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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 109 records · Page 6
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

EIF3H knockdown inhibits malignant melanoma through regulating cell proliferation, apoptosis and cell cycle

Highlights: • EIF3H is highly expressed in malignant melanoma and correlated with tumor stage and risk of lymphatic metastasis. • Knockdown of EIF3H inhibits cell proliferation, cell migration and promotes cell apoptosis of melanoma in vitro. • Knockdown of EIF3H suppresses tumor growth in vivo. Malignant melanoma (MM) causes 80% of skin cancer-related deaths and becomes the most lethal type of skin cancer. The molecular mechanism of MM is still not clear. This study aimed to reveal the relationship between MM and EIF3H. Clinical specimens were collected to preliminarily explore the role of EIF3H in MM. MM cell lines with EIF3H knockdown were constructed for investigating the effects of EIF3H on cell proliferation, apoptosis, cell cycle and cell motility. Mice xenograft model was constructed for verification in vivo. We found that EIF3H was obviously upregulated in MM tissues compared with normal skin tissues, which was correlated with tumor stage and risk of lymphatic metastasis. The in vitro results indicated that silencing EIF3H in MM cells could significantly suppress cell proliferation, promote cell apoptosis and induce cell cycle arrest. Moreover, EIF3H knockdown significantly restrained cell motility through regulating EMT-related proteins. The effects of EIF3H knockdown were also verified in mice xenograft model, which were represented by slower growth rate, smaller volume and lighter weight of tumors. Therefore, EIF3H was identified as a critical factor in the development and progression of MM which may be used as a novel therapeutic target in the treatment of MM.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

The NRG3/ERBB4 signaling cascade as a novel therapeutic target for canine glioma

Highlights: • miR-190a was downregulated in canine brain tumors. • miR-190a functioned as a tumor-suppressor through targeting NRG3. • A pan-ERBB inhibitor exhibited a tumor suppressive effect in canine glioma cells. Canine glioma is a common brain tumor with poor prognosis despite surgery and/or radiation therapy. Therefore, newer and more effective treatment modalities are needed. Neuregulin 3 (NRG3) has known to be a ligand of ERBB4. This study aimed to investigate the usefulness of the NRG3/ERBB4 signaling cascade as a novel therapeutic target in canine glioma. We found out that microRNA (miR)-190a was downregulated in canine brain tumor tissues, including glioma and meningioma. miR-190a directly targeted NRG3 and inhibited the growth of canine glioma cells. The level of p-Akt, which is a downstream target of ERBB4 signaling, was decreased by transfection with miR-190a. NRG3 silencing also suppressed cell growth and decreased the levels of p-Akt and p-ERK1/2, and NRG3 overexpression exhibited opposed effects in canine glioma J3T-1 cells. The mRNA level of erbb4 was significantly upregulated in glioma tissues compared with that in normal brain tissues and meningioma tissues. Furthermore, compared with gefitinib and lapatinib, afatinib exerted a greater inhibitory effect on the growth of canine glioma cells. In conclusion, NRG3/ERBB4 signaling is negatively regulated by miR-190a and contributes to the growth of canine glioma cells, indicating that it may be a promising therapeutic target in canine glioma.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Adipose-specific knockout of Protein Kinase D1 suppresses de novo lipogenesis in mice via SREBP1c-dependent signaling

Highlights: • Protein Kinase D1 is a functional regulator in adipocyte and white adipose tissue. • PKD1 drives de novo lipogenesis in adipocyte. • SREBP1c signaling regulates lipogenesis in adipocyte. • PKD1-mediated de novo lipogenesis relies on SREBP1c. Having healthy adipose tissue is essential for metabolic health, as excessive adipose tissue in the body can cause its dysregulation and driving chronic metabolic diseases. Protein kinase D1 (PKD1) is considered to be a key kinase in signal transduction, which regulates multiple cellular functions, but its physiological functions in adipose are still not fully understood. This study aimed at elucidating the function of adipocyte PKD1 on lipogenesis. From RNA-Sequencing data, we found that the fatty acid biosynthesis pathway in white adipose tissue lacking PKD1 was significantly affected. Critical rate-limiting enzymes for de novo lipogenesis in adipocytes, such as FASN, ACCα, and SCD1, were significantly repressed after deleting PKD1 in vivo and in vitro. Further studies revealed that blockade of PKD1 significantly increased phosphorylation of SREBP1c at serine 372 site. Co-immunoprecipitation analysis showed that PKD1 interacts with SREBP1c in vitro and in vivo. Importantly, overexpression of SREBP1c reversed the inhibition of FASN and ACCα expression caused by PKD1 silencing. Together, adipocyte PKD1 promotes de novo lipogenesis via SREBP1c-dependent manner in visceral white adipose tissue and might provide a new target for the development of anti-obesity therapies.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Mitoferrin 2 deficiency prevents mitochondrial iron overload-induced endothelial injury and alleviates atherosclerosis

