scholarly journals mTOR Driven Gene Transcription Is Required for Cholesterol Production in Neurons of the Developing Cerebral Cortex

2021 ◽  
Vol 22 (11) ◽  
pp. 6034
Author(s):  
Martin Schüle ◽  
Tamer Butto ◽  
Sri Dewi ◽  
Laura Schlichtholz ◽  
Susanne Strand ◽  
...  
Keyword(s):  
Cerebral Cortex ◽  
Neurodevelopmental Disorders ◽  
Cholesterol Biosynthesis ◽  
Single Gene ◽  
Autism Spectrum ◽  
Altered Expression ◽  
Mtorc1 Signaling ◽  
Mtor Activity

Dysregulated mammalian target of rapamycin (mTOR) activity is associated with various neurodevelopmental disorders ranging from idiopathic autism spectrum disorders (ASD) to syndromes caused by single gene defects. This suggests that maintaining mTOR activity levels in a physiological range is essential for brain development and functioning. Upon activation, mTOR regulates a variety of cellular processes such as cell growth, autophagy, and metabolism. On a molecular level, however, the consequences of mTOR activation in the brain are not well understood. Low levels of cholesterol are associated with a wide variety of neurodevelopmental disorders. We here describe numerous genes of the sterol/cholesterol biosynthesis pathway to be transcriptionally regulated by mTOR complex 1 (mTORC1) signaling in vitro in primary neurons and in vivo in the developing cerebral cortex of the mouse. We find that these genes are shared targets of the transcription factors SREBP, SP1, and NF-Y. Prenatal as well as postnatal mTORC1 inhibition downregulated expression of these genes which directly translated into reduced cholesterol levels, pointing towards a substantial metabolic function of the mTORC1 signaling cascade. Altogether, our results indicate that mTORC1 is an essential transcriptional regulator of the expression of sterol/cholesterol biosynthesis genes in the developing brain. Altered expression of these genes may be an important factor contributing to the pathogenesis of neurodevelopmental disorders associated with dysregulated mTOR signaling.

scholarly journals mTOR inhibition in primary neurons and the developing brain represses transcription of cholesterol biosynthesis genes and alters cholesterol levels

2020 ◽  
Author(s):  
Martin Schüle ◽  
Tamer Butto ◽  
Sri Dewi ◽  
Susanne Strand ◽  
Susanne Gerber ◽  
...  
Keyword(s):  
Neurodevelopmental Disorders ◽  
Cholesterol Biosynthesis ◽  
Autism Spectrum ◽  
Developing Brain ◽  
Primary Neurons ◽  
Mtor Inhibition ◽  
Altered Expression ◽  
Cholesterol Levels ◽  
The Brain ◽  
Mtor Activity

AbstractDysregulated mammalian target of rapamycin (mTOR) activity is associated with various neurodevelopmental disorders ranging from idiopathic autism spectrum disorders to monogenic syndromes as for example Tuberous sclerosis complex. Thus, maintaining mTOR activity levels in a physiological range is essential for brain development and functioning. Upon activation, mTOR regulates a variety of cellular processes such as cell growth, autophagy and metabolism. On a molecular level, however, the consequences of mTOR activation are not well understood, especially in the brain. Thus, while it was shown that in cells outside the central nervous system mTORC1 activity is necessary for activating gene transcription of different metabolic pathways this mechanism is ill defined in the brain.By combining mTORC1 inhibition with RNA-sequencing we identified numerous genes of the sterol/cholesterol biosynthesis pathway to be downstream targets of mTORC1 in vitro in primary neurons and in vivo in the developing cerebral cortex of the mouse. Of note, reduced expression of these genes upon mTORC1 inhibition translated into reduced cholesterol levels. We further show that while mTORC1 does not regulate chromatin accessibility or RNA stability of these genes it drives transcription of their DNA. Using a bioinformatics approach, we identified binding sites for the transcription factors SREBP, SP1 and NF-Y to be enriched in the promoters of mTORC1 target genes and confirmed binding of NF-YA by ChIP-qPCR. Altogether, our results indicate that mTORC1 is an important regulator of the expression of sterol/cholesterol biosynthesis genes in the developing brain. Altered expression of these genes may be an important contributing factor in the pathogenesis of neurodevelopmental disorders associated with dysregulated mTOR signaling.

