A Population-Based Study of Genes Previously Implicated in Breast Cancer.

BACKGROUND: Population-based estimates of the risk of breast cancer associated with germline pathogenic variants in cancer-predisposition genes are critically needed for risk assessment and management in women with inherited pathogenic variants. METHODS: In a population-based case-control study, we performed sequencing using a custom multigene amplicon-based panel to identify germline pathogenic variants in 28 cancer-predisposition genes among 32,247 women with breast cancer (case patients) and 32,544 unaffected women (controls) from population-based studies in the Cancer Risk Estimates Related to Susceptibility (CARRIERS) consortium. Associations between pathogenic variants in each gene and the risk of breast cancer were assessed. RESULTS: Pathogenic variants in 12 established breast cancer-predisposition genes were detected in 5.03% of case patients and in 1.63% of controls. Pathogenic variants in BRCA1 and BRCA2 were associated with a high risk of breast cancer, with odds ratios of 7.62 (95% confidence interval [CI], 5.33 to 11.27) and 5.23 (95% CI, 4.09 to 6.77), respectively. Pathogenic variants in PALB2 were associated with a moderate risk (odds ratio, 3.83; 95% CI, 2.68 to 5.63). Pathogenic variants in BARD1, RAD51C, and RAD51D were associated with increased risks of estrogen receptor-negative breast cancer and triple-negative breast cancer, whereas pathogenic variants in ATM, CDH1, and CHEK2 were associated with an increased risk of estrogen receptor-positive breast cancer. Pathogenic variants in 16 candidate breast cancer-predisposition genes, including the c.657_661del5 founder pathogenic variant in NBN, were not associated with an increased risk of breast cancer. CONCLUSIONS: This study provides estimates of the prevalence and risk of breast cancer associated with pathogenic variants in known breast cancer-predisposition genes in the U.S. population. These estimates can inform cancer testing and screening and improve clinical management strategies for women in the general population with inherited pathogenic variants in these genes. (Funded by the National Institutes of Health and the Breast Cancer Research Foundation.).

Augmentation of tumor expression of HLA-DR, CXCL9, and CXCL10 may improve olfactory neuroblastoma immunotherapeutic responses.

BACKGROUND: Olfactory neuroblastoma is a rare malignancy of the anterior skull base typically treated with surgery and adjuvant radiation. Although outcomes are fair for low-grade disease, patients with high-grade, recurrent, or metastatic disease oftentimes respond poorly to standard treatment methods. We hypothesized that an in-depth evaluation of the olfactory neuroblastoma tumor immune microenvironment would identify mechanisms of immune evasion in high-grade olfactory neuroblastoma as well as rational targetable mechanisms for future translational immunotherapeutic approaches. METHODS: Multispectral immunofluorescence and RNAScope evaluation of the tumor immune microenvironment was performed on forty-seven clinically annotated olfactory neuroblastoma samples. A retrospective chart review was performed and clinical correlations assessed. RESULTS: A significant T cell infiltration was noted in olfactory neuroblastoma samples with a stromal predilection, presence of myeloid-derived suppressor cells, and sparse natural killer cells. A striking decrease was observed in MHC-I expression in high-grade olfactory neuroblastoma compared to low-grade disease, representing a mechanism of immune evasion in high-grade disease. Mechanistically, the immune effector stromal predilection appears driven by low tumor cell MHC class II (HLA-DR), CXCL9, and CXCL10 expression as those tumors with increased tumor cell expression of each of these mediators correlated with significant increases in T cell infiltration. CONCLUSION: These data suggest that immunotherapeutic strategies that augment tumor cell expression of MHC class II, CXCL9, and CXCL10 may improve parenchymal trafficking of immune effector cells in olfactory neuroblastoma and augment immunotherapeutic responses.

Increased interaction between endoplasmic reticulum and mitochondria following sleep deprivation.