Endothelial dysfunction is an early step in the development of atherosclerotic cardiovascular disease. Iron overload can lead to excessive mitochondrial reactive oxygen species (mtROS) production, resulting in mitochondrial dysfunction and vascular endothelial cell (EC) damage. Mitoferrin 2 (Mfrn2) is an iron transporter in the inner mitochondrial membrane. This study aimed to assess whether Mfrn2 and mitochondrial iron overload were involved in atherosclerosis progression and to explore the potential mechanism. We observed significant upregulation of Mfrn2 in the arteries of high-fat diet (HFD)-fed Apolipoprotein E{sup −/-} (ApoE{sup −/-}) mice and in TNF-α-induced mouse aortic endothelial cells (MAECs). Mfrn2 gene silencing inhibited mitochondrial iron overload, stabilized mitochondrial membrane potential and improved mitochondrial function in TNF-α-induced MAECs. Vascular EC-specific knockdown of Mfrn2 in ApoE{sup −/-} mice markedly decreased atherosclerotic lesion formation and the levels of ICAM-1 in aortas and reduced monocyte infiltration into the vascular wall. Furthermore, TNF-α increased the binding of 14-3-3 epsilon (ε) and Mfrn2, preventing Mfrn2 degradation and leading to mitochondrial iron overload in ECs, while 14-3-3ε overexpression increased Mfrn2 stability by inhibiting its ubiquitination. Together, our results reveal that Mfrn2 deficiency attenuates endothelial dysfunction by decreasing iron levels within the mitochondria and mitochondrial dysfunction. These findings may provide new insights into preventive and therapeutic strategies against vascular endothelial dysfunction in atherosclerotic disease.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Indole-3-carbinol ameliorates necroptosis and inflammation of intestinal epithelial cells in mice with ulcerative colitis by activating aryl hydrocarbon receptor

Highlights: • AHR activated by I3C plays an important role in protecting IECs from necroptosis and inflammation, in vivo and in vitro. • AHR could ubiquitinate RIPK1, promote IAPs expression and inhibit NF-κB activation in NCM460 cells. • I3C is a good candidate as a natural product to prevent and treat UC. Ulcerative colitis (UC) is a disease characterized by inflammation and disruption of the intestinal epithelial barrier. Necroptosis plays a critical role in disease progression. Indole-3-carbinol (I3C), a natural dietary agonist of aryl hydrocarbon receptor (AHR), has shown alleviating effects on UC. However, its mechanisms of action have not been comprehensively elucidated. Therefore, we aimed at investigating the protective role of I3C in DSS-induced colitis mice models. I3C significantly ameliorated body weight loss, colon length shortening and colonic pathological damage in colitis mice, reduced disease activity index (DAI) and histological (HI) scores, as well as alleviated colonic necroptosis and inflammation. In vitro, I3C attenuated necroptosis and inflammation of colonoids and NCM460 cells. AHR, activated by I3C, inhibits activation of receptor-interacting protein kinase 1 (RIPK1) and the subsequent assembly of necrosome in a time-dependent manner, as well as suppressing NF-κB activation and decreasing TNF-α, IL-1β, IL-6 and IL-8 expression. Silencing of AHR aggravated necroptosis and inflammation of NCM460 cells, and did not be ameliorated by I3C. Furthermore, AHR activation induces the expression of inhibitor of apoptosis proteins (IAPs) and the ubiquitination of RIPK1. In conclusion, I3C exerts a protective effect in DSS-induced colitis mice models by alleviating the necroptosis and inflammation of IECs through activating AHR.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

