Gabriela Salinas

Effects of Long-Term Environmental Enrichment on Anxiety, Memory, Hippocampal Plasticity and Overall Brain Gene Expression in C57BL6 Mice

There is ample evidence that physical activity exerts positive effects on a variety of brain functions by facilitating neuroprotective processes and influencing neuroplasticity. Accordingly, numerous studies have shown that continuous exercise can successfully diminish or prevent the pathology of neurodegenerative diseases such as Alzheimer’s disease in transgenic mouse models. However, the long-term effect of physical activity on brain health of aging wild-type (WT) mice has not yet been studied in detail. Here, we show that prolonged physical and cognitive stimulation, mediated by an enriched environment (EE) paradigm for a duration of 11 months, leads to reduced anxiety and improved spatial reference memory in C57BL6 WT mice. While the number of CA1 pyramidal neurons remained unchanged between standard housed (SH) and EE mice, the number of dentate gyrus (DG) neurons, as well as the CA1 and DG volume were significantly increased in EE mice. A whole-brain deep sequencing transcriptome analysis, carried out to better understand the molecular mechanisms underlying the observed effects, revealed an up-regulation of a variety of genes upon EE, mainly associated with synaptic plasticity and transcription regulation. The present findings corroborate the impact of continuous physical activity as a potential prospective route in the prevention of age-related cognitive decline and neurodegenerative disorders.

mRNA quality control is bypassed for immediate export of stress-responsive transcripts.

Cells grow well only in a narrow range of physiological conditions. Surviving extreme conditions requires the instantaneous expression of chaperones that help to overcome stressful situations. To ensure the preferential synthesis of these heat-shock proteins, cells inhibit transcription, pre-mRNA processing and nuclear export of non-heat-shock transcripts, while stress-specific mRNAs are exclusively exported and translated. How cells manage the selective retention of regular transcripts and the simultaneous rapid export of heat-shock mRNAs is largely unknown. In Saccharomyces cerevisiae, the shuttling RNA adaptor proteins Npl3, Gbp2, Hrb1 and Nab2 are loaded co-transcriptionally onto growing pre-mRNAs. For nuclear export, they recruit the export-receptor heterodimer Mex67–Mtr2 (TAP–p15 in humans). Here we show that cellular stress induces the dissociation of Mex67 and its adaptor proteins from regular mRNAs to prevent general mRNA export. At the same time, heat-shock mRNAs are rapidly exported in association with Mex67, without the need for adapters. The immediate co-transcriptional loading of Mex67 onto heat-shock mRNAs involves Hsf1, a heat-shock transcription factor that binds to heat-shock-promoter elements in stress-responsive genes. An important difference between the export modes is that adaptor-protein-bound mRNAs undergo quality control, whereas stress-specific transcripts do not. In fact, regular mRNAs are converted into uncontrolled stress-responsive transcripts if expressed under the control of a heat-shock promoter, suggesting that whether an mRNA undergoes quality control is encrypted therein. Under normal conditions, Mex67 adaptor proteins are recruited for RNA surveillance, with only quality-controlled mRNAs allowed to associate with Mex67 and leave the nucleus. Thus, at the cost of error-free mRNA formation, heat-shock mRNAs are exported and translated without delay, allowing cells to survive extreme situations.

The SCN Clock Governs Circadian Transcription Rhythms in Murine Epididymal White Adipose Tissue.

The circadian master pacemaker in the suprachiasmatic nucleus (SCN) orchestrates peripheral clocks in various organs and synchronizes them with external time, including those in adipose tissue, which displays circadian oscillations in various metabolic and endocrine outputs. Because our knowledge about the instructive role of the SCN clock on peripheral tissue function is based mainly on SCN lesion studies, we here used an alternative strategy employing the Cre/loxP system to functionally delete the SCN clock in mice. We performed whole-genome microarray hybridizations of murine epididymal white adipose tissue (eWAT) RNA preparations to characterize the role of the SCN clock in eWAT circadian transcriptome regulation. Most of the rhythmic transcripts in control animals were not rhythmic in SCN mutants, but a significant number of transcripts were rhythmic only in mutant eWAT. Core clock genes were rhythmic in both groups, but as was reported before for other tissues, rhythms were dampened and phase advanced in mutant animals. In SCN-mutant mice, eWAT lost the rhythm of metabolic pathway–related transcripts, while transcripts gaining rhythms in SCN-mutant mice were associated with various immune functions. These data reveal a complex interaction of SCN-derived and local circadian signals in the regulation of adipose transcriptome programs.

Global analysis of asymmetric RNA enrichment in oocytes reveals low conservation between closely related Xenopus species

Subcellular localization of mRNAs contributes to the generation of cellular asymmetries and cell fate determination. A comparative global analysis is given of animally and vegetally enriched RNAs in oocytes from two closely related Xenopus species.