BACKGROUND: Prolonged cellular activity may overload cell function, leading to high rates of protein synthesis and accumulation of misfolded or unassembled proteins, which cause endoplasmic reticulum (ER) stress and activate the unfolded protein response (UPR) to re-establish normal protein homeostasis. Previous molecular work has demonstrated that sleep deprivation (SD) leads to ER stress in neurons, with a number of ER-specific proteins being upregulated to maintain optimal cellular proteostasis. It is still not clear which cellular processes activated by sleep deprivation lead to ER- stress, but increased cellular metabolism, higher request for protein synthesis, and over production of oxygen radicals have been proposed as potential contributing factors. Here, we investigate the transcriptional and ultrastructural ER and mitochondrial modifications induced by sleep loss. RESULTS: We used gene expression analysis in mouse forebrains to show that SD was associated with significant transcriptional modifications of genes involved in ER stress but also in ER-mitochondria interaction, calcium homeostasis, and mitochondrial respiratory activity. Using electron microscopy, we also showed that SD was associated with a general increase in the density of ER cisternae in pyramidal neurons of the motor cortex. Moreover, ER cisternae established new contact sites with mitochondria, the so-called mitochondria associated membranes (MAMs), important hubs for molecule shuttling, such as calcium and lipids, and for the modulation of ATP production and redox state. Finally, we demonstrated that Drosophila male mutant flies (elav > linker), in which the number of MAMs had been genetically increased, showed a reduction in the amount and consolidation of sleep without alterations in the homeostatic sleep response to SD. CONCLUSIONS: We provide evidence that sleep loss induces ER stress characterized by increased crosstalk between ER and mitochondria. MAMs formation associated with SD could represent a key phenomenon for the modulation of multiple cellular processes that ensure appropriate responses to increased cell metabolism. In addition, MAMs establishment may play a role in the regulation of sleep under baseline conditions.

Phylogenomics revealed migration routes and adaptive radiation timing of Holarctic malaria mosquito species of the Maculipennis Group.

BACKGROUND: Phylogenetic analyses of closely related species of mosquitoes are important for better understanding the evolution of traits contributing to transmission of vector-borne diseases. Six out of 41 dominant malaria vectors of the genus Anopheles in the world belong to the Maculipennis Group, which is subdivided into two Nearctic subgroups (Freeborni and Quadrimaculatus) and one Palearctic (Maculipennis) subgroup. Although previous studies considered the Nearctic subgroups as ancestral, details about their relationship with the Palearctic subgroup, and their migration times and routes from North America to Eurasia remain controversial. The Palearctic species An. beklemishevi is currently included in the Nearctic Quadrimaculatus subgroup adding to the uncertainties in mosquito systematics. RESULTS: To reconstruct historic relationships in the Maculipennis Group, we conducted a phylogenomic analysis of 11 Palearctic and 2 Nearctic species based on sequences of 1271 orthologous genes. The analysis indicated that the Palearctic species An. beklemishevi clusters together with other Eurasian species and represents a basal lineage among them. Also, An. beklemishevi is related more closely to An. freeborni, which inhabits the Western United States, rather than to An. quadrimaculatus, a species from the Eastern United States. The time-calibrated tree suggests a migration of mosquitoes in the Maculipennis Group from North America to Eurasia about 20-25 million years ago through the Bering Land Bridge. A Hybridcheck analysis demonstrated highly significant signatures of introgression events between allopatric species An. labranchiae and An. beklemishevi. The analysis also identified ancestral introgression events between An. sacharovi and its Nearctic relative An. freeborni despite their current geographic isolation. The reconstructed phylogeny suggests that vector competence and the ability to enter complete diapause during winter evolved independently in different lineages of the Maculipennis Group. CONCLUSIONS: Our phylogenomic analyses reveal migration routes and adaptive radiation timing of Holarctic malaria vectors and strongly support the inclusion of An. beklemishevi into the Maculipennis Subgroup. Detailed knowledge of the evolutionary history of the Maculipennis Subgroup provides a framework for examining the genomic changes related to ecological adaptation and susceptibility to human pathogens. These genomic variations may inform researchers about similar changes in the future providing insights into the patterns of disease transmission in Eurasia.

Exploiting docetaxel-induced tumor cell necrosis with tumor targeted delivery of IL-12.