SMYD2 suppresses p53 activity to promote glucose metabolism in cervical cancer

Reprogrammed energy metabolism, especially the Warburg effect, is emerged as a hallmark of cancer. The protein lysine methyltransferase SMYD2 functions as an oncogene and is implicated in various malignant phenotypes of human cancers. However, the role of SMYD2 in tumor metabolism is still largely unknown. Here, we report that SMYD2 is highly expressed in human cervical cancer and its aberrant expression is linked to a poor prognosis. Bioinformatic analysis revealed a novel link between SMYD2 expression and aerobic glycolysis. Through loss-of-function experiments, we demonstrated that SMYD2 knockdown or inhibition induced a metabolic shift from aerobic glycolysis to oxidative phosphorylation, as evidenced by glucose uptake, lactate production, extracellular acidification, and the oxygen consumption rate. In contrast, SMYD2 overexpression promoted glycolytic metabolism in cervical cancer cells. Moreover, SMYD2 was required for tumor growth in cervical cancer and this oncogenic activity was largely glycolysis-dependent. Mechanistically, SMYD2 altered the methylation status of p53 and inhibited its transcriptional activity. Genetic silencing of p53 largely abrogated the effects of SMYD2 in promoting aerobic glycolysis. Taken together, our findings reveal a novel function of SMYD2 in regulating the Warburg effect in cervical cancer.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

MIER3 induces epithelial-mesenchymal transition and promotes breast cancer cell aggressiveness via forming a co-repressor complex with HDAC1/HDAC2/Snail

Highlights: • The expression of MIER3 was correlated with poor outcome of breast cancer patients. • MIER3 induced EMT and promoted cell migration. • MIER3 interacted with HDAC1/HDAC2 and Snail to form a repressive complex. • The repressive complex bound to E-cadherin promoter and reduced its acetylation. Breast cancer is one of the most frequently diagnosed cancers and the leading cause of cancer death in women. MIER3 (Mesoderm induction early response 1, family member3) is considered as a potential oncogene for breast cancer. However, the role of MIER3 in breast cancer remain largely unknown. The expression of MIER3 was detected and the relationship between its expression and clinicopathological characteristics was also analyzed. The effect of MIER3 on proliferation and migration of breast cancer cells was detected in vitro and in vivo. Western blot, IF, and Co-IP were employed to detect the relationship between MIER3, HDAC1, HDAC2, and Snail. ChIP assay was performed to determine the binding of MIER3/HDAC1/HDAC2/Snail complex to the promoter of E-cadherin. In this study, we found that MIER3 was upregulated in breast cancer tissue and closely associated with poor prognosis of patients. MIER3 could promote the proliferation, migration, and epithelial-mesenchymal transition (EMT) of breast cancer cells. Further studies showed that MIER3 interacted with HDAC1/HDAC2 and Snail to form a repressive complex which could bind to E-cadherin promoter and was related to its deacetylation. Our study concluded that MIER3 was involved in forming a co-repressor complex with HDAC1/HDAC2/Snail to promote EMT by silencing E-cadherin.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

GLIDR promotes the progression of glioma by regulating the miR-4677-3p/MAGI2 axis

Gliomas are the most common and fatal primary brain tumors. Growing evidence suggests that long non-coding RNAs (lncRNAs) constitute novel and potential therapeutic targets for glioma. However, the biological role of glioblastoma down-regulated RNA (GLIDR) in glioma remains largely elusive. In the current study, we used quantitative real-time polymerase chain reaction (qRT-PCR) to detect GLIDR expression in glioma cells. Cell counting kit 8 (CCK-8) assay, colony formation assay, JC-1 staining, and flow cytometry were used to evaluate the role of GLIDR in proliferation and apoptosis of glioma cells. Western blotting was performed to assess the effect of GLIDR on the level of apoptosis-related proteins. In addition, bioinformatics prediction, RNA immunoprecipitation (RIP), RNA pull-down, and luciferase reporter gene assays were used to study the regulatory mechanisms of GLIDR in glioma. GLIDR was found to be highly expressed in glioma cells and silencing of GLIDR inhibited cell proliferation and promoted apoptosis. Functionally, GLIDR bound to miR-4677-3p that directly targeted membrane-associated guanylate kinase, WW, and PDZ domain-containing protein 2 (MAGI2). Our data showed that GLIDR affects the proliferation and apoptosis of glioma cells by targeting miR-4677-3p to regulate the expression of MAGI2. In conclusion, our study determined the oncogenic role of GLIDR in glioma, which may provide a new perspective for the treatment of glioma.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

