Transcription Control Pathways Research Articles

Transcription factors SltA and CrzA reversely regulate calcium homeostasis under calcium-limited conditions.

Calcium ions are ubiquitous intracellular signaling molecules for many signaling pathways regulating the fungal response to stress and antifungal drugs. The concentration of intracellular calcium is tightly regulated in its storage, release, and distribution. CrzA is the best-studied transcription factor that regulates this process under sufficient calcium or other external signals. However, CrzA was excluded from nuclei and then lost transcriptional activation under calcium-limited conditions. The regulators in the Ca2+ signaling pathway under calcium-limited conditions remain unclear. Here, we identified SltA as a key regulator in the Ca2+ signaling pathway under calcium-limited conditions, and the underlying mechanisms were further explored in Aspergillus fumigatus. These findings reveal a transcriptional control pathway that precisely regulates calcium homeostasis under calcium-limited conditions.

Daily heart rate variability biofeedback practice sessions affect plasma Αβ40 and Αβ42 levels in healthy younger and older adults: A randomized clinical trial (N = 108)

AbstractBackgroundGenetic mutations affecting β‐amyloid (Αβ) production suggest that decreasing Αβ levels in healthy adults could prevent AD. However, evidence for behavioral interventions that decrease Αβ levels is lacking. The current study examined whether heart rate variability (HRV) biofeedback affects plasma Αβ levels. This intervention involves breathing slowly at a pace that maximizes the amplitude of heart rate oscillations. During slow paced breathing, the vagus nerve receives strong signals from stretch receptors in blood vessels and the lungs. These afferent signals stimulate ascending vagus nerve signaling that activate ‘safety’ signaling pathways, suppressing stress and arousal pathways in the brain and body. Experiencing adversity increases production of Aβ and risk of AD. We hypothesized that, in healthy adults, daily sessions involving slow paced breathing during HRV biofeedback could decrease Aβ by suppressing these adversity‐related processes.MethodHealthy adults (N = 54 age 18‐35; N = 54 age 55‐80) practiced HRV biofeedback daily for five weeks. They were randomized to use slow paced breathing and HRV biofeedback to increase heart rate oscillations (Osc+ condition) or to use personalized strategies and HRV biofeedback to try to decrease heart rate oscillations (Osc‐ condition). Pre‐ and post‐intervention plasma Aβ40 and Aβ42 levels were quantified blind to condition. Gene expression analysis of peripheral blood samples of the 54 younger adults using bioinformatics‐based analyses of genome‐wide RNA profiles examined activity of pro‐inflammatory, neuroendocrine, and antiviral transcription control pathways relevant to the conserved transcriptional response to adversity (CTRA). We predicted that reductions in the CTRA across the 5‐week trial would be associated with reduced plasma Aβ.ResultBoth younger and older adults showed significant differences between Osc+ and Osc‐ conditions in change in plasma Aβ40 and Aβ42 levels (Figure 1). The Osc+ condition decreased Αβ while the Osc‐ condition increased Αβ. Individual changes in plasma Aβ from pre‐ to post‐intervention were associated with changes in expression of the CTRA in the predicted direction.ConclusionHRV biofeedback affects plasma Aβ and these effects are associated with changes in expression of genes upregulated during adversity. Future longer‐term studies should test whether reducing plasma Αβ via the Osc+ intervention reduces risk of developing AD.

Multiomics Analysis Reveals Novel Genetic Determinants for Lens Differentiation, Structure, and Transparency.

Recent advances in next-generation sequencing and data analysis have provided new gateways for identification of novel genome-wide genetic determinants governing tissue development and disease. These advances have revolutionized our understanding of cellular differentiation, homeostasis, and specialized function in multiple tissues. Bioinformatic and functional analysis of these genetic determinants and the pathways they regulate have provided a novel basis for the design of functional experiments to answer a wide range of long-sought biological questions. A well-characterized model for the application of these emerging technologies is the development and differentiation of the ocular lens and how individual pathways regulate lens morphogenesis, gene expression, transparency, and refraction. Recent applications of next-generation sequencing analysis on well-characterized chicken and mouse lens differentiation models using a variety of omics techniques including RNA-seq, ATAC-seq, whole-genome bisulfite sequencing (WGBS), chip-seq, and CUT&RUN have revealed a wide range of essential biological pathways and chromatin features governing lens structure and function. Multiomics integration of these data has established new gene functions and cellular processes essential for lens formation, homeostasis, and transparency including the identification of novel transcription control pathways, autophagy remodeling pathways, and signal transduction pathways, among others. This review summarizes recent omics technologies applied to the lens, methods for integrating multiomics data, and how these recent technologies have advanced our understanding ocular biology and function. The approach and analysis are relevant to identifying the features and functional requirements of more complex tissues and disease states.