There is strong evidence that chemotherapy can induce tumor necrosis which can be exploited for the targeted delivery of immuno-oncology agents into the tumor microenvironment (TME). We hypothesized that docetaxel, a chemotherapeutic agent that induces necrosis, in combination with the bifunctional molecule NHS-IL-12 (M9241), which delivers recombinant IL-12 through specific targeting of necrotic regions in the tumor, would provide a significant antitumor benefit in the poorly inflamed murine tumor model, EMT6 (breast), and in the moderately immune-infiltrated tumor model, MC38 (colorectal). Docetaxel, as monotherapy or in combination with NHS-IL-12, promoted tumor necrosis, leading to the improved accumulation and retention of NHS-IL-12 in the TME. Significant antitumor activity and prolonged survival were observed in cohorts receiving docetaxel and NHS-IL-12 combination therapy in both the MC38 and EMT6 murine models. The therapeutic effects were associated with increased tumor infiltrating lymphocytes and were dependent on CD8+ T cells. Transcriptomics of the TME of mice receiving the combination therapy revealed the upregulation of genes involving crosstalk between innate and adaptive immunity factors, as well as the downregulation of signatures of myeloid cells. In addition, docetaxel and NHS-IL-12 combination therapy effectively controlled tumor growth of PD-L1 wild-type and PD-L1 knockout MC38 in vivo, implying this combination could be applied in immune checkpoint refractory tumors, and/or tumors regardless of PD-L1 status. The data presented herein provide the rationale for the design of clinical studies employing this combination or similar combinations of agents.

Patterns of genetic differentiation imply distinct phylogeographic history of the mosquito species Anopheles messeae and Anopheles daciae in Eurasia.

Detailed knowledge of phylogeography is important for control of mosquito species involved in the transmission of human infectious diseases. Anopheles messeae is a geographically widespread and genetically diverse dominant vector of malaria in Eurasia. A closely related species, An. daciae, was originally distinguished from An. messeae based on five nucleotide substitutions in its ribosomal DNA (rDNA). However, the patterns of phylogeographic history of these species in Eurasia remain poorly understood. Here, using internal transcribed spacer 2 (ITS2) of rDNA and karyotyping for the species identification we determined the composition of five Anopheles species in 28 locations in Eurasia. Based on the frequencies of 11 polymorphic chromosomal inversions used as genetic markers, a large-scale population genetics analysis was performed of 1932 mosquitoes identified as An. messeae, An. daciae and their hybrids. The largest genetic differences between the species were detected in the X sex chromosome suggesting a potential involvement of this chromosome in speciation. The frequencies of autosomal inversions in the same locations differed by 13%-45% between the species demonstrating a restricted gene flow between the species. Overall, An. messeae was identified as a diverse species with a more complex population structure than An. daciae. The clinal gradients in frequencies of chromosomal inversions were determined in both species implicating their possible involvement in climate adaptations. The frequencies of hybrids were low ~1% in northern Europe but high up to 50% in south-eastern populations. Thus, our study revealed critical differences in patterns of phylogeographic history between An. messeae and An. daciae in Eurasia. This knowledge will help to predict the potential of the malaria transmission in the northern territories of the continent.

Targeting Immunosuppressive Adenosine Signaling: A Review of Potential Immunotherapy Combination Strategies.

The tumor microenvironment regulates many aspects of cancer progression and anti-tumor immunity. Cancer cells employ a variety of immunosuppressive mechanisms to dampen immune cell function in the tumor microenvironment. While immunotherapies that target these mechanisms, such as immune checkpoint blockade, have had notable clinical success, resistance is common, and there is an urgent need to identify additional targets. Extracellular adenosine, a metabolite of ATP, is found at high levels in the tumor microenvironment and has potent immunosuppressive properties. Targeting members of the adenosine signaling pathway represents a promising immunotherapeutic modality that can potentially synergize with conventional anti-cancer treatment strategies. In this review, we discuss the role of adenosine in cancer, present preclinical and clinical data on the efficacy adenosine pathway inhibition, and discuss possible combinatorial approaches.

Effects of Eph/ephrin signalling and human Alzheimer's disease-associated EphA1 on Drosophila behaviour and neurophysiology.