scholarly journals Neuroanatomy and behavior in mice with a haploinsufficiency of AT-rich interactive domain 1B (ARID1B) throughout development

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
J. Ellegood ◽  
S. P. Petkova ◽  
A. Kinman ◽  
L. R. Qiu ◽  
A. Adhikari ◽  
...  
Keyword(s):  
Corpus Callosum ◽  
Ex Vivo ◽  
Chromatin Modification ◽  
Autism Spectrum ◽  
Repetitive Behaviors ◽  
Social Deficits ◽  
Altered Expression ◽  
Developmental Growth ◽  
And Behavior

Abstract Background One of the causal mechanisms underlying neurodevelopmental disorders (NDDs) is chromatin modification and the genes that regulate chromatin. AT-rich interactive domain 1B (ARID1B), a chromatin modifier, has been linked to autism spectrum disorder and to affect rare and inherited genetic variation in a broad set of NDDs. Methods A novel preclinical mouse model of Arid1b deficiency was created and validated to characterize and define neuroanatomical, behavioral and transcriptional phenotypes. Neuroanatomy was assessed ex vivo in adult animals and in vivo longitudinally from birth to adulthood. Behavioral testing was also performed throughout development and tested all aspects of motor, learning, sociability, repetitive behaviors, seizure susceptibility, and general milestones delays. Results We validated decreased Arid1b mRNA and protein in Arid1b+/− mice, with signatures of increased axonal and synaptic gene expression, decreased transcriptional regulator and RNA processing expression in adult Arid1b+/− cerebellum. During neonatal development, Arid1b+/− mice exhibited robust impairments in ultrasonic vocalizations (USVs) and metrics of developmental growth. In addition, a striking sex effect was observed neuroanatomically throughout development. Behaviorally, as adults, Arid1b+/− mice showed low motor skills in open field exploration and normal three-chambered approach. Arid1b+/− mice had learning and memory deficits in novel object recognition but not in visual discrimination and reversal touchscreen tasks. Social interactions in the male–female social dyad with USVs revealed social deficits on some but not all parameters. No repetitive behaviors were observed. Brains of adult Arid1b+/− mice had a smaller cerebellum and a larger hippocampus and corpus callosum. The corpus callosum increase seen here contrasts previous reports which highlight losses in corpus callosum volume in mice and humans. Limitations The behavior and neuroimaging analyses were done on separate cohorts of mice, which did not allow a direct correlation between the imaging and behavioral findings, and the transcriptomic analysis was exploratory, with no validation of altered expression beyond Arid1b. Conclusions This study represents a full validation and investigation of a novel model of Arid1b+/− haploinsufficiency throughout development and highlights the importance of examining both sexes throughout development in NDDs.

scholarly journals 1455. Potential Mechanisms of Cefiderocol MIC Increase in Enterobacterales in In Vitro Resistance Acquisition Studies

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S730-S730
Author(s):  
Yoshinori Yamano ◽  
Rio Nakamura ◽  
Miki Takemura ◽  
Roger Echols
Keyword(s):  
Iron Transport ◽  
Resistance Mechanisms ◽  
Altered Expression ◽  
Carbapenem Resistant ◽  
Mueller Hinton ◽  
Two Component ◽  
Mueller Hinton Agar ◽  
Related Proteins