De-ubiquitination of p300 by USP12 Critically Enhances METTL3 Expression and Ang II-induced cardiac hypertrophy

Highlights: • The expression of USP12 is increased in Ang II-induced cardiac hypertrophy in vitro and in vivo. • USP12 aggravates Ang II-induced cardiomyocyte hypertrophy via upregulating METTL3. • USP12 promoted METTL3 expression via modulating p300. • USP12 suppresses the ubiquitination of p300 to block its degradation. Stresses, such as neurohumoral activation, induced pathological cardiac hypertrophy is the main risk factor for heart failure. The ubiquitin-proteasome system (UPS) plays a key role in maintaining protein homeostasis and cardiac function. However, research on the role and mechanism of deubiquitinating enzymes (DUBs) in cardiac hypertrophy is limited. Here, we observe that the deubiquitinating enzyme ubiquitin-specific protease 12(USP12) is upregulated in Ang II-induced hypertrophic hearts and primary neonatal rat cardiomyocytes (NRCMs). Inhibition of USP12 ameliorate Ang II-induced myocardial hypertrophy, while overexpression of USP12 have the opposite effect. USP12 deficiency also significantly attenuate the phenotype of Ang II-induced cardiac hypertrophy in vivo. Moreover, we demonstrate that USP12 aggravate Ang II-induced cardiac hypertrophy by enhancing METTL3, a methyltransferase which catalyze N{sup 6}-methyladenosine (m{sup 6}A) modification on messenger RNA and acts as a harmful factor in pathological cardiac hypertrophy. Upregulation of METTL3 reverse the reduction of myocardial hypertrophy induced by USP12 silencing in NRCMs. In contrast, knockdown of METTL3 attenuate the aggravation of myocardial hypertrophy in USP12-overexpressing NRCMs. Furthermore, we discover that USP12 promote the expression of METTL3 via upregulating p300. Mechanistically, USP12 binds and stabilizes p300, thereby activating the transcription of its downstream gene METTL3. Finally, our data show that USP12 is partially dependent on the stabilization of p300 to activate METTL3 expression and promote myocardial hypertrophy. Taken together, our results demonstrate that USP12 acts as a pro-hypertrophic deubiquitinating enzyme via enhancing p300/METTL3 axis, indicating that targeting USP12 could be a potential treatment strategy for pathological cardiac hypertrophy.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

MIG/CXCL9 exacerbates the progression of metabolic-associated fatty liver disease by disrupting Treg/Th17 balance

CD4{sup +}CD25{sup +} regulatory T (Treg) cells and Th17 cells play important roles in the progression of metabolic-associated fatty liver disease (MAFLD). However, the contribution of monokine induced by interferon-gamma (MIG)/CXCL9 to the Treg/Th17 imbalance in MAFLD is only partially understood. In the present study, we detected increased levels of MIG/CXCL9 and a Treg/Th17 imbalance in the setting of metabolic-associated steatohepatitis (MASH). Recombinant adeno-associated virus-mediated gene transfer and silencing of MIG/CXCL9 expression in mice alleviated MASH and increased the Treg/Th17 ratio. Furthermore, the percentage of Th17 cells, but not Treg cells, differentiated from splenic CD4{sup +} T cells was significantly increased by administration of MIG/CXCL9. MIG/CXCL9 also promoted Th17 cell proliferation, and its effects were dose dependent. Levels of phosphorylated c-Jun N-terminal kinase (JNK) decreased dramatically when MIG/CXCL9 was inhibited in a murine MASH model. In cultured Treg cells, phosphorylated JNK levels decreased dose-dependently in response to MIG/CXCL9 inhibition, but increased in cultured Th17 cells. This effect was blocked in the presence of a JNK inhibitor. These findings underline the fundamental importance of MIG/CXCL9 in maintaining the Treg/Th17 balance in MAFLD and provide the foundations for a novel approach to preventing and treating MAFLD.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