Alzheimer's disease (AD) is the most prevalent neurodegenerative disease placing a great burden on people living with it, carers and society. Yet, the underlying patho-mechanisms remain unknown and treatments limited. To better understand the molecular changes associated with AD, genome-wide association studies (GWAS) have identified hundreds of candidate genes linked to the disease, like the receptor tyrosine kinase EphA1. However, demonstration of whether and how these genes cause pathology is largely lacking. Here, utilising fly genetics, we generated the first Drosophila model of human wild-type and P460L mutant EphA1 and tested the effects of Eph/ephrin signalling on AD-relevant behaviour and neurophysiology. We show that EphA1 mis-expression did not cause neurodegeneration, shorten lifespan or affect memory but flies mis-expressing the wild-type or mutant receptor were hyper-aroused, had reduced sleep, a stronger circadian rhythm and increased clock neuron activity and excitability. Over-expression of endogenous fly Eph and RNAi-mediated knock-down of Eph and its ligand ephrin affected sleep architecture and neurophysiology. Eph over-expression led to stronger circadian morning anticipation while ephrin knock-down impaired memory. A dominant negative form of the GTPase Rho1, a potential intracellular effector of Eph, led to hyper-aroused flies, memory impairment, less anticipatory behaviour and neurophysiological changes. Our results demonstrate a role of Eph/ephrin signalling in a range of behaviours affected in AD. This presents a starting point for studies into the underlying mechanisms of AD including interactions with other AD-associated genes, like Rho1, Ankyrin, Tau and APP with the potential to identify new targets for treatment.

Prospective investigation of herpesvirus infection and risk of glioma.

Glioma is an aggressive neoplasm of the brain with poorly understood etiology. A limited number of pathogens have been examined as glioma risk factors, but data from prospective studies with infection status determined before disease are lacking. Herpesviruses comprise a large family of DNA viruses that infect humans and are linked to a range of chronic diseases. We conducted a prospective evaluation of the association between antibody to six human herpesviruses and glioma risk in the Janus Serum Bank (Janus) and the Cancer Prevention Study-II (CPS-II). In Janus and CPS-II, the risk for glioma was not related to seroprevalence of herpes simplex virus-1, varicella zoster virus, or human herpes viruses 6A or 6B. In Janus, seropositivity to either the Epstein Barr virus (EBV) EA[D] or VCAp18 antigen was associated with a lower risk of glioma (ORs: 0.55 [95% CI 0.32-0.94] and 0.57 [95% CI 0.38-0.85]). This inverse association was consistent by histologic subtype and was observed for gliomas diagnosed up to two decades following antibody measurement. In Janus, seropositivity to at least one of three examined cytomegalovirus (CMV) antigens (pp150, pp52, pp28) was associated with an increased risk of nonglioblastoma (OR: 2.08 [95% CI 1.07-4.03]). This association was limited to tumors diagnosed within 12 years of antibody measurement. In summary, we report evidence of an inverse association between exposure to EBV and glioma. We further report that CMV exposure may be related to a higher likelihood of the nonglioblastoma subtype.

In vivo expansion of functionally integrated GABAergic interneurons by targeted increase in neural progenitors.

A central hypothesis for brain evolution is that it might occur via expansion of progenitor cells and subsequent lineage-dependent formation of neural circuits. Here, we report in vivo amplification and functional integration of lineage-specific circuitry in Drosophila Levels of the cell fate determinant Prospero were attenuated in specific brain lineages within a range that expanded not only progenitors but also neuronal progeny, without tumor formation. Resulting supernumerary neural stem cells underwent normal functional transitions, progressed through the temporal patterning cascade, and generated progeny with molecular signatures matching source lineages. Fully differentiated supernumerary gamma-amino butyric acid (GABA)-ergic interneurons formed functional connections in the central complex of the adult brain, as revealed by in vivo calcium imaging and open-field behavioral analysis. Our results show that quantitative control of a single transcription factor is sufficient to tune neuron numbers and clonal circuitry, and provide molecular insight into a likely mechanism of brain evolution.

Hereditary spastic paraparesis presenting as cerebral palsy due to ADD3 variant with mechanistic insight provided by a Drosophila γ-adducin model.

