AAV delivery of shRNA against IRS1 in GABAergic neurons in rat hippocampus impairs spatial memory in females and male rats

This is a pre-print of an article published in Brain Structure and Function. The final authenticated version is available online at: https://doi.org/10.1007/s00429-020-02155-xBrain insulin resistance is a major factor leading to impaired cognitive function and it is considered as the onset of Alzheimer´s disease. Insulin resistance is intimately linked to inflammatory conditions, many studies have revealed how pro-inflammatory cytokines lead to insulin resistance, by inhibiting IRS1 function. Thus, the dysfunction of insulin signaling is concomitant with inflammatory biomarkers. However, the specific effect of IRS1 impaired function in otherwise healthy brain has not been dissected out. So, we decided in our study, to study the specific role of IRS1 in the hippocampus, in the absence of comorbidities. To that end, shRNA against rat and human IRS1 was designed and tested in cultured HEK cells to evaluate mRNA levels and specificity. The best candidate sequence was encapsulated in an AAV vector (strain DJ8) under the control of the cytomegalovirus promoter and together with the green fluorescent protein gene as a reporter. AAV-CMV-shIRS1-EGFP and control AAV-CMV-EGFP were inoculated into the dorsal hippocampus of female and male Wistar rats. One month later, animals undertook a battery of behavioral paradigms evaluating spatial and social memory and anxiety. Our results suggest that females displayed increased susceptibility to AAV-shIRS1 in the novel recognition object paradigm; whereas both females and males show impaired performance in the T maze when infected with AAV-shIRS1 compared to control. Anxiety parameters were not affected by AAV-shIRS1 infection. We observed specific fluorescence within the hilum of the dentate gyrus, in immuno-characterized parvalbumin and somatostatin neurons. AAV DJ8 did not enter astrocytes. Intense green fibers were found in the fornix, mammillary bodies, and in the medial septum indicating that hippocampal efferent had been efficiently targeted by the AAV DJ8 infection. We observed that AAV-shIRS1 reduced significantly synaptophysin labeling in hippocampal-septal projections compared to controls. These results support that, small alterations in the insulin/IGF1 pathway in specific hippocampal circuitries can underlie alterations in synaptic plasticity and affect behavior, in the absence of inflammatory condition

Proyecto de asentamiento ex-novo barrio de Intaca (Mozambique)

El objetivo general del presente proyecto es contribuir al uso racional del suelo con fines urbanos, disminuyendo los niveles de informalidad en la vivienda y contando con el esfuerzo propio y la ayuda mutua como bases para el desarrollo del proyecto, posibilitando un plan vital integrado a través de la satisfacción de las necesidades de asentamiento, residencia y producción. El objetivo específico consiste en desarrollar un proyecto de asentamiento ex-novo para 670 familias en un área de 72 ha ubicada en el barrio de Intaca, a 23 km del centro de la ciudad de Matola. El proyecto incluye la comprensión del microterritorio, el análisis de los problemas existentes y el planteamiento de una solución integrada que los resuelva, pasando por las fases de análisis de la elección del sitio, parcelación, urbanización, y edificación. Es fundamental contar tanto con la fuerza de los pobladores (última fase principalmente), como con la colaboración de instituciones locales desde el comienzo

Abscisic Acid Supplementation Rescues High Fat Diet-Induced Alterations in Hippocampal Inflammation and IRSs Expression

