Glioma stem cells (GSCs) are a subpopulation of stem-like cells that contribute to glioblastoma (GBM) aggressiveness, recurrence, and resistance to radiation and chemotherapy. Therapeutically ...targeting the GSC population may improve patient survival, but unique vulnerabilities need to be identified.
We isolate GSCs from well-characterized GBM patient-derived xenografts (PDX), characterize their stemness properties using immunofluorescence staining, profile their epigenome including 5mC, 5hmC, 5fC/5caC, and two enhancer marks, and define their transcriptome. Fetal brain-derived neural stem/progenitor cells are used as a comparison to define potential unique and common molecular features between these different brain-derived cells with stem properties. Our integrative study reveals that abnormal expression of ten-eleven-translocation (TET) family members correlates with global levels of 5mC and 5fC/5caC and may be responsible for the distinct levels of these marks between glioma and neural stem cells. Heterogenous transcriptome and epigenome signatures among GSCs converge on several genes and pathways, including DNA damage response and cell proliferation, which are highly correlated with TET expression. Distinct enhancer landscapes are also strongly associated with differential gene regulation between glioma and neural stem cells; they exhibit unique co-localization patterns with DNA epigenetic mark switching events. Upon differentiation, glioma and neural stem cells exhibit distinct responses with regard to TET expression and DNA mark changes in the genome and GSCs fail to properly remodel their epigenome.
Our integrative epigenomic and transcriptomic characterization reveals fundamentally distinct yet potentially targetable biologic features of GSCs that result from their distinct epigenomic landscapes.
With the epidemic of obesity, nonalcoholic fatty liver disease (NAFLD) has become the most common pediatric liver disease. The influence of a perinatal obesity‐inducing diet (OID) on the development ...and progression of NAFLD in offspring is important but incompletely studied. Hence, we fed breeding pairs of C57BL/6J mice during gestation and lactation (perinatally) either chow or an OID rich in fat, fructose, and cholesterol (FFC). The offspring were weaned to either chow or an FFC diet, generating four groups: perinatal (p)Chow‐Chow, pChow‐FFC, pFFC‐Chow, and pFFC‐FFC. Mice were sacrificed at 10 weeks of age. We examined the whole‐liver transcriptome by RNA sequencing (RNA‐seq) and whole‐liver genome methylation by reduced representation bisulfite sequencing (RRBS). Our results indicated that the pFFC‐FFC mice had a significant increase in hepatic steatosis, injury, inflammation, and fibrosis, as assessed histologically and biochemically. We identified 189 genes that were differentially expressed and methylated in the pFFC‐FFC mice versus the pChow‐FFC mice. Gene set enrichment analysis identified hepatic fibrosis/hepatic stellate cell activation as the top canonical pathway, suggesting that the differential DNA methylation events in the mice exposed to the FFC diet perinatally were associated with a profibrogenic transcriptome. To verify that this finding was consistent with perinatal nutritional reprogramming of the methylome, we exposed pFFC‐Chow mice to an FFC diet in adulthood. These mice developed significant hepatic steatosis, injury, inflammation, and more importantly fibrosis when compared to the appropriate controls. Conclusion: Perinatal exposure to an OID primes the immature liver for an accentuated fibrosing nonalcoholic steatohepatitis (NASH) phenotype, likely through nutritional reprogramming of the offspring methylome. These data have potential clinical implications for monitoring children of obese mothers and risk stratification of children with NAFLD.
Perinatal exposure to high fat, fructose, and cholesterol (FFC) in mice primes the fetal liver to fibrosing NASH, likely secondary to nutritional reprograming of the liver methylome. This association is further confirmed, upon re‐exposure to the FFC diet in adulthood, where mice develop an accentuated, rapidly progressive NASH phenotype.