Cerebral palsy (CP) causes neurological disability in early childhood. Hypoxic-ischaemic injury plays a major role in its aetiology, nevertheless, genetic and epigenetic factors may contribute to the clinical presentation. Mutations in ADD3 (encoding γ-adducin) gene have been described in a monogenic form of spastic quadriplegic cerebral palsy (OMIM 601568). We studied a 16-year-old male with spastic diplegia. Several investigations including neurometabolic testing, brain and spine magnetic resonance imaging (MRI) and CGH-Array were normal. Further, clinical genetics assessment and whole exome sequencing (WES) gave the diagnosis. We generated an animal model using Drosophila to study the effects of γ-adducin loss and gain of function. WES revealed a biallelic variant in the ADD3 gene, NM_016824.5(ADD3): c.1100G > A, p.(Gly367Asp). Mutations in this gene have been described as an ultra-rare autosomal recessive, which is a known form of inherited cerebral palsy. Molecular modelling suggests that this mutation leads to a loss of structural integrity of γ-adducin and is therefore expected to result in a decreased level of functional protein. Pan-neuronal over-expression or knock-down of the Drosophila ortholog of ADD3 called hts caused a reduction of life span and impaired locomotion thereby phenocopying aspects of the human disease. Our animal experiments present a starting point to understand the biological processes underpinning the clinical phenotype and pathogenic mechanisms, to gain insights into potential future methods for treating or preventing ADD3 related spastic quadriplegic cerebral palsy.

The Drosophila ortholog of the schizophrenia-associated CACNA1A and CACNA1B voltage-gated calcium channels regulate memory, sleep and circadian rhythms.

Schizophrenia exhibits up to 80% heritability. A number of genome wide association studies (GWAS) have repeatedly shown common variants in voltage-gated calcium (Cav) channel genes CACNA1C, CACNA1I and CACNA1G have a major contribution to the risk of the disease. More recently, studies using whole exome sequencing have also found that CACNA1B (Cav2.2 N-type) deletions and rare disruptive variants in CACNA1A (Cav2.1 P/Q-type) are associated with schizophrenia. The negative symptoms of schizophrenia include behavioural defects such as impaired memory, sleep and circadian rhythms. It is not known how variants in schizophrenia-associated genes contribute to cognitive and behavioural symptoms, thus hampering the development of treatment for schizophrenia symptoms. In order to address this knowledge gap, we studied behavioural phenotypes in a number of loss of function mutants for the Drosophila ortholog of the Cav2 gene family called cacophony (cac). cac mutants showed several behavioural features including decreased night-time sleep and hyperactivity similar to those reported in human patients. The change in timing of sleep-wake cycles suggested disrupted circadian rhythms, with the loss of night-time sleep being caused by loss of cac just in the circadian clock neurons. These animals also showed a reduction in rhythmic circadian behaviour a phenotype that also could be mapped to the central clock. Furthermore, reduction of cac just in the clock resulted in a lengthening of the 24 h period. In order to understand how loss of Cav2 function may lead to cognitive deficits and underlying cellular pathophysiology we targeted loss of function of cac to the memory centre of the fly, called the mushroom bodies (MB). This manipulation was sufficient to cause reduction in both short- and intermediate-term associative memory. Memory impairment was accompanied by a decrease in Ca2+ transients in response to a depolarizing stimulus, imaged in the MB presynaptic terminals. This work shows loss of cac Cav2 channel function alone causes a number of cognitive and behavioural deficits and underlying reduced neuronal Ca2+ transients, establishing Drosophila as a high-throughput in vivo genetic model to study the Cav channel pathophysiology related to schizophrenia.

Toxoplasma gondii infection and the risk of adult glioma in two prospective studies.