The oncogenic role of meiosis-specific Aurora kinase C in mitotic cells

Highlights: • AURKC is expressed in meiotic cells and some cancers. • AURKC expression increases cell migration and transformation of human mitotic cells. • Deletion of AURKC in U2OS cells decreases cell migration and soft agar growth. • Transcriptome analysis reveals that AURKC alters the extracellular matrix. Aberrant expression of meiosis-specific genes in cancer has recently emerged as a driver of some cancer formation. Aurora kinase C (AURKC) is a member of the Aurora kinase family of proteins known to regulate chromosome segregation during cell divisions. AURKC is normally expressed in meiotic cells; however, elevated levels of AURKC mRNA and protein are frequently measured in cancer cells. To understand the function of AURKC in cancer cells, expression was induced in noncancerous, human retina pigmented epithelial cells. While AURKC expression did not alter cell proliferation over 72 h, it did increase cell migration and anchorage independent growth in soft agar suggesting an oncogenic role in mitotically dividing cells. To evaluate AURKC as a potential therapeutic target, a frameshift mutation in the gene was introduced in U2OS osteosarcoma cells using CRISPR-Cas9 technology resulting in a premature stop codon. Cancer cells lacking AURKC displayed no change in cell proliferation over 72 h but did migrate less and formed fewer colonies in soft agar. Whole transcriptome sequencing analysis uncovered over 400 differentially expressed genes in U2OS cells with and without AURKC. GO analysis revealed alterations in proteinaceous extracellular matrix genes including COL1A1. These data indicate that therapeutics targeting AURKC could decrease cancer cell metastasis and disease progression. Because AURKC is transcriptionally silenced in normal mitotic cells, its disruption could specifically target cancer cells limiting the toxic side effects associated with current therapeutics.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

G protein nucleolar 3 promotes Non-Hodgkin lymphoma progression by activating the Wnt/β-catenin signaling pathway

G protein nucleolar 3 (GNL3), which acts as an oncoprotein in various carcinomas, is associated with tumor progression; however, little is known regarding GNL3 function in non-Hodgkin lymphoma (NHL). In this study, we first used in silico analysis to determine associations between GNL3 and diffuse large B-cell lymphoma (DLBCL). We then examined the effect of GNL3 on NHL progression, including cell proliferation, apoptosis, and cell cycle progression, and determined its underlying molecular mechanism using in vitro lymphoma cell lines and in vivo mouse xenograft models. We found that GNL3 mRNA levels were markedly higher in DLBCL tissues than in normal tissues, with these higher levels associated with poor prognosis. Additionally, GNL3 overexpression promoted NHL cell proliferation and cell cycle progression and reduced apoptosis in vitro, and enhanced tumorigenesis in an in vivo xenograft model. Moreover, we found that GNL3 upregulated the levels of Wnt/β-catenin signaling pathway-related factors and downstream target genes, whereas the opposite result was observed in GNL3-silenced cells. Furthermore, a rescue experiment using a Wnt/β-catenin inhibitor (XAV939) confirmed that GNL3 promotes NHL progression by activating the Wnt/β-catenin signaling pathway. These findings demonstrated that GNL3 functions as an oncogenic driver in NHL via the Wnt/β-catenin pathway.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

CLEC12B suppresses lung cancer progression by inducing SHP-1 expression and inactivating the PI3K/AKT signaling pathway