Effects of repeated long-term psychosocial stress and acute cannabinoid exposure on mouse corticostriatal circuitries: implications for neuropsychiatric disorders

Vulnerability to psychiatric manifestations is achieved by the influence of genetic and environment including stress and cannabis consumption. Here, we used a psychosocial stress model based on resident‐intruder confrontations to study the brain corticostriatal‐function, since deregulation of corticostriatal circuitries has been reported in many psychiatric disorders. CB1 receptors are widely expressed in the central nervous system and particularly, in both cortex and striatum brain structures.

Pre-infection transcript levels of FAM26F in peripheral blood mononuclear cells inform about overall plasma viral load in acute and post-acute phase after simian immunodeficiency virus infection

CD8+ cells from simian immunodeficiency virus (SIV)-infected long-term non-progressors and some uninfected macaques can suppress viral replication in vitro without killing the infected cells. The aim of this study was to identify factors responsible for non-cytolytic viral suppression by transcriptional profiling and to investigate their potential impact on SIV replication. Results of microarray experiments and further validation with cells from infected and uninfected macaques revealed that FAM26F RNA levels distinguished CD8+ cells of controllers and non-controllers (P=0.001). However, FAM26F was also expressed in CD4+ T-cells and B-cells. FAM26F expression increased in lymphocytes after in vitro IFN-γ treatment on average 40-fold, and ex vivo FAM26F RNA levels in peripheral blood mononuclear cells correlated with plasma IFN-γ but not with IFN-α. Baseline FAM26F expression appeared to be stable for months, albeit the individual expression levels varied up to tenfold. Investigating its role in SIV-infection revealed that FAM26F was upregulated after infection (P<0.0008), but did not directly correlate with viral load in contrast to MX1 and CXCL10. However, pre-infection levels of FAM26F correlated inversely with overall plasma viral load (AUC) during the acute and post-acute phases of infection (e.g. AUC weeks post infection 0–8; no AIDS vaccine: P<0.0001, Spearman rank correlation coefficient (rs)=−0.89, n=16; immunized with an AIDS vaccine: P=0.033, rs=−0.43; n=25). FAM26F transcript levels prior to infection can provide information about the pace and strength of the antiviral immune response during the early stage of infection. FAM26F expression represented, in our experiments, one of the earliest prognostic markers, and could supplement major histocompatibility complex (MHC)-typing to predict disease progression before SIV-infection.

Hepatic gene therapy rescues high-fat diet responses in circadian Clock mutant mice.

Objective Circadian Clock gene mutant mice show dampened 24-h feeding rhythms and an increased sensitivity to high-fat diet (HFD) feeding. Restricting HFD access to the dark phase counteracts its obesogenic effect in wild-type mice. The extent to which altered feeding rhythms are causative for the obesogenic phenotype of Clock mutant mice, however, remains unknown. Methods Metabolic parameters of wild-type (WT) and ClockΔ19 mutant mice (MT) were investigated under ad libitum and nighttime restricted HFD feeding. Liver circadian clock function was partially rescued by hydrodynamic tail vein delivery of WT-Clock DNA vectors in mutant mice and transcriptional, metabolic, endocrine and behavioral rhythms studied. Results Nighttime-restricted feeding restored food intake, but not body weight regulation in MT mice under HFD, suggesting Clock-dependent metabolic dysregulation downstream of circadian appetite control. Liver-directed Clock gene therapy partially restored liver circadian oscillator function and transcriptome regulation without affecting centrally controlled circadian behaviors. Under HFD, MT mice with partially restored liver clock function (MT-LR) showed normalized body weight gain, rescued 24-h food intake rhythms, and WT-like energy expenditure. This was associated with decreased nighttime leptin and daytime ghrelin levels, reduced hepatic lipid accumulation, and improved glucose tolerance. Transcriptome analysis revealed that hepatic Clock rescue in MT mice affected a range of metabolic pathways. Conclusion Liver Clock gene therapy improves resistance against HFD-induced metabolic impairments in mice with circadian clock disruption. Restoring or stabilizing liver clock function might be a promising target for therapeutic interventions in obesity and metabolic disorders.