Toxoplasma gondii (T gondii) is a common parasite that shows affinity to neural tissue and may lead to the formation of cysts in the brain. Previous epidemiologic studies have suggested an association between glioma and increased prevalence of T gondii infection, but prospective studies are lacking. Therefore, we examined the association between prediagnostic T gondii antibodies and risk of glioma in two prospective cohorts using a nested case-control study design. Cases and matched controls were selected from the American Cancer Society's Cancer Prevention Study-II Nutrition Cohort (CPSII-NC) (n = 37 cases and 74 controls) and the Norwegian Cancer Registry's Janus Serum Bank (Janus) (n = 323 cases and 323 controls). Blood samples collected prior to diagnosis were analyzed for antibodies to two T gondii surface antigens (p22 and sag-1), with individuals considered seropositive if antibodies to either antigen were detected. Conditional logistic regression was used to calculate odds ratios (OR) and 95% confidence intervals (95% CI) for each cohort. In both cohorts, a suggestive increase in glioma risk was observed among those infected with T gondii (OR: 2.70; 95% CI: 0.96-7.62 for CPSII-NC; OR: 1.32, 95% CI: 0.85-2.07 for Janus), particularly among participants with high antibody titers specific to the sag-1 antigen (CPSII-NC OR: 3.35, 95% CI: 0.99-11.38; Janus OR: 1.79, 95% CI: 1.02-3.14). Our findings provide the first prospective evidence of an association between T gondii infection and risk of glioma. Further studies with larger case numbers are needed to confirm a potential etiologic role for T gondii in glioma.

Impaired Pre-Motor Circuit Activity and Movement in a Drosophila Model of KCNMA1-Linked Dyskinesia.

BACKGROUND: Paroxysmal dyskinesias (PxDs) are characterized by involuntary movements and altered pre-motor circuit activity. Causative mutations provide a means to understand the molecular basis of PxDs. Yet in many cases, animal models harboring corresponding mutations are lacking. Here we utilize the fruit fly, Drosophila, to study a PxD linked to a gain-of-function (GOF) mutation in the KCNMA1/hSlo1 BK potassium channel. OBJECTIVES: We aimed to recreate the equivalent BK (big potassium) channel mutation in Drosophila. We sought to determine how this mutation altered action potentials (APs) and synaptic release in vivo; to test whether this mutation disrupted pre-motor circuit function and locomotion; and to define neural circuits involved in locomotor disruption. METHODS: We generated a knock-in Drosophila model using homologous recombination. We used electrophysiological recordings and calcium-imaging to assess AP shape, neurotransmission, and the activity of the larval pre-motor central pattern generator (CPG). We used video-tracking and automated systems to measure movement, and developed a genetic method to limit BK channel expression to defined circuits. RESULTS: Neuronal APs exhibited reduced width and an enhanced afterhyperpolarization in the PxD model. We identified calcium-dependent reductions in neurotransmitter release, dysfunction of the CPG, and corresponding alterations in movement, in model larvae. Finally, we observed aberrant locomotion and dyskinesia-like movements in adult model flies, and partially mapped the impact of GOF BK channels on movement to cholinergic neurons. CONCLUSION: Our model supports a link between BK channel GOF and hyperkinetic movements, and provides a platform to dissect the mechanistic basis of PxDs. © 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

The new Internal Transcribed Spacer 2 diagnostic tool clarifies the taxonomic position and geographic distribution of the North American malaria vector Anopheles punctipennis.

BACKGROUND: The malaria mosquito Anopheles punctipennis, a widely distributed species in North America, is capable of transmitting human malaria and is actively involved in the transmission of the ungulate malaria parasite Plasmodium odocoilei. However, molecular diagnostic tools based on Internal Transcribed Spacer 2 (ITS2) of ribosomal DNA are lacking for this species. Anopheles punctipennis is a former member of the Anopheles maculipennis complex but its systematic position remains unclear. METHODS: In this study, ITS2 sequences were obtained from 276 An. punctipennis specimens collected in the eastern and midwestern United States and a simple and robust Restriction Fragment Length Polymorphism approach for species identification was developed. The maximum-likelihood phylogenetic tree was constructed based on ITS2 sequences available through this study and from GenBank for 20 species of Anopheles. RESULTS: The analysis demonstrated a consistent ITS2 sequence length and showed no indications of intragenomic variation among the samples based on ITS2, suggesting that An. punctipennis represents a single species in the studied geographic locations. In this study, An. punctipennis was found in urban, rural, and forest settings, suggesting its potential broad role in pathogen transmission. Phylogeny based on ITS2 sequence comparison demonstrated the close relationship of this species with other members of the Maculipennis group. CONCLUSIONS: This study developed molecular tools based on ITS2 sequences for the malaria vector An. punctipennis and clarified the phylogenetic position of the species within the Maculipennis group.