The Effect of Human and Bovine Milk Osteopontin on Intestinal Caco-2 Cells: A Transcriptome Comparison

Osteopontin (OPN) is a multifunctional protein abundantly present in human milk, whereas the concentration is significantly lower in bovine milk. Human and bovine milk OPN are structurally similar and both proteins resist gastric digestion and reach the intestines in a bioactive form. Intervention studies have indicated the beneficial effects of supplementing infant formula with bovine milk OPN and several in vivo and in vitro studies have shown that bovine milk OPN positively influences intestinal development. To investigate the functional relationship, we compared the effect of simulated gastrointestinal digested human and bovine milk OPN on gene expression in Caco-2 cells. After incubation, total RNA was extracted and sequenced and transcripts were mapped to the human genome. Human and bovine milk OPN regulated the expression of 239 and 322 genes, respectively. A total of 131 genes were similarly regulated by the OPNs. As a control, a whey protein fraction with a high content of alpha-lactalbumin had a very limited transcriptional impact on the cells. Enrichment data analysis showed that biological processes related to the ubiquitin system, DNA binding, and genes associated with transcription and transcription control pathways were affected by the OPNs. Collectively, this study shows that human and bovine milk OPN have a significant and highly comparable effect on the intestinal transcriptome.

Immune transcriptional profiles in mothers with clinically elevated depression and anxiety symptoms several years post-delivery.

Most research on maternal mental health focuses on the perinatal period and does not extend beyond 12 months postpartum. However, emerging evidence suggests that for some women (30%-50%), psychological symptoms may persist beyond the first year postpartum or even emerge later increasing the risk of chronic mood and anxiety symptoms. Despite the high prevalence rates and devastating maternal-child consequences, studies examining maternal depression, anxiety, and post-traumatic stress disorder (PTSD) beyond the first year postpartum are rare and our understanding of the underlying biological mechanisms is incomplete. Inflammatory processes are thought to be involved in the pathophysiology of depression, anxiety, & PTSD outside of the postpartum period. Therefore, the purpose of the current investigation was to examine the relationship between depression, anxiety, and PTSD two to three years post-delivery, and transcriptional control pathways relevant to inflammatory and antiviral processes. Women over 18 years of age enrolled in ongoing research studies at Cedars Sinai Medical Center who were 2-3 years postpartum were invited to participate in the current study. Women (N = 33) reported on their levels of depression, anxiety, and PTSD and provided a blood sample approximately 2-3 years post-delivery. Bioinformatic analyses of differential gene expression (DGEs) to infer transcription factor activity were used. Gene expression was assayed by RNA sequencing and TELiS bioinformatics analysis of transcription factor-binding motifs in the promoters of differentially expressed genes. DGE analyses revealed that women with clinically elevated symptoms of depression, anxiety and PTSD (n = 16) showed upregulation of genes activated by transcription control pathways associated with inflammation (NF-Κ B, p = 0.004; JUN, p = 0.02), including β-adrenergic responsive CREB (p = 0.01) and reduced activation of genes associated with the antiviral response (IRFs, p = 0.02) and the glucocorticoid signaling pathway (GR, p = 0.02) compared to women without clinical symptoms (n = 17). This is one of the first investigations into the immune signaling pathways involved in depression, anxiety, and PTSD two to three years post-delivery. The results of this study suggest that clinically elevated symptoms of depression, anxiety, and PTSD two to three years post-delivery are associated with a gene expression profile characterized by upregulated expression of pro-inflammatory genes and downregulated expression of antiviral genes. The data also point to two potential stress responsive pathways linking symptoms to increased inflammatory signaling in immune cells: sympathetic nervous system mediated β-adrenergic signaling and reduced hypothalamic pituitary adrenal axis activity. Together, these findings highlight the need for investigations into maternal mental health beyond the first year postpartum.

Glutamine transport as a possible regulator of nitrogen catabolite repression in Saccharomyces cerevisiae.

