Retrograde Bone Morphogenetic Protein Signaling Shapes a Key Circadian Pacemaker Circuit

The neuropeptide pigment-dispersing factor (PDF) synchronizes molecular oscillations within circadian pacemakers in the Drosophila brain. It is expressed in the small ventral lateral neurons (sLNvs) and large ventral lateral neurons, the former being indispensable for maintaining behavioral rhythmicity under free-running conditions. How PDF circuits develop the specific connectivity traits that endow such global behavioral control remains unknown. Here, we show that mature sLNv circuits require PDF signaling during early development, acting through its cognate receptor PDFR at postsynaptic targets. Yet, axonal defects by PDF knockdown are presynaptic and become apparent only after metamorphosis, highlighting a delayed response to a signal released early on. Presynaptic expression of constitutively active bone morphogenetic protein (BMP) receptors prevents pdfr mutants misrouting phenotype, while sLNv-restricted downregulation of BMP signaling components phenocopied pdf(01). Thus, we have uncovered a novel mechanism that provides an early "tagging" of synaptic targets that will guide circuit refinement later in development.Fil: Gorostiza, Ezequiel Axel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Ceriani, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentin

Dorsal clock neurons claw their way out to control sleep in Drosophila

The drive to sleep is strongly influenced by time of day, with temporal information conveyed through the circadian clock. In pursuit of the neural mechanisms underlying this process, in this issue of Neuron, Sun et al. identify a novel circuit that links circadian output neurons to sleep-promoting neurons within the mushroom bodies.Fil: Duhart, José Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Ceriani, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin

A Novel Genetic Screen Identifies Modifiers of Age-Dependent Amyloid β Toxicity in the Drosophila Brain

The accumulation of amyloid β peptide (Aβ) in the brain of Alzheimer's disease (AD) patients begins many years before clinical onset. Such process has been proposed to be pathogenic through the toxicity of Aβ soluble oligomers leading to synaptic dysfunction, phospho-tau aggregation and neuronal loss. Yet, a massive accumulation of Aβ can be found in approximately 30% of aged individuals with preserved cognitive function. Therefore, within the frame of the "amyloid hypothesis", compensatory mechanisms and/or additional neurotoxic or protective factors need to be considered and investigated. Here we describe a modifier genetic screen in Drosophila designed to identify genes that modulate toxicity of Aβ42 in the CNS. The expression of Aβ42 led to its accumulation in the brain and a moderate impairment of negative geotaxis at 18 days post-eclosion (d.p.e) as compared with genetic or parental controls. These flies were mated with a collection of lines carrying chromosomal deletions and negative geotaxis was assessed at 5 and 18 d.p.e. Our screen is the first to take into account all of the following features, relevant to sporadic AD: (1) pan-neuronal expression of wild-type Aβ42; (2) a quantifiable complex behavior; (3) Aβ neurotoxicity associated with progressive accumulation of the peptide; and (4) improvement or worsening of climbing ability only evident in aged animals. One hundred and ninety-nine deficiency (Df) lines accounting for ~6300 genes were analyzed. Six lines, including the deletion of 52 Drosophila genes with human orthologs, significantly modified Aβ42 neurotoxicity in 18-day-old flies. So far, we have validated CG11796 and identified CG17249 as a strong candidate (whose human orthologs are HPD and PRCC, respectively) by using RNAi or mutant hemizygous lines. PRCC encodes proline-rich protein PRCC (ppPRCC) of unknown function associated with papillary renal cell carcinoma. HPD encodes 4-hydroxyphenylpyruvate dioxygenase (HPPD), a key enzyme in tyrosine degradation whose Df causes autosomal recessive Tyrosinemia type 3, characterized by mental retardation. Interestingly, lines with a partial Df of HPD ortholog showed increased intraneuronal accumulation of Aβ42 that coincided with geotaxis impairment. These previously undetected modifiers of Aβ42 neurotoxicity in Drosophila warrant further study to validate their possible role and significance in the pathogenesis of sporadic AD.Fil: Belfiori Carrasco, Lautaro Francisco. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Marcora, Maria Silvina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Bocai, Nadia Irina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Ceriani, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Morelli, Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Castaño, Eduardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin

Genome-Wide Expression Analysis in Drosophila Reveals Genes Controlling Circadian Behavior

In Drosophila, a number of key processes such as emergence from the pupal case, locomotor activity, feeding, olfaction, and aspects of mating behavior are under circadian regulation. Although we have a basic understanding of how the molecular oscillations take place, a clear link between gene regulation and downstream biological processes is still missing. To identify clock-controlled output genes, we have used an oligonucleotide-based high-density array that interrogates gene expression changes on a whole genome level. We found genes regulating various physiological processes to be under circadian transcriptional regulation, ranging from protein stability and degradation, signal transduction, heme metabolism, detoxification, and immunity. By comparing rhythmically expressed genes in the fly head and body, we found that the clock has adapted its output functions to the needs of each particular tissue, implying that tissue-specific regulation is superimposed on clock control of gene expression. Finally, taking full advantage of the fly as a model system, we have identified and characterized a cycling potassium channel protein as a key step in linking the transcriptional feedback loop to rhythmic locomotor behavior.Fil: Ceriani, Maria Fernanda. The Scripps Research Institute; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Hogenesch, John B.. Genomics Institute of the Novartis Research Foundation; Estados UnidosFil: Yanovsky, Marcelo Javier. The Scripps Research Institute; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Panda, Satchidananda. Genomics Institute of the Novartis Research Foundation; Estados Unidos. The Scripps Research Institute; Estados UnidosFil: Straume, Martin. University Of Virginia; Estados UnidosFil: Kay, Steve A.. Genomics Institute of the Novartis Research Foundation; Estados Unidos. The Scripps Research Institute; Estados Unido

Optical study of laser biospeckle activity in leaves of Jatropha curcas L. A noninvasive analysis of foliar endophyte colonization

Currently, the detection of endophytic fungi is determined mostly by invasive methods, including direct isolation of fungal organismsfrom plant tissue in growth media, molecular detection of endophyticfungi DNA from plant material by PCR, or evaluation under microscopy techniques.In this work we explore the potential of laser biospeckle activity (LBSA) to be usedfor the detection of endophytic colonization of leaves of a promising energy crop, Jatropha curcas L. We compared the laser biospeckle activityof endophyte infected and uninfected J. curcas leaves. The differences between blade and veins (including midrib) of the studied leaves was validated and growth parameters of the studied plants were also analyzed using the normalized weigthed generalized differencescoefficient (nWGD). The obtained results showeda relationship between the endophytic burden of leaves and the LBS, suggesting that LSBA is a useful tools to detect endophytic colonization in situ.Also, the increasedwater movements inside leaves promoted by endophytic colonizationcould be explainby the obtained data

High-frequency neuronal bursting is essential for circadian and sleep behaviors in drosophila

Circadian rhythms have been extensively studied in Drosophila; however, still little is known about how the electrical properties of clock neurons are specified. We have performed a behavioral genetic screen through the downregulation of candidate ion channels in the lateral ventral neurons (LNvs) and show that the hyperpolarization-activated cation current Ih is important for the behaviors that the LNvs influence: temporal organization of locomotor activity, analyzed in males, and sleep, analyzed in females. Using whole-cell patch clamp electrophysiology we demonstrate that small LNvs (sLNvs) are bursting neurons, and that Ih is necessary to achieve the high-frequency bursting firing pattern characteristic of both types of LNvs in females. Since firing in bursts has been associated to neuropeptide release, we hypothesized that Ih would be important for LNvs communication. Indeed, herein we demonstrate that Ih is fundamental for the recruitment of pigment dispersing factor (PDF) filled dense core vesicles (DCVs) to the terminals at the dorsal protocerebrum and for their timed release, and hence for the temporal coordination of circadian behaviors.Fil: Fernández, Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigación en Biomedicina de Buenos Aires - Instituto Partner de la Sociedad Max Planck; ArgentinaFil: Frenkel, Lia. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Biociencias, Biotecnología y Biología Traslacional; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Colque, Carina Celeste. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Ricciuti, Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigación en Biomedicina de Buenos Aires - Instituto Partner de la Sociedad Max Planck; ArgentinaFil: Hahm, Bryan. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigación en Biomedicina de Buenos Aires - Instituto Partner de la Sociedad Max Planck; ArgentinaFil: Cerredo, Karina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigación en Biomedicina de Buenos Aires - Instituto Partner de la Sociedad Max Planck; ArgentinaFil: Muraro, Nara Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigación en Biomedicina de Buenos Aires - Instituto Partner de la Sociedad Max Planck; ArgentinaFil: Ceriani, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin

Amyloid peptides ABri and ADan show differential neurotoxicity in transgenic Drosophila models of familial British and Danish dementia

BACKGROUND: Familial British and Familial Danish dementias (FBD and FDD, respectively) are associated with mutations in the BRI2 gene. Processing of the mutated BRI2 protein leads to the accumulation in the brain of the 34-mer amyloid Bri (ABri) and amyloid Dan (ADan) peptides, accompanied by neurofibrillary tangles. Recently, transgenic mice successfully reproduced different aspects of FDD, while modeling of FBD in vivo has been more difficult. In this work we have modeled FBD and FDD in Drosophila and tested the hypothesis that ABri and ADan are differentially neurotoxic. RESULTS: By using site-directed insertion, we generated transgenic lines carrying ABri, ADan, Bri2-23 (the normal product of wild-type BRI2 processing) and amyloid-β (Aβ) 1-42 as a well-characterized neurotoxic peptide, alone or with a His-tag. Therefore, we avoided random insertion effects and were able to compare levels of accumulation accurately. Peptides were expressed with the GAL4-Upstream Activating Sequence (UAS) system using specific drivers. Despite low levels of expression, toxicity in the eye was characterized by mild disorganization of ommatidia and amyloid peptides accumulation. The highest toxicity was seen for ADan, followed by Aβ42 and ABri. Pan-neuronal expression in the CNS revealed an age-dependent toxicity of amyloid peptides as determined by the ability of flies to climb in a geotaxis paradigm when compared to Bri2-23. This effect was stronger for ADan, detected at 7 days post-eclosion, and followed by ABri and Aβ42, whose toxicity became evident after 15 and 21 days, respectively. Histological analysis showed mild vacuolization and thioflavine-S-negative deposits of amyloid peptides. In contrast, the over-expression of amyloid peptides in the specific subset of lateral neurons that control circadian locomotor activity showed no toxicity. CONCLUSIONS: Our results support the differential neurotoxicity of ADan and ABri in the Drosophila eye and CNS at low expression levels. Such differences may be partially attributed to rates of aggregation and accumulation. In the CNS, both peptides appear to be more neurotoxic than wild-type Aβ42. These Drosophila models will allow a systematic and unambiguous comparison of differences and similarities in the mechanisms of toxicity of diverse amyloid peptides associated with dementia.Fil: Marcora, Maria Silvina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Fernandez Gamba, Agata Claudia. Fundación Instituto Leloir; ArgentinaFil: Avendaño, Luz A. Fundación Instituto Leloir; ArgentinaFil: Rotondaro, Cecilia. Fundación Instituto Leloir; ArgentinaFil: Podhajcer, Osvaldo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Vidal, Rubén. Indiana University School of Medicine. Indiana Alzheimer Disease Center. Department of Pathology and Laboratory Medicine; Estados UnidosFil: Morelli, Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Ceriani, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Castaño, Eduardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentin

Guidelines for Genome-Scale Analysis of Biological Rhythms

Genome biology approaches have made enormous contributions to our understanding of biological rhythms, particularly in identifying outputs of the clock, including RNAs, proteins, and metabolites, whose abundance oscillates throughout the day. These methods hold significant promise for future discovery, particularly when combined with computational modeling. However, genome-scale experiments are costly and laborious, yielding “big data” that are conceptually and statistically difficult to analyze. There is no obvious consensus regarding design or analysis. Here we discuss the relevant technical considerations to generate reproducible, statistically sound, and broadly useful genome-scale data. Rather than suggest a set of rigid rules, we aim to codify principles by which investigators, reviewers, and readers of the primary literature can evaluate the suitability of different experimental designs for measuring different aspects of biological rhythms. We introduce CircaInSilico, a web-based application for generating synthetic genome biology data to benchmark statistical methods for studying biological rhythms. Finally, we discuss several unmet analytical needs, including applications to clinical medicine, and suggest productive avenues to address them

Impact of COVID-19 on cardiovascular testing in the United States versus the rest of the world

Objectives: This study sought to quantify and compare the decline in volumes of cardiovascular procedures between the United States and non-US institutions during the early phase of the coronavirus disease-2019 (COVID-19) pandemic. Background: The COVID-19 pandemic has disrupted the care of many non-COVID-19 illnesses. Reductions in diagnostic cardiovascular testing around the world have led to concerns over the implications of reduced testing for cardiovascular disease (CVD) morbidity and mortality. Methods: Data were submitted to the INCAPS-COVID (International Atomic Energy Agency Non-Invasive Cardiology Protocols Study of COVID-19), a multinational registry comprising 909 institutions in 108 countries (including 155 facilities in 40 U.S. states), assessing the impact of the COVID-19 pandemic on volumes of diagnostic cardiovascular procedures. Data were obtained for April 2020 and compared with volumes of baseline procedures from March 2019. We compared laboratory characteristics, practices, and procedure volumes between U.S. and non-U.S. facilities and between U.S. geographic regions and identified factors associated with volume reduction in the United States. Results: Reductions in the volumes of procedures in the United States were similar to those in non-U.S. facilities (68% vs. 63%, respectively; p = 0.237), although U.S. facilities reported greater reductions in invasive coronary angiography (69% vs. 53%, respectively; p < 0.001). Significantly more U.S. facilities reported increased use of telehealth and patient screening measures than non-U.S. facilities, such as temperature checks, symptom screenings, and COVID-19 testing. Reductions in volumes of procedures differed between U.S. regions, with larger declines observed in the Northeast (76%) and Midwest (74%) than in the South (62%) and West (44%). Prevalence of COVID-19, staff redeployments, outpatient centers, and urban centers were associated with greater reductions in volume in U.S. facilities in a multivariable analysis. Conclusions: We observed marked reductions in U.S. cardiovascular testing in the early phase of the pandemic and significant variability between U.S. regions. The association between reductions of volumes and COVID-19 prevalence in the United States highlighted the need for proactive efforts to maintain access to cardiovascular testing in areas most affected by outbreaks of COVID-19 infection

Guidelines for Genome-Scale Analysis of Biological Rhythms

Genome biology approaches have made enormous contributions to our understanding of biological rhythms, particularly in identifying outputs of the clock, including RNAs, proteins, and metabolites, whose abundance oscillates throughout the day. These methods hold significant promise for future discovery, particularly when combined with computational modeling. However, genome-scale experiments are costly and laborious, yielding ‘big data’ that is conceptually and statistically difficult to analyze. There is no obvious consensus regarding design or analysis. Here we discuss the relevant technical considerations to generate reproducible, statistically sound, and broadly useful genome scale data. Rather than suggest a set of rigid rules, we aim to codify principles by which investigators, reviewers, and readers of the primary literature can evaluate the suitability of different experimental designs for measuring different aspects of biological rhythms. We introduce CircaInSilico, a web-based application for generating synthetic genome biology data to benchmark statistical methods for studying biological rhythms. Finally, we discuss several unmet analytical needs, including applications to clinical medicine, and suggest productive avenues to address them