Immortalization of common marmoset monkey fibroblasts by piggyBac transposition of hTERT

Following a certain type-specific number of mitotic divisions, terminally differentiated cells undergo proliferative senescence, thwarting efforts to expand different cell populations in vitro for the needs of scientific research or medical therapies. The primary cause of this phenomenon is the progressive shortening of the telomeres and the subsequent activation of cell cycle control pathways leading to a block of cell proliferation. Restoration of telomere length by transgenic expression of telomerase reverse transcriptase (TERT) usually results in bypassing of the replicative senescence and ultimately in cell immortalization. To date, there have not been any reports regarding immortalization of cells from common marmoset (Callithrix jacchus), an important non-human primate model for various human diseases, with the use of exogenous human TERT (hTERT). In this study, marmoset fibroblasts were successfully immortalized with transposon-integrated transgenic hTERT and expanded in vitro for over 500 population doublings. Calculation of population doubling levels (PDL) showed that the derived hTERT-transgenic lines had significantly higher proliferation potential than the wild-type fibroblasts, which reached only a maximum of 46 doublings. However, the immortalized cells exhibited differences in the morphology compared with the control fibroblasts and transcriptome analysis also revealed changes in the gene expression patterns. Finally, the karyotypes of all hTERT-transgenic cell lines showed various aberrations such as presence of extra Chromosome 17, isochromosome 21q, or tetraploidy. By single-cell expansion of the least affected monoclonal immortalized line, one sub-clonal line with normal karyotype was established, suggesting the possibility to derive immortal marmoset cells with normal karyotypes. The results of this study are an important step towards the development and optimization of methods for the production of immortalized cells from common marmoset monkeys.

Expression of the DNA-Binding Factor TOX Promotes the Encephalitogenic Potential of Microbe-Induced Autoreactive CD8+ T Cells

SUMMARY Infections are thought to trigger CD8+ cytotoxic T lymphocyte (CTL) responses during autoimmunity. However, the transcriptional programs governing the tissue‐destructive potential of CTLs remain poorly defined. In a model of central nervous system (CNS) inflammation, we found that infection with lymphocytic choriomeningitis virus (LCMV), but not Listeria monocytogenes (Lm), drove autoimmunity. The DNA‐binding factor TOX was induced in CTLs during LCMV infection and was essential for their encephalitogenic properties, and its expression was inhibited by interleukin‐12 during Lm infection. TOX repressed the activity of several transcription factors (including Id2, TCF‐1, and Notch) that are known to drive CTL differentiation. TOX also reduced immune checkpoint sensitivity by restraining the expression of the inhibitory checkpoint receptor CD244 on the surface of CTLs, leading to increased CTL‐mediated damage in the CNS. Our results identify TOX as a transcriptional regulator of tissue‐destructive CTLs in autoimmunity, offering a potential mechanistic link to microbial triggers. Graphical Abstract Figure. No Caption available. HighlightsLCMV‐primed but not Listeria‐primed CD8+ T cells exert encephalitogenic activityLCMV infection induces the DNA‐binding factor TOX in CNS‐infiltrating CD8+ T cellsLoss of TOX abrogates the tissue‐destructive ability of CD8+ T cellsTOX restrains terminal differentiation of effector CD8+ T cells &NA; Little is known about the transcriptional programs that drive the tissue destructive capacity of effector CD8+ T cells during autoimmunity. In an animal model of CNS inflammation, Page et al. demonstrate that expression of the DNA‐binding factor TOX promotes the encephalitogenic potential of pathogen‐primed CD8+ T cells and that TOX expression is determined by the microbial context of CTL priming.

PI3K: A master regulator of brain metastasis-promoting macrophages/microglia

Mutations and activation of the PI3K signaling pathway in breast cancer cells have been linked to brain metastases. However, here we describe that in some breast cancer brain metastases samples the protein expression of PI3K signaling components is restricted to the metastatic microenvironment. In contrast to the therapeutic effects of PI3K inhibition on the breast cancer cells, the reaction of the brain microenvironment is less understood. Therefore we aimed to quantify the PI3K pathway activity in breast cancer brain metastasis and investigate the effects of PI3K inhibition on the central nervous system (CNS) microenvironment. First, to systematically quantify the PI3K pathway activity in breast cancer brain metastases, we performed a prospective biomarker study using a reverse phase protein array (RPPA). The majority, namely 30 out of 48 (62.5%) brain metastatic tissues examined, revealed high PI3K signaling activity that was associated with a median overall survival (OS) of 9.41 months, while that of patients, whose brain metastases showed only moderate or low PI3K activity, amounted to only 1.93 and 6.71 months, respectively. Second, we identified PI3K as a master regulator of metastasis‐promoting macrophages/microglia during CNS colonization; and treatment with buparlisib (BKM120), a pan‐PI3K Class I inhibitor with a good blood‐brain‐barrier penetrance, reduced their metastasis‐promoting features. In conclusion, PI3K signaling is active in the majority of breast cancer brain metastases. Since PI3K inhibition does not only affect the metastatic cells but also re‐educates the metastasis‐promoting macrophages/microglia, PI3K inhibition may hold considerable promise in the treatment of brain metastasis and the respective microenvironment.