Nitrogen catabolite repression (NCR) is a major transcriptional control pathway governing nitrogen use in yeast, with several hundred of target genes identified to date. Early and extensive studies on NCR led to the identification of the 4 GATA zinc finger transcription factors, but the primary mechanism initiating NCR is still unclear up till now. To identify novel players of NCR, we have undertaken a genetic screen in an NCR-relieved gdh1Δ mutant, which led to the identification of four genes directly linked to protein ubiquitylation. Ubiquitylation is an important way of regulating amino acid transporters and our observations being specifically observed in glutamine-containing media, we hypothesized that glutamine transport could be involved in establishing NCR. Stabilization of Gap1 at the plasma membrane restored NCR in gdh1Δ cells and AGP1 (but not GAP1) deletion could relieve repression in the ubiquitylation mutants isolated during the screen. Altogether, our results suggest that deregulated glutamine transporter function in all three weak nitrogen derepressed (wnd) mutants restores the repression of NCR-sensitive genes consecutive to GDH1 deletion.

Synthesis and Activity of Triazole-Adenosine Analogs as Protein Arginine Methyltransferase 5 Inhibitors.

Protein arginine methyltransferase 5 (PRMT5) is an attractive molecular target in anticancer drug discovery due to its extensive involvement in transcriptional control, RNA processing, and other cellular pathways that are causally related to tumor initiation and progression. In recent years, various compounds have been screened or designed to target either the substrate- or cofactor-binding site of PRMT5. To expand the diversity of chemotypes for inhibitory binding to PRMT5 and other AdoMet-dependent methyltransferases, in this work, we designed a series of triazole-containing adenosine analogs aimed at targeting the cofactor-binding site of PRMT5. Triazole rings have commonly been utilized in drug discovery due to their ease of synthesis and functionalization as bioisosteres of amide bonds. Herein, we utilized the electronic properties of the triazole ring as a novel way to specifically target the cofactor-binding site of PRMT5. A total of about 30 compounds were synthesized using the modular alkyne-azide cycloaddition reaction. Biochemical tests showed that these compounds exhibited inhibitory activity of PRMT5 at varying degrees and several showed single micromolar potency, with clear selectivity for PRMT5 over PRMT1. Docking-based structural analysis showed that the triazole ring plays a key role in binding to the characteristic residue Phe327 in the active pocket of PRMT5, explaining the compounds’ selectivity for this type-II enzyme. Overall, this work provides new structure–activity relationship information on the design of AdoMet analogs for selective inhibition of PRMT5. Further structural optimization work will further improve the potency of the top leads.

RNA metabolism and links to inflammatory regulation and disease.

Inflammation is vital to protect the host against foreign organism invasion and cellular damage. It requires tight and concise gene expression for regulation of pro- and anti-inflammatory gene expression in immune cells. Dysregulated immune responses caused by gene mutations and errors in post-transcriptional regulation can lead to chronic inflammatory diseases and cancer. The mechanisms underlying post-transcriptional gene expression regulation include mRNA splicing, mRNA export, mRNA localisation, mRNA stability, RNA/protein interaction, and post-translational events such as protein stability and modification. The majority of studies to date have focused on transcriptional control pathways. However, post-transcriptional regulation of mRNA in eukaryotes is equally important and related information is lacking. In this review, we will focus on the mechanisms involved in the pre-mRNA splicing events, mRNA surveillance, RNA degradation pathways, disorders or symptoms caused by mutations or errors in post-transcriptional regulation during innate immunity especially toll-like receptor mediated pathways.

Differential Gene Expression in Circulating T-Cells in Long-Term Adolescent/Young Adult Hodgkin Lymphoma (AYAHL) Survivors and Their Unaffected Twins