Abstract Background Cefiderocol (CFDC) is a novel siderophore, iron-chelating cephalosporin, which is transported into bacteria via iron transporters. CFDC has potent in vitro and in vivo activity against all aerobic Gram-negative bacteria, including carbapenem-resistant strains. To date, clinical isolates with cefiderocol MIC >4 µg/mL have been found infrequently, in which the presence of a few β-lactamases or altered iron transport was found. We investigated potential new mechanisms causing CFDC MIC increases in non-clinical studies. Methods The mutation positions were determined by whole genome sequencing using four K. pneumoniae mutants including two KPC producers and one NDM producer that had shown CFDC MIC increases in previous in vitro resistance-acquisition studies. The mutant strains were obtained at the frequency of 10-7 to < 10-8 by spreading bacteria on standard Mueller‒Hinton agar medium containing CFDC at concentrations of 10× MIC, with or without apo-transferrin (20 μg/mL). CFDC MIC was determined by broth microdilution using iron-depleted cation-adjusted Mueller-Hinton broth based on Clinical and Laboratory Standards Institute guidelines. The emergence of MIC increase mutants was also assessed by in vitro chemostat models under humanized plasma pharmacokinetic exposures of CFDC. Results The possible resistance mechanisms were investigated. Mutation of baeS or envZ, sensors of two-component regulation systems, were found in three or two mutants among the tested four isolates, respectively, and caused the MIC to increase by 4–32-fold. The altered expression level of specific genes by the baeS or envZ mutation could affect CFDC susceptibility, but the specific genes have not been identified. In addition, the mutation of exbD, an accessory protein related to iron transport, was identified in one case and caused the MIC to increase by >8-fold. In vitro chemostat studies using two isolates (one NDM producer and one KPC producer) showed no resistance acquisition during 24-hour exposure. Table. Overview of mutation emergence in five isolates of K. pneumoniae Conclusion The mutation of two-component regulation systems (BaeSR and OmpR/EnvZ) and iron transport-related proteins were shown to be possible mechanisms causing CFDC MIC increases, but these mutants did not appear under human exposures. Disclosures Yoshinori Yamano, PhD, Shionogi & Co., Ltd. (Employee) Rio Nakamura, BSc, Shionogi & Co., Ltd. (Employee) Miki Takemura, MSc, Shionogi & Co., Ltd. (Employee) Roger Echols, MD, Shionogi Inc. (Consultant)

scholarly journals The BET Inhibitor JQ1 Augments the Antitumor Efficacy of Gemcitabine in Preclinical Models of Pancreatic Cancer

Cancers ◽  
2021 ◽  
Vol 13 (14) ◽  
pp. 3470
Author(s):  
Aubrey L. Miller ◽  
Patrick L. Garcia ◽  
Samuel C. Fehling ◽  
Tracy L. Gamblin ◽  
Rebecca B. Vance ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Dna Damage ◽  
Cholesterol Biosynthesis ◽  
Antitumor Efficacy ◽  
Rna Seq ◽  
Bet Inhibitor ◽  
Bet Inhibitors ◽  
Xenograft Models

Gemcitabine is used to treat pancreatic cancer (PC), but is not curative. We sought to determine whether gemcitabine + a BET bromodomain inhibitor was superior to gemcitabine, and identify proteins that may contribute to the efficacy of this combination. This study was based on observations that cell cycle dysregulation and DNA damage augment the efficacy of gemcitabine. BET inhibitors arrest cells in G1 and allow increases in DNA damage, likely due to inhibition of expression of DNA repair proteins Ku80 and RAD51. BET inhibitors (JQ1 or I-BET762) + gemcitabine were synergistic in vitro, in Panc1, MiaPaCa2 and Su86 PC cell lines. JQ1 + gemcitabine was more effective in vivo than either drug alone in patient-derived xenograft models (P < 0.01). Increases in the apoptosis marker cleaved caspase 3 and DNA damage marker γH2AX paralleled antitumor efficacy. Notably, RNA-seq data showed that JQ1 + gemcitabine selectively inhibited HMGCS2 and APOC1 ~6-fold, compared to controls. These proteins contribute to cholesterol biosynthesis and lipid metabolism, and their overexpression supports tumor cell proliferation. IPA data indicated that JQ1 + gemcitabine selectively inhibited the LXR/RXR activation pathway, suggesting the hypothesis that this inhibition may contribute to the observed in vivo efficacy of JQ1 + gemcitabine.

scholarly journals A study of deregulated MMR pathways and anticancer potential of curcuma derivatives using computational approach

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Priyanjali Bhattacharya ◽  
Trupti N. Patel
Keyword(s):  
Mismatch Repair ◽  
Protein Function ◽  
Curcuma Longa ◽  
Signaling Cascades ◽  
Altered Expression ◽  
Anticancer Properties ◽  
Medicinal Properties ◽  
Multiple Signaling