Accumulated evidence indicates that neuroinflammation induces insulin resistance in the brain. Moreover, both processes are intimately linked to neurodegenerative disorders, including Alzheimer’s disease. Potential mechanisms underlying insulin resistance include serine phosphorylation of the insulin receptor substrate (IRS) or insulin receptor (IR) misallocation. However, only a few studies have focused on IRS expression in the brain and its modulation in neuroinflammatory processes. This study used the high-fat diet (HFD) model of neuroinflammation to study the alterations of IR, an insulin-like growth factor receptor (IGF1R) and IRS expressions in the hippocampus. We observed that HFD effectively reduced mRNA and protein IRS2 expression. In contrast, a HFD induced the upregulation of the IRS1 mRNA levels, but did not alter an IR and IGF1R expression. As expected, we observed that a HFD increased hippocampal tumor necrosis factor alpha (TNFα) and amyloid precursor protein (APP) levels while reducing brain-derived neurotrophic factor (BDNF) expression and neurogenesis. Interestingly, we found that TNFα correlated positively with IRS1 and negatively with IRS2, whereas APP levels correlated positively only with IRS1 but not IRS2. These results indicate that IRS1 and IRS2 hippocampal expression can be affected differently by HFD-induced neuroinflammation. In addition, we aimed to establish whether abscisic acid (ABA) can rescue hippocampal IRS1 and IRS2 expression, as we had previously shown that ABA supplementation prevents memory impairments and improves neuroinflammation induced by a HFD. In this study, ABA restored HFD-induced hippocampal alterations, including IRS1 and IRS2 expression, TNFα, APP, and BDNF levels and neurogenesis. In conclusion, this study highlights different regulations of hippocampal IRS1 and IRS2 expression using a HFD, indicating the important differences of these scaffolding proteins, and strongly supports ABA therapeutic effects

ENO regulates tomato fruit size through the floral meristem development network

A dramatic evolution of fruit size has accompanied the domestication and improvement of fruit-bearing crop species. In tomato (Solanum lycopersicum), naturally occurring cis-regulatory mutations in the genes of the CLAVATA-WUSCHEL signaling pathway have led to a significant increase in fruit size generating enlarged meristems that lead to flowers with extra organs and bigger fruits. In this work, by combining mapping-by-sequencing and CRISPR/Cas9 genome editing methods, we isolated EXCESSIVE NUMBER OF FLORAL ORGANS (ENO), an AP2/ERF transcription factor which regulates floral meristem activity. Thus, the ENO gene mutation gives rise to plants that yield larger multilocular fruits due to an increased size of the floral meristem. Genetic analyses indicate that eno exhibits synergistic effects with mutations at the LOCULE NUMBER (encoding SlWUS) and FASCIATED (encoding SlCLV3) loci, two central players in the evolution of fruit size in the domestication of cultivated tomatoes. Our findings reveal that an eno mutation causes a substantial expansion of SlWUS expression domains in a flower-specific manner. In vitro binding results show that ENO is able to interact with the GGC-box cis-regulatory element within the SlWUS promoter region, suggesting that ENO directly regulates SlWUS expression domains to maintain floral stem-cell homeostasis. Furthermore, the study of natural allelic variation of the ENO locus proved that a cis-regulatory mutation in the promoter of ENO had been targeted by positive selection during the domestication process, setting up the background for significant increases in fruit locule number and fruit size in modern tomatoes

Giant tortoise genomes provide insights into longevity and age-related disease

© 2018, The Author(s), under exclusive licence to Springer Nature Limited. Giant tortoises are among the longest-lived vertebrate animals and, as such, provide an excellent model to study traits like longevity and age-related diseases. However, genomic and molecular evolutionary information on giant tortoises is scarce. Here, we describe a global analysis of the genomes of Lonesome George—the iconic last member of Chelonoidis abingdonii—and the Aldabra giant tortoise (Aldabrachelys gigantea). Comparison of these genomes with those of related species, using both unsupervised and supervised analyses, led us to detect lineage-specific variants affecting DNA repair genes, inflammatory mediators and genes related to cancer development. Our study also hints at specific evolutionary strategies linked to increased lifespan, and expands our understanding of the genomic determinants of ageing. These new genome sequences also provide important resources to help the efforts for restoration of giant tortoise populations