Lung cancer is the leading cause of cancer mortality worldwide. CLEC12B, a C-type lectin-like receptor, is low-expressed in lung cancer tissues. However, the function of CLEC12B in lung cancer and its underlying mechanism remain unclear. Here, an obvious down-regulation of CLEC12B was observed in lung cancer cells compared with the normal lung epithelial cells. CLEC12B over-expression suppressed cell viability and cell cycle entry in lung cancer, along with the reduction of PCNA and cyclin D1 expressions, while silencing CLEC12B possessed the opposite effects. Over-expression of CLEC12B promoted lung cancer cell apoptosis, accompanied by decreased Bcl-2 and increased Bax, cleaved caspase-3 and cleaved caspase-9. Moreover, CLEC12B decreased phosphorylation of PI3K-p85 and AKT proteins. By contrast, CLEC12B knockdown activated the PI3K/AKT pathway. In vivo, CLEC12B inhibited tumor growth in lung cancer, which can be reversed by CLEC12B inhibition. Co-IP and immunofluorescence assays confirmed the interaction between CLEC12B and SHP-1, and CLEC12B over-expression increased SHP-1 level. Furthermore, knocking down SHP-1 abrogated the above biological phenotypes caused by CLEC12B elevation. Taken together, our findings demonstrate that CLEC12B serves as a tumor-suppressing gene in lung cancer through positively regulating SHP-1 expression, which may be mediated by the PI3K/AKT signaling pathway.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2022

Poly( N -vinylpyrrolidone)- block -Poly(dimethylsiloxane)- block -Poly( N -vinylpyrrolidone) Triblock Copolymer Polymersomes for Delivery of PARP1 siRNA to Breast Cancers

Nearly 20% of HER2-positive breast cancers develop resistance to HER2-targeted therapies requiring the use of advanced therapies. Silencing RNA therapy may be a powerful modality for treating resistant HER2 cancers due to its high specificity and low toxicity. However, the systemic administration of siRNAs requires a safe and efficient delivery platform because of siRNA’s low stability in physiological fluids, inefficient cellular uptake, immunoreactivity, and rapid clearance. We have developed theranostic polymeric vesicles to overcome these hurdles for encapsulation and delivery of small functional molecules and PARP1 siRNA for in vivo delivery to breast cancer tumors. The 100 nm polymer vesicles were assembled from biodegradable and non-ionic poly(N-vinylpyrrolidone) 14 -block-poly(dimethylsiloxane) 47 -block-poly(N-vinylpyrrolidone) 14 triblock copolymer PVPON 14 -PDMS 47 -PVPON 14 using nanoprecipitation and thin-film hydration. We demonstrated that the vesicles assembled from the copolymer covalently tagged with the Cy5.5 fluorescent dye for in vivo imaging could also encapsulate the model drug with high loading efficiency (40%). The dye-loaded vesicles were accumulated in tumors after 18 h circulation in 4TR breast tumor-bearing mice via passive targeting. We found that PARP1 siRNA encapsulated into the vesicles was released intact (13%) into solution by the therapeutic ultrasound treatment as quantified by gel electrophoresis. Additionally, the PARP1 siRNA-loaded polymersomes inhibited the proliferation of MDA-MB-361TR cells by 34% after 6 days of treatment by suppressing the NF-kB signaling pathway, unlike their scrambled siRNA-loaded counterparts. Finally, the treatment by PARP1 siRNA-loaded vesicles prolonged the survival of the mice bearing 4T1 breast cancer xenografts, with the 4-fold survival increase, unlike the untreated mice after 3 weeks following the treatment. These biodegradable, non-ionic PVPON 14 -PDMS 47 -PVPON 14 polymeric nanovesicles capable of the efficient encapsulation and delivery of PARP1 siRNA to successfully knock down PARP1 in vivo can provide an advanced platform for the development of precision-targeted therapeutic carriers, which could help develop highly effective drug delivery nanovehicles for breast cancer gene therapy.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Controlled Organization of Inorganic Materials Using Biological Molecules for Activating Therapeutic Functionalities

Precise control over the assembly of biocompatible three-dimensional (3D) nanostructures would allow for programmed interactions within the cellular environment. Nucleic acids can be used as programmable crosslinkers to direct the assembly of quantum dots (QDs) and tuned to demonstrate different interparticle binding strategies. In this work, morphologies of self-assembled QDs are evaluated via gel electrophoresis, transmission electron microscopy, small-angle X-ray scattering, and dissipative particle dynamics simulations, with all results being in good agreement. The controlled assembly of 3D QD organizations is demonstrated in cells via the colocalized emission of multiple assembled QDs, and their immunorecognition is assessed via enzyme-linked immunosorbent assays. RNA interference inducers are also embedded into the interparticle binding strategy to be released in human cells only upon QD assembly, which is demonstrated by specific gene silencing. The programmability and intracellular activity of QD assemblies offer a strategy for nucleic acids to imbue the structure and therapeutic function into the formation of complex networks of nanostructures, while the photoluminescent properties of the material allow for optical tracking in cells in vitro.