Drosophila Ionotropic Receptor 25a mediates circadian clock resetting by temperature.

Circadian clocks are endogenous timers adjusting behaviour and physiology with the solar day. Synchronized circadian clocks improve fitness and are crucial for our physical and mental well-being. Visual and non-visual photoreceptors are responsible for synchronizing circadian clocks to light, but clock-resetting is also achieved by alternating day and night temperatures with only 2-4 °C difference. This temperature sensitivity is remarkable considering that the circadian clock period (~24 h) is largely independent of surrounding ambient temperatures. Here we show that Drosophila Ionotropic Receptor 25a (IR25a) is required for behavioural synchronization to low-amplitude temperature cycles. This channel is expressed in sensory neurons of internal stretch receptors previously implicated in temperature synchronization of the circadian clock. IR25a is required for temperature-synchronized clock protein oscillations in subsets of central clock neurons. Extracellular leg nerve recordings reveal temperature- and IR25a-dependent sensory responses, and IR25a misexpression confers temperature-dependent firing of heterologous neurons. We propose that IR25a is part of an input pathway to the circadian clock that detects small temperature differences. This pathway operates in the absence of known 'hot' and 'cold' sensors in the Drosophila antenna, revealing the existence of novel periphery-to-brain temperature signalling channels.

Erythrocyte levels of cadmium and lead and risk of B-cell non-Hodgkin lymphoma and multiple myeloma.

Cadmium and lead are persistent environmental toxins that are known or probable carcinogens, based on evidence for causality for nonhematologic cancers. Associations of these metals with risk of non-Hodgkin lymphoma (NHL) and multiple myeloma (MM) are unknown but biologically plausible. To examine the associations of circulating levels of lead and cadmium exposure with risk of B-cell NHL (B-NHL) and multiple myeloma, we conducted a nested case-control study among 299 incident B-cell NHLs and 76 MM cases within the Cancer Prevention Study-II Nutrition Cohort (CPS-II NC). Each case was incidence-density matched to two eligible controls on age, race, sex and blood draw date. Conditional logistic regression was used to estimate relative risks (RR) and 95% confidence intervals (CI) for lymphoid malignancies overall and stratified by subtype. We observed a significant positive association between high erythrocyte lead concentration and risk of lymphoid malignancies overall (RR = 1.16, 95% CI: 1.02-1.33 per 17.6 µg/L (1 standard deviation [SD])) and follicular lymphoma in particular (RR = 1.80, 95% CI: 1.15-2.80 per SD). In contrast, there was no association between erythrocyte cadmium and risk of B-NHL (RR = 0.89, 95% CI: 0.75-1.06 per 0.37 µg/L [1 SD]), or any B-NHL subtypes; but a strong inverse association with MM risk (RR = 0.59, 95% CI: 0.38-0.89, per SD). Results from our study suggest a positive association between erythrocyte lead level and risk of lymphoid malignancies and a possible inverse association between cadmium and myeloma. Additional research is needed to confirm and further explore these findings.

Relationship Between Muscle-Strengthening Activity and Cause-Specific Mortality in a Large US Cohort.