AbstractPlant derived products have steadily gained momentum in treatment of cancer over the past decades. Curcuma and its derivatives, in particular, have diverse medicinal properties including anticancer potential with proven safety as supported by numerous in vivo and in vitro studies. A defective Mis-Match Repair (MMR) is implicated in solid tumors but its role in haematologic malignancies is not keenly studied and the current literature suggests that it is limited. Nonetheless, there are multiple pathways interjecting the mismatch repair proteins in haematologic cancers that may have a direct or indirect implication in progression of the disease. Here, through computational analysis, we target proteins that are involved in rewiring of multiple signaling cascades via altered expression in cancer using various curcuma derivatives (Curcuma longa L. and Curcuma caesia Roxb.) which in turn, profoundly controls MMR protein function. These biomolecules were screened to identify their efficacy on selected targets (in blood-related cancers); aberrations of which adversely impacted mismatch repair machinery. The study revealed that of the 536 compounds screened, six of them may have the potential to regulate the expression of identified targets and thus revive the MMR function preventing genomic instability. These results reveal that there may be potential plant derived biomolecules that may have anticancer properties against the tumors driven by deregulated MMR-pathways.

scholarly journals Biological Relevance of Colony Morphology and Phenotypic Switching by Burkholderia pseudomallei

2006 ◽  
Vol 189 (3) ◽  
pp. 807-817 ◽  
Author(s):  
Narisara Chantratita ◽  
Vanaporn Wuthiekanun ◽  
Khaemaporn Boonbumrung ◽  
Rachaneeporn Tiyawisutsri ◽  
Mongkol Vesaratchavest ◽  
...  
Keyword(s):  
Burkholderia Pseudomallei ◽  
Vaccine Development ◽  
Colony Morphology ◽  
Phenotypic Switching ◽  
Type I ◽  
Type Ii ◽  
Altered Expression ◽  
Replication Fitness

ABSTRACT Melioidosis is a notoriously protracted illness and is difficult to cure. We hypothesize that the causative organism, Burkholderia pseudomallei, undergoes a process of adaptation involving altered expression of surface determinants which facilitates persistence in vivo and that this is reflected by changes in colony morphology. A colony morphotyping scheme and typing algorithm were developed using clinical B. pseudomallei isolates. Morphotypes were divided into seven types (denoted I to VII). Type I gave rise to other morphotypes (most commonly type II or III) by a process of switching in response to environmental stress, including starvation, iron limitation, and growth at 42°C. Switching was associated with complex shifts in phenotype, one of which (type I to type II) was associated with a marked increase in production of factors putatively associated with in vivo concealment. Isogenic types II and III, derived from type I, were examined using several experimental models. Switching between isogenic morphotypes occurred in a mouse model, where type II appeared to become adapted for persistence in a low-virulence state. Isogenic type II demonstrated a significant increase in intracellular replication fitness compared with parental type I after uptake by epithelial cells in vitro. Isogenic type III demonstrated a higher replication fitness following uptake by macrophages in vitro, which was associated with a switch to type II. Mixed B. pseudomallei morphologies were common in individual clinical specimens and were significantly more frequent in samples of blood, pus, and respiratory secretions than in urine and surface swabs. These findings have major implications for therapeutics and vaccine development.

scholarly journals Study of Staphylococcus aureusPathogenic Genes by Transfer and Expression in the Less Virulent Organism Streptococcus gordonii

2001 ◽  
Vol 69 (2) ◽  
pp. 657-664 ◽  
Author(s):  
P. Stutzmann Meier ◽  
J. M. Entenza ◽  
P. Vaudaux ◽  
P. Francioli ◽  
M. P. Glauser ◽  
...  
Keyword(s):  
Single Gene ◽  
Individual Factors ◽  
Gene Products ◽  
Streptococcus Gordonii ◽  
Pathogenic Role ◽  
Gain Of Function ◽  
Function Strategy