Host factor PLAC8 is required for pancreas infection by SARS-CoV-2

Although mounting evidence demonstrated that pancreas is infected by SARS-CoV-2 the severity and pathophysiology of pancreatic COVID-19 disease are still unclear. Here we investigated the consequences of SARS-CoV-2 infection of the pancreas and the role of Placenta-associated protein-8 (PLAC8). Our data showed pancreatic damage in patients who died from COVID-19. Notably, circulating pancreatic enzymes stratified patients according to COVID-19 severity and outcome. PLAC8 expression was associated with SARS-CoV-2 infection in postmortem analysis of COVID-19 patients and functional assays demonstrated the requirement of PLAC8 in SARS-CoV-2 pancreatic infection. Full SARS-CoV-2 infectious virus revealed a requirement of PLAC8 for efficient viral infection of pancreatic cell lines. Finally, we observed colocalization of PLAC8 and SARS-CoV-2 in the pancreas of deceased patients. In conclusion, our data confirm the human pancreas as a SARS-CoV-2 target and demonstrate the requirement of PLAC8 for SARS-CoV-2 pancreatic infection thereby opening new target opportunities for COVID-19-associated pancreatic pathogenesis.N

Genetic interactions of the unfinished flower development (ufd) mutant support a significant role of the tomato UFD gene in regulating floral organogenesis

Tomato (Solanum lycopersicum L.) is a major vegetable crop that also constitutes a model species for the study of plant developmental processes. To gain insight into the control of flowering and floral development, a novel tomato mutant, unfinished flower development (ufd), whose inflorescence and flowers were unable to complete their normal development was characterized using double mutant and gene expression analyses. Genetic interactions of ufd with mutations affecting inflorescence fate (uniflora, jointless and single flower truss) were additive and resulted in double mutants displaying the inflorescence structure of the non-ufd parental mutant and the flower phenotype of the ufd mutant. In addition, ufd mutation promotes an earlier inflorescence meristem termination. Taken together, both results indicated that UFD is not involved in the maintenance of inflorescence meristem identity, although it could participate in the regulatory system that modulates the rate of meristem maturation. Regarding the floral meristem identity, the falsiflora mutation was epistatic to the ufd mutation even though FALSIFLORA was upregulated in ufd inflorescences. In terms of floral organ identity, the ufd mutation was epistatic to macrocalyx, and MACROCALYX expression was differently regulated depending on the inflorescence developmental stage. These results suggest that the UFD gene may play a pivotal role between the genes required for flowering initiation and inflorescence development (such as UNIFLORA, FALSIFLORA, JOINTLESS and SINGLE FLOWER TRUSS) and those required for further floral organ development such as the floral organ identity genes

IRS1 expression in hippocampus is age-dependent and is required for mature spine maintenance and neuritogenesis

Insulin and insulin-like growth factor type I (IGF-1) play prominent roles in brain activity throughout the lifespan. Insulin/IGF1 signaling starts with the activation of the intracellular insulin receptor substrates (IRS). In this work, we performed a comparative study of IRS1 and IRS2, together with the IGF1 (IGF1R) and insulin (IR) receptor expression in the hippocampus and prefrontal cortex during development. We found that IRS1 and IRS2 expression is prominent during development and declines in the aged hippocampus, contrary to IR, which increases in adulthood and aging. In contrast, IGF1R expression is unaffected by age. Expression patterns are similar in the prefrontal cortex. Neurite development occurs postnatally in the rodent hippocampus and cortex, and it declines in the mature and aged brain and is influenced by trophic factors. In our previous work, we demonstrated that knockdown of IRS1 by shRNA impairs learning and reduces synaptic plasticity in a rat model, as measured by synaptophysin puncta in axons. In this study, we report that shIRS1 alters spine maturation in adult hilar hippocampal neurons. Lastly, to understand the role of IRS1 in neuronal neurite tree, we transfect shIRS1 into primary neuronal cultures and observed that shIRS1 reduced neurite branching and neurite length. Our results demonstrate that IRS1/2 and insulin/IGF1 receptors display different age-dependent expression profiles and that IRS1 is required for spine maturation, demonstrating a novel role for IRS1 in synaptic plasticity