomes including permissive sexual behavior. Findings suggest that the younger sisters of parenting teens are at very high risk of early pregnancy and that this risk becomes increasingly pronounced across time.

Preliminary characterisation of digestive proteases of the green mirid, Creontiades dilutus (Hemiptera: Miridae).

Protease activities in the secreted saliva, salivary glands and midgut of the green mirid, Creontiades dilutus, were investigated. The saliva and salivary glands had more protease activity than the midgut, but no differences in protease activity levels were detected between male and female mirids, adult mirids and third instar nymphs, or between fed and starved mirids. In the salivary glands, chymotrypsin-like serine proteases predominated, as characterised by inhibitor specificity, basic pH optima, and hydrolysis of N-benzoyl-L-tyrosine p-nitroanilide and N-succinyl-ala-ala-pro-leu p-nitroanilide. The pH optimum of midgut extracts was acidic (pH 4), implying that acidic proteases predominate. However, protease activity was inhibited substantially by both aprotinin and E-64, suggesting the presence of both serine and cysteine proteases in the midgut of the green mirid.

Technical and economic strategies of maintenance dialysis in the UK and Nordic Countries.

The technical and economic strategies of maintenance dialysis in the UK and Nordic Countries are described. In particular the concept of 'Therapy Cost', or therapy pricing, as it is also known, in the UK and the concept of 'Patient Flow', which basically plots the flow of patients through the dialysis process, in the Nordic Region are discussed.

Characterisation of the helicostatin peptide precursor gene from Helicoverpa armigera (Lepidoptera: Noctuidae).

The gene encoding the helicostatin peptide family was isolated from a Helicoverpa armigera genomic DNA library. The deduced precursor sequence allowed unambiguous identification of all helicostatin peptides and verified the sequences of eight peptides previously isolated. The gene consists of at least three exons and encodes a precursor of 225 amino acids that contains three blocks of tandemly arranged helicostatin peptides including seven copies of the C-terminal sequence -YXFGL followed by a single Gly residue for carboxylamidation. Complete endoproteolytic processing at all possible dibasic cleavage sites would generate the seven helicostatin octapeptides previously purified from larval extracts. If processing was not complete at the third pair of basic amino acids the octadecapeptide (helicostatin IIa) would also be released. Two novel putative helicostatin peptide sequences were identified; YSKFNFGL and ERDMHRFSFGL, both of which had the C-terminal pentapeptide -FXFGL in place of the more usual -YXFGL sequence. Comparison of the helicostatin precursor with that of the cockroaches, locust and flies revealed variation in size, sequence and organisation of the 'allatostatin' precursors across different insect orders. In situ hybridisation histochemistry established that helicostatins are expressed in neurones of the central nervous system and endocrine cells of the midgut, indicating that the helicostatins are true brain-gut peptides. Northern blot analysis identified a single transcript of 1.6 kb in mRNA from whole larvae, isolated central nervous system and gut tissue.

Past physical abuse is significantly correlated with pregnancy as an adolescent.

STUDY OBJECTIVE: To determine which type or types of childhood abuse are associated with adolescent pregnancy and other adverse health outcomes. DESIGN: 100 females between the ages of 12 and 24 years of age, attending either an adolescent/young adult clinic (for ages 12 to 24), or teen obstetrics clinic (for ages 13 to 18) at an urban academic medical center, were recruited to participate in an interview study during their clinic visit. The clinician or research assistant used a 186-item structured interview form to record answers to detailed questions on abuse, in addition to traditional questions about the patient's feelings on home, education, activities, drugs and depression, sex and suicide (HEADS). RESULTS: 32% of subjects reported a history of sexual abuse, 29% reported physical abuse, and 46% reported past emotional abuse. The frequency of any past abuse was significantly higher in girls who had ever been pregnant (29 of 50, 58%), compared with "never pregnant" girls (19 of 49, 38%)( P < .05). A past history of physical abuse (P = .04), but not sexual or emotional abuse, was significantly associated with pregnancy as a teen. Other factors significantly correlated with teen pregnancy were: frequent alcohol use, older age, and Mexican-American or African-American ethnicity. Past history of abuse was significantly correlated with depression, suicidal thoughts, absent father, school failure, alcohol and tobacco use, and delinquency. CONCLUSIONS: A history of past physical abuse is strongly associated with adolescent pregnancy, and questions about all types of abuse should be routinely asked of adolescent patients.

Regulation of lepidopteran foregut movement by allatostatins and allatotropin from the frontal ganglion.