ABSTRACT Because Staphylococcus aureus strains contain multiple virulence factors, studying their pathogenic role by single-gene inactivation generated equivocal results. To circumvent this problem, we have expressed specific S. aureus genes in the less virulent organism Streptococcus gordonii and tested the recombinants for a gain of function both in vitro and in vivo. Clumping factor A (ClfA) and coagulase were investigated. Both gene products were expressed functionally and with similar kinetics during growth by streptococci and staphylococci. ClfA-positive S. gordoniiwas more adherent to platelet-fibrin clots mimicking cardiac vegetations in vitro and more infective in rats with experimental endocarditis (P < 0.05). Moreover, deletingclfA from clfA-positive streptococcal transformants restored both the low in vitro adherence and the low in vivo infectivity of the parent. Coagulase-positive transformants, on the other hand, were neither more adherent nor more infective than the parent. Furthermore, coagulase did not increase the pathogenicity ofclfA-positive streptococci when both clfA andcoa genes were simultaneously expressed in an artificial minioperon in streptococci. These results definitively attribute a role for ClfA, but not coagulase, in S. aureus endovascular infections. This gain-of-function strategy might help solve the role of individual factors in the complex the S. aureus-host relationship.

scholarly journals Bcl-2 antagonist apogossypol (NSC736630) displays single-agent activity in Bcl-2–transgenic mice and has superior efficacy with less toxicity compared with gossypol (NSC19048)

Blood ◽  
2008 ◽  
Vol 111 (6) ◽  
pp. 3211-3219 ◽  
Author(s):  
Shinichi Kitada ◽  
Christina L. Kress ◽  
Maryla Krajewska ◽  
Lee Jia ◽  
Maurizio Pellecchia ◽  
...  
Keyword(s):  
B Cells ◽  
Transgenic Mice ◽  
Parent Compound ◽  
Single Agent ◽  
Maximum Tolerated Dose ◽  
Low Grade ◽  
Altered Expression ◽  
Cell Counts

Abstract Altered expression of Bcl-2 family proteins plays central roles in apoptosis dysregulation in cancer and leukemia, promoting malignant cell expansion and contributing to chemoresistance. In this study, we compared the toxicity and efficacy in mice of natural product gossypol and its semisynthetic derivative apo-gossypol, compounds that bind and inhibit antiapoptotic Bcl-2 family proteins. Daily oral dosing studies showed that mice tolerate doses of apogossypol 2- to 4-times higher than gossypol. Hepatotoxicity and gastrointestinal toxicity represented the major adverse activities of gossypol, with apogossypol far less toxic. Efficacy was tested in transgenic mice in which Bcl-2 is overexpressed in B cells, resembling low-grade follicular lymphoma in humans. In vitro, Bcl-2–expressing B cells from transgenic mice were more sensitive to cytotoxicity induced by apogossypol than gossypol, with LD50 values of 3 to 5 μM and 7.5 to 10 μM, respectively. In vivo, using the maximum tolerated dose of gossypol for sequential daily dosing, apogossypol displayed superior activity to gossypol in terms of reducing splenomegaly and reducing B-cell counts in spleens of Bcl-2–transgenic mice. Taken together, these studies indicate that apogossypol is superior to parent compound gossypol with respect to toxicology and efficacy, suggesting that further development of this compound for cancer therapy is warranted.

scholarly journals Functional human genes typically exhibit epigenetic conservation

PLoS ONE ◽  
2021 ◽  
Vol 16 (9) ◽  
pp. e0253250
Author(s):  
Daniel Rud ◽  
Paul Marjoram ◽  
Kimberly Siegmund ◽  
Darryl Shibata
Keyword(s):  
Cell Lines ◽  
Drug Sensitivity ◽  
Single Gene ◽  
Functional Genes ◽  
Human Tissues ◽  
Gene Conservation ◽  
Individual Culture ◽  
Human Genes

Recent DepMap CRISPR-Cas9 single gene disruptions have identified genes more essential to proliferation in tissue culture. It would be valuable to translate these finding with measurements more practical for human tissues. Here we show that DepMap essential genes and other literature curated functional genes exhibit cell-specific preferential epigenetic conservation when DNA methylation measurements are compared between replicate cell lines and between intestinal crypts from the same individual. Culture experiments indicate that epigenetic drift accumulates through time with smaller differences in more functional genes. In NCI-60 cell lines, greater targeted gene conservation correlated with greater drug sensitivity. These studies indicate that two measurements separated in time allow normal or neoplastic cells to signal through conservation which human genes are more essential to their survival in vitro or in vivo.

Sign in / Sign up

Export Citation Format

Share Document