36 MATERIALS SCIENCE↗
DOE OSTI2021

Structures of monomeric and dimeric PRC2:EZH1 reveal flexible modules involved in chromatin compaction

Polycomb repressive complex 2 (PRC2) is a histone methyltransferase critical for maintaining gene silencing during eukaryotic development. In mammals, PRC2 activity is regulated in part by the selective incorporation of one of two paralogs of the catalytic subunit, EZH1 or EZH2. Each of these enzymes has specialized biological functions that may be partially explained by differences in the multivalent interactions they mediate with chromatin. Here, we present two cryo-EM structures of PRC2:EZH1, one as a monomer and a second one as a dimer bound to a nucleosome. When bound to nucleosome substrate, the PRC2:EZH1 dimer undergoes a dramatic conformational change. We demonstrate that mutation of a divergent EZH1/2 loop abrogates the nucleosome-binding and methyltransferase activities of PRC2:EZH1. Finally, we show that PRC2:EZH1 dimers are more effective than monomers at promoting chromatin compaction, and the divergent EZH1/2 loop is essential for this function, thereby tying together the methyltransferase, nucleosome-binding, and chromatin-compaction activities of PRC2:EZH1. We speculate that the conformational flexibility and the ability to dimerize enable PRC2 to act on the varied chromatin substrates it encounters in the cell.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

The intrinsically disordered protein TgIST from Toxoplasma gondii inhibits STAT1 signaling by blocking cofactor recruitment

Signal transducer and activator of transcription (STAT) proteins communicate from cell-surface receptors to drive transcription of immune response genes. The parasite Toxoplasma gondii blocks STAT1-mediated gene expression by secreting the intrinsically disordered protein TgIST that traffics to the host nucleus, binds phosphorylated STAT1 dimers, and occupies nascent transcription sites that unexpectedly remain silenced. Here we define a core region within internal repeats of TgIST that is necessary and sufficient to block STAT1-mediated gene expression. Cellular, biochemical, mutational, and structural data demonstrate that the repeat region of TgIST adopts a helical conformation upon binding to STAT1 dimers. The binding interface is defined by a groove formed from two loops in the STAT1 SH2 domains that reorient during dimerization. TgIST binding to this newly exposed site at the STAT1 dimer interface alters its conformation and prevents the recruitment of co-transcriptional activators, thus defining the mechanism of blocked transcription.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

The cholesterol transporter NPC1 is essential for epigenetic regulation and maturation of oligodendrocyte lineage cells

The intracellular cholesterol transporter NPC1 functions in late endosomes and lysosomes to efflux unesterified cholesterol, and its deficiency causes Niemann–Pick disease Type C, an autosomal recessive lysosomal disorder characterized by progressive neurodegeneration and early death. Here, we use single-nucleus RNA-seq on the forebrain of Npc1 –/– mice at P16 to identify cell types and pathways affected early in pathogenesis. Our analysis uncovers significant transcriptional changes in the oligodendrocyte lineage during developmental myelination, accompanied by diminished maturation of myelinating oligodendrocytes. We identify upregulation of genes associated with neurogenesis and synapse formation in Npc1 –/– oligodendrocyte lineage cells, reflecting diminished gene silencing by H3K27me3. Npc1 –/ – oligodendrocyte progenitor cells reproduce impaired maturation in vitro, and this phenotype is rescued by treatment with GSK-J4, a small molecule inhibitor of H3K27 demethylases. Moreover, mobilizing stored cholesterol in Npc1 –/– mice by a single administration of 2-hydroxypropyl-β-cyclodextrin at P7 rescues myelination, epigenetic marks, and oligodendrocyte gene expression. Our findings highlight an important role for NPC1 in oligodendrocyte lineage maturation and epigenetic regulation, and identify potential targets for therapeutic intervention.

59 BASIC BIOLOGICAL SCIENCES↗