Mobile markets (MM) bring affordable, quality, healthy foods to high-need, low-food access communities. However, little is known about food insecurity of MM customers. This manuscript evaluates food ...insecurity prevalence in MM customers and assesses associations between food insecurity and MM use, food-related characteristics and behaviors, and fruit and vegetable (FV) intake. Customers (N = 302) completed cross-sectional surveys in summer 2019 that assessed: food security, food availability, cooking attitude, self-efficacy for healthy cooking, self-efficacy for cooking and eating FV, social connectedness, and FV intake. Descriptive and multivariate analyses were used to describe and assess associations with food insecurity and FV intake. Results show most MM customers were food insecure (85%). In logistic regression models adjusted for sociodemographic characteristics, long-term MM use (OR = 0.77, CI = 0.60–0.997), access to affordable, quality foods (OR = 0.81, CI = 0.71–0.93), and self-efficacy for both cooking healthy foods (OR = 0.88, CI = 0.80–0.97) and cooking and eating FV (OR = 0.90, CI = 0.82–0.98) were associated with lower odds of food insecurity; negative cooking attitudes (OR = 1.12, CI = 1.02–1.24) were associated with higher odds of food insecurity. Being food insecure (β = −1.37, SE=0.43, p < 0.01) was associated with poorer FV intake; this association attenuated slightly (β = −1.22, SE=0.43, p < 0.01) when length of MM use was added to the general linear model, which was also associated with higher fruit and vegetable intake (β = 0.26, SE=0.10, p = 0.01). Results suggest the MM reaches customers experiencing high levels of food insecurity and long-term MM use is associated with lower food insecurity and higher FV intake. Relationships between food insecurity and several food characteristics/behaviors provide insight for potential targets for wrap-around interventions to address food insecurity among customers. Findings suggest longitudinal evaluation of the MM's impact on food security and other food-related characteristics/behaviors is warranted.
To evaluate the outcomes of increasing mobile market service from mostly biweekly in 2019 to weekly in 2021.
Repeated, cross-sectional customer intercept surveys.
Mobile market customers in Summers ...2019 (N = 302) and 2021 (N = 72).
Mobile food markets bring affordable, high-quality foods to communities that lack such access.
Outcomes included food security, fruit/vegetable intake, and food-related characteristics and behaviors. General linear and logistic regression models were used to assess associations between outcomes and survey year and length of mobile market shopping. Models were adjusted for economic assistance use, race, and ethnicity.
No outcomes were significantly different between 2019 (with mostly biweekly service) and 2021 (with weekly service). Length of mobile market shopping (e.g., >2 years, 1-2 years, etc.) was positively associated with affordable, quality food access (β = 0.20, SE = 0.10, p = .03) and fruit/vegetable intake (β = 0.28, SE = 0.08, p < .001) as well as lower odds of food insecurity in the last 12 months (aOR = 0.79, 95% CI = 0.64, 0.99).
Despite COVID-19 interrupting scheduled market service, the length of time that a survey respondent identified as a full-service mobile market customer was associated with higher food access and fruit/vegetable intake and reduced food insecurity odds. These findings suggest promise and encourage further evaluation.
Here, we report the successful design, construction, and characterization of a 770-kilobase synthetic yeast chromosome II (synII). Our study incorporates characterization at multiple levels-including ...phenomics, transcriptomics, proteomics, chromosome segregation, and replication analysis-to provide a thorough and comprehensive analysis of a synthetic chromosome. Our Trans-Omics analyses reveal a modest but potentially relevant pervasive up-regulation of translational machinery observed in synII, mainly caused by the deletion of 13 transfer RNAs. By both complementation assays and SCRaMbLE (synthetic chromosome rearrangement and modification by
-mediated evolution), we targeted and debugged the origin of a growth defect at 37°C in glycerol medium, which is related to misregulation of the high-osmolarity glycerol response. Despite the subtle differences, the synII strain shows highly consistent biological processes comparable to the native strain.
Proliferative chronic myelomonocytic leukemia (pCMML), an aggressive CMML subtype, is associated with dismal outcomes. RAS pathway mutations, mainly NRAS
, define the pCMML phenotype as demonstrated ...by our exome sequencing, progenitor colony assays and a Vav-Cre-Nras
mouse model. Further, these mutations promote CMML transformation to acute myeloid leukemia. Using a multiomics platform and biochemical and molecular studies we show that in pCMML RAS pathway mutations are associated with a unique gene expression profile enriched in mitotic kinases such as polo-like kinase 1 (PLK1). PLK1 transcript levels are shown to be regulated by an unmutated lysine methyl-transferase (KMT2A) resulting in increased promoter monomethylation of lysine 4 of histone 3. Pharmacologic inhibition of PLK1 in RAS mutant patient-derived xenografts, demonstrates the utility of personalized biomarker-driven therapeutics in pCMML.
Introduction: Truncating mutations in the Additional Sex Combs-Like 1 (ASXL1) gene are associated with a proliferative disease phenotype and poor survival outcomes across the spectrum of myeloid ...malignancies including chronic myelomonocytic leukemia (CMML). ASXL1 is thought to act as a chromatin modifier regulating transcriptional activity, however the exact mechanisms and resulting chromatin states remain controversial. We interrogated the epigenome of 16 patients with ASXL1-mutant and -wildtype CMML using a multiomics approach.
Methods: Bone marrow mononuclear cells from patients with CMML (8 ASXL1-mutant, 8 -wildtype) were subjected to targeted NGS of DNA, whole transcriptome shotgun sequencing (RNA-seq), immunoprecipitation (IP) of DNA (hydroxy-)methyl residues (DIP-seq), IP of the histone modifications H3K4me1, H3K4me3, and H3K27me3 (ChIP-seq), and DNA transposase accessibility studies (ATAC-seq). After quality control all samples were sequenced on an Illumina HiSeq 4000 before further processing and data analysis. Global assessments of DNA (hydroxy-)methylation, DNA accessibility, and histone modifications between ASXL1-mutant and -wildtype CMML were performed. Differential gene expression was performed to define the up-regulated genes in ASXL1-mutant disease. The promoter regions of these up-regulated genes (defined as transcription start site ±3kb) were compared using the aforementioned multiomics approach. Epigenomic modification of the promoter region facilitating up-regulation of transcription was defined as the presence of a signal peak in ASXL1-mutant disease (in the absence of a signal peak in -wildtype disease) or 25% increase in a common signal peak (H3K4me1/3, 5hmC), the presence of a unique signal peak in ASXL1-mutant disease (ATAC), or the absence of a signal peak in ASXL1-mutant disease (in the presence of a signal peak in -wildtype disease), or 25% decrease in a common signal peak (H3K27me3, 5mC).
Results: Sixteen patients with CMML, median age 69 years (48 - 77), 63% male, were included. Half of the patients had proliferative disease (pCMML) and half of them had truncating frameshift mutations in ASXL1 (heatmap). All ASXL1 variant allele frequencies were compatible with heterozygosity (31 - 48%). The burden of co-mutations was similar between ASXL1-wildtype and ASXL1-mutant disease (21 versus 23 per group; no difference in the median number of co-mutations, p = 0.684). The spectrum of co-mutations was typical for CMML, involving spliceosome components, epigenetic regulators, chromatin regulators, and cell signaling molecules (heatmap). There was a predominant up-regulation of gene expression in ASXL1-mutant patients: 707 genes up- and 124 down-regulated (volcano plot, FDR < 0.05 for all genes). Functional annotation of the up-regulated genes showed cell division, mitotic nuclear division, sister chromatid cohesion, DNA replication, and G1/S transition to be the 5 most enriched processes (accounting for 29% of all up-regulated genes, FDR < 1x10-10 for all terms). The up-regulated genes included several potential therapeutic targets and HOXA family members (including HOXA9). There were global increases in H3K4me1/3, 5mC, and 5hmC, decreases in H3K27me3, as well as a more relaxed chromatin conformation (bar graphs). Many of these epigenomic changes affected non-coding regions. When focusing on the promoter regions of the 707 up-regulated genes there was evidence of one or more of the interrogated epigenomic mechanisms facilitating transcription for 519 of the genes (73%). The most abundant mechanism was histone modification, followed by changes in DNA (hydroxy-)methylation, and increased chromatin accessibility, with considerable overlap (Venn diagram). For HOXA9, a known driver of leukemogenesis, the data supported a loss of H3K27me3 as the most prominent among the interrogated epigenomic regulatory mechanisms (average signal tracks).
Conclusions: The transcriptome and chromatin conformation of ASXL1-mutant CMML are skewed towards proliferation and mirror the aggressive disease phenotype observed in practice. There is evidence of histone modification as well as changes in DNA methylation, and chromatin conformation facilitating transcriptional activity including known leukemogenic drivers. Additional regulatory mechanisms such as gene body methylation and enhancer elements require further exploration.
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Patnaik:Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.
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