INTRODUCTION: Muscle-strengthening activity (MSA) has beneficial effects on hypertension, glucose homeostasis, and other health conditions; however, its association with mortality is not as well understood. METHODS: We analyzed data from the Cancer Prevention Study-II Nutrition Cohort (data collection 1982-2014), a prospective US cohort that consisted of 72,462 men and women who were free of major chronic diseases; 18,034 of the cohort died during 13 years of follow-up (2001-2014). We used Cox proportional hazards modeling, controlling for various potential confounding factors, to calculate hazard ratios (HRs) and 95% confidence intervals (CIs) for MSA (none, >0 to <1 h/wk, 1 to <2 h/wk, and ≥2 h/wk) in relation to mortality risk, independent of and in combination with aerobic physical activity. RESULTS: The association between MSA and mortality appeared to be nonlinear (quadratic trend P value, <.001). After multivariable adjustment and comparison with no MSA, engaging in less than 2 hours per week of MSA was associated with lowered all-cause mortality (>0 to <1 h/wk: HR = 0.88, 95% CI, 0.82-0.94; 1 to <2 h/wk: HR = 0.90, 95% CI, 0.84-0.97), but engaging in 2 or more hours per week was not associated with reduced risk (HR = 1.01; 95% CI, 0.92-1.09). Associations were similar but not significant for cancer mortality. Engaging in >0 to <1 hr/wk of MSA was associated with a 19% lower risk (HR = 0.81; 95% CI, 0.71-0.92) of cardiovascular disease mortality, but more time spent in MSA was not associated with reduced risk (quadratic trend P value =.005). Associations did not vary by amount of moderate-to-vigorous aerobic physical activity. CONCLUSION: Engaging in ≥2 hours per week of MSA was associated with lower all-cause mortality, independent of aerobic activity. Reasons for the lack of association with higher amounts of MSA are unclear. Our findings support recommending muscle-strengthening activities for overall health.

Shaw and Shal voltage-gated potassium channels mediate circadian changes in Drosophila clock neuron excitability.

As in mammals, Drosophila circadian clock neurons display rhythms of activity with higher action potential firing rates and more positive resting membrane potentials during the day. This rhythmic excitability has been widely observed but, critically, its regulation remains unresolved. We have characterized and modelled the changes underlying these electrical activity rhythms in the lateral ventral clock neurons (LNvs). We show that currents mediated by the voltage-gated potassium channels Shaw (Kv3) and Shal (Kv4) oscillate in a circadian manner. Disruption of these channels, by expression of dominant negative (DN) subunits, leads to changes in circadian locomotor activity and shortens lifespan. LNv whole-cell recordings then show that changes in Shaw and Shal currents drive changes in action potential firing rate and that these rhythms are abolished when the circadian molecular clock is stopped. A whole-cell biophysical model using Hodgkin-Huxley equations can recapitulate these changes in electrical activity. Based on this model and by using dynamic clamp to manipulate clock neurons directly, we can rescue the pharmacological block of Shaw and Shal, restore the firing rhythm, and thus demonstrate the critical importance of Shaw and Shal. Together, these findings point to a key role for Shaw and Shal in controlling circadian firing of clock neurons and show that changes in clock neuron currents can account for this. Moreover, with dynamic clamp we can switch the LNvs between morning-like and evening-like states of electrical activity. We conclude that changes in Shaw and Shal underlie the daily oscillation in LNv firing rate.

Neuronal overexpression of Alzheimer's disease and Down's syndrome associated DYRK1A/minibrain gene alters motor decline, neurodegeneration and synaptic plasticity in Drosophila.

Down syndrome (DS) is characterised by abnormal cognitive and motor development, and later in life by progressive Alzheimer's disease (AD)-like dementia, neuropathology, declining motor function and shorter life expectancy. It is caused by trisomy of chromosome 21 (Hsa21), but how individual Hsa21 genes contribute to various aspects of the disorder is incompletely understood. Previous work has demonstrated a role for triplication of the Hsa21 gene DYRK1A in cognitive and motor deficits, as well as in altered neurogenesis and neurofibrillary degeneration in the DS brain, but its contribution to other DS phenotypes is unclear. Here we demonstrate that overexpression of minibrain (mnb), the Drosophila ortholog of DYRK1A, in the Drosophila nervous system accelerated age-dependent decline in motor performance and shortened lifespan. Overexpression of mnb in the eye was neurotoxic and overexpression in ellipsoid body neurons in the brain caused age-dependent neurodegeneration. At the larval neuromuscular junction, an established model for mammalian central glutamatergic synapses, neuronal mnb overexpression enhanced spontaneous vesicular transmitter release. It also slowed recovery from short-term depression of evoked transmitter release induced by high-frequency nerve stimulation and increased the number of boutons in one of the two glutamatergic motor neurons innervating the muscle. These results provide further insight into the roles of DYRK1A triplication in abnormal aging and synaptic dysfunction in DS.