Background: We and others have demonstrated persistent differences in T cell function and cytokine responses in adolescent/ young adult Hodgkin lymphoma (AYAHL) survivors and their family members. To identify the transcription control pathways involved, we examined mRNA profiles from 17 pairs of long term AYA HL survivors and their unaffected twins.Methods: Blood samples were collected remotely from AYAHL survivors and their like-sex unaffected co-twin controls and shipped to the Epeldegui laboratory at UCLA. B and T cells were negatively separated using magnetic beads. RNA was extracted and sequenced using a high-efficiency mRNA-targeted assay system (Lexogen QuantSeq 3' FWD), with an average 6.7 million reads per sample mapped to the hg38 human transcriptome (99%) using the STAR aligner. Raw read counts for each gene transcript were normalized to transcripts per million total mapped reads and log2 transformed for analysis using linear statistical models including a random effect of twin pair. Analyses examined average expression of pre-specified sets of 17 genes in general immunologic activation (i.e., AP-1- and NF-kB-family transcripts) and 28 Type I IFN-responsive genes (e.g., IFI-, OAS-, and MX-family), as well as TELiS promoter-based bioinformatics analyses of all genes showing >2-fold empirical differences in gene expression (focusing on activation-induced transcription factors, NF-kB and AP-1, and Type 1 interferon-related IRF transcription factors). Body mass index, history of therapeutic radiation, zygosity, sex, age at diagnosis and age at blood collection were included in the model.Results: The average age at diagnosis was 25 years with blood collected 33 years post-diagnosis. In T-cells, AYAHL survivors showed marginally lower expression of 12 activation-induced genes (average 0.41-fold, p = .038) and markedly lower expression of 28 Type 1 interferon-related genes (average 0.33-fold, p < .001) relative to their unaffected co-twins. B cells showed similar reductions in activation-induced genes (0.33-fold, p < .001) but no difference in Type 1 interferon genes (1.02-fold, p =. 942). Promoter-based bioinformatics analyses of all 151 genes found to be differentially expressed by >2-fold in T cells between AYAHL survivors and their unaffected co-twin controls implicated IRF-family transcription factors in mediating observed differences, with IRF3 showing the most significant signal of decreased activity (0.68-fold, p = .059). We also identified marked up-regulation of multiple GATA family transcription factors (GATA3: p < .001; GATA1: p = .009; GATA2: p = .032).Conclusions: Transcriptional indicators of immunologic activation and Type 1 interferon activity show persistent down-regulation in T lymphocytes from long-term adult AYAHL survivors compared to their unaffected like-sex co-twin controls. Incidental findings also identified persistent activation of other gene regulatory pathways mediating T-cell differentiation (e.g., GATA family). We have previously demonstrated higher Epstein-Barr virus (EBV) copy number in AYAHL long-term survivors compared to their unaffected co-twin controls, as well as lower levels of circulating interleukin-12, consistent with a decrease in the ability to control viral replication. The GATA family plays a major role in commitment to T cell lineage (GATA3) and upregulation of the T-helper-type 2 response, and possibly in control of EBV viral infection (GATA2), processes relevant to AYAHL. These differences further support the premise that abnormalities in T-cells play a strong role in etiology, pathogenesis, and survivorship. DisclosuresNo relevant conflicts of interest to declare.

The Transcription Factor SomA Synchronously Regulates Biofilm Formation and Cell Wall Homeostasis in Aspergillus fumigatus.

Polysaccharides are key components of both the fungal cell wall and biofilm matrix. Despite having distinct assembly and regulation pathways, matrix exopolysaccharide and cell wall polysaccharides share common substrates and intermediates in their biosynthetic pathways. It is not clear, however, if the biosynthetic pathways governing the production of these polysaccharides are cooperatively regulated. Here, we demonstrate that cell wall stress promotes production of the exopolysaccharide galactosaminogalactan (GAG)-depend biofilm formation in the major fungal pathogen of humans Aspergillus fumigatus and that the transcription factor SomA plays a crucial role in mediating this process. A core set of SomA target genes were identified by transcriptome sequencing and chromatin immunoprecipitation coupled to sequencing (ChIP-Seq). We identified a novel SomA-binding site in the promoter regions of GAG biosynthetic genes agd3 and ega3, as well as its regulators medA and stuA Strikingly, this SomA-binding site was also found in the upstream regions of genes encoding the cell wall stress sensors, chitin synthases, and β-1,3-glucan synthase. Thus, SomA plays a direct regulation of both GAG and cell wall polysaccharide biosynthesis. Consistent with these findings, SomA is required for the maintenance of normal cell wall architecture and compositions in addition to its function in biofilm development. Moreover, SomA was found to globally regulate glucose uptake and utilization, as well as amino sugar and nucleotide sugar metabolism, which provides precursors for polysaccharide synthesis. Collectively, our work provides insight into fungal adaptive mechanisms in response to cell wall stress where biofilm formation and cell wall homeostasis were synchronously regulated.IMPORTANCE The cell wall is essential for fungal viability and is absent from human hosts; thus, drugs disrupting cell wall biosynthesis have gained more attention. Caspofungin is a member of a new class of clinically approved echinocandin drugs to treat invasive aspergillosis by blocking β-1,3