The frontal ganglion and associated neuronal pathways in larvae of the noctuid moth Helicoverpa armigera have been studied immunocytochemically with antisera against the endogenous neuropeptides, the allatostatins (helicostatins), and allatotropin. Two pairs of large ganglionic neurones contain allatostatin immunoreactivity, with the anteriormost of these pairs showing colocalisation with allatotropin. Allatostatin and allatotropin axons exit the frontal ganglion in the recurrent nerve and traverse the surface of the crop to give terminal arborisations around the stomodeal valve. There is a greater degree of lateral branching of allatotropin axons compared with allatostatin axons over the crop musculature. In vitro experiments show that the two types of peptides have antagonistic effects on the spontaneous myoactivity of the crop musculature. Allatotropin is myostimulatory at concentrations as low as 10(-16) M, enhancing both frequency and amplitude of peristaltic waves of contraction. All members of the helicostatin family inhibit peristalsis completely at concentrations of 10(-7)-10(-6) M and, to varying degrees, at 10(-10)-10(-8) M. On the basis of this evidence, it is suggested that peptidergic neurones of the frontal ganglion play a major part in regulating foregut motility through the antagonistic actions of the allatostatins and allatotropin.

Adolescents' pregnancy intentions: relations to life situations and caretaking behaviors prenatally and 2 years postpartum.

PURPOSE: This study explores if and how adolescents' pregnancy intentions relate to life situations and health-related behaviors prenatally and up to 2 years postpartum. METHODS: Adolescent girls who reported that they had "wanted a baby" (n = 75) as their reason for pregnancy were compared with those who reported that the pregnancy "just happened" (n = 79) at four separate time periods: prenatally, at 6 and 24 months postpartum, and at 18 months postpartum for teens who became pregnant again subsequent to the study pregnancy. RESULTS: Those who stated that they wanted a baby were more likely to be Hispanic, married, and out of school before becoming pregnant. They were less likely to receive welfare as their primary means of support and to have run away from home in the past than teens who stated that their pregnancy just happened. Self-reported reason for pregnancy was unrelated to repeat pregnancy by 18 months postpartum, but those who had wanted the study baby were less likely to undergo elective termination of a subsequent pregnancy and less likely to become pregnant by a different partner. The groups diverged at 24 months postpartum when those who wanted a baby were more likely to be married to the father of the baby, be financially supported by him, receive child care assistance from him, and have attempted or succeeded at breastfeeding the study child. CONCLUSION: Self-reported reason for pregnancy reveals many important characteristics of pregnant adolescents both at the time of presentation and up to 2 years postpartum. Young women in this study who reported intentional pregnancy seem to fare better with regard to their financial status and their relationship with the father of the baby.

Subacute and chronic toxicity studies of triethylenetetramine dihydrochloride (TJA-250) by oral administration to F-344 rats.

Triethylenetetramine dihydrochloride (trientine-2HCl, TJA-250), a copper chelating agent used to treat Wilson's disease, was administered orally to male and female F-344 rats for 4 or 8 weeks at dosages of 0, 100, 350 or 1200 mg/kg/day or for 26 weeks at dosages of 50, 175 or 600 mg/kg/day. 4 or 8-week study. Two males receiving 1200 mg/kg/day died during week 8 of treatment. In males receiving 1200 mg/kg/day during weeks 5 to 8 of treatment, body weight gain and food consumption were decreased and hunched posture and thin build were observed. During week 4 or 8 of treatment urinalysis revealed, for males receiving 100 mg/kg/day or animals receiving 350 mg/kg/day or more, increased electrolyte outputs possibly due to the hydrochloride nature of trientine-2HCl, with low plasma alkaline phosphatase activities evident in animals receiving 350 or 1200 mg/kg/day. After 4 and 8 weeks, and during 8 weeks of treatment, high lung weights and bronchiolar epithelium hypertrophy and broncho-alveolar pneumonia were recorded for animals receiving 1200 mg/kg/day, and submucosal acute inflammation within the glandular region of the stomach was recorded for males receiving 350 or 1200 mg/kg/day and in all treated female groups. 26-week study. One male receiving 175 mg/kg/day and three males receiving 600 mg/kg/day died, showing lung changes. The body weight gain of animals receiving 600 mg/kg/day was slightly decreased. Blood chemistry and urinalysis examinations showed changes similar to those indicated in the 4- or 8-week study. The low plasma copper concentrations seen in males receiving 600 mg/kg/day, the slightly low liver copper concentrations found in animals receiving 600 or 175 mg/kg/day and the high urinary copper concentrations found in all treated groups, are attributed to the pharmacological action of trientine-2HCl. Histopathology revealed a dosage-related incidence and severity of focal chronic interstitial pneumonitis accompanied by fibrosis of the alveolar walls in females receiving 175 mg/kg/day or more and all treated male groups, but no significant pathological changes in the stomach. Apart from the histological changes found in the lung, all the above changes were reversible. In conclusion, the NOAEL of trientine-2HCl in this 26-week study was considered to be 50 mg/kg/day for females and less than 50 mg/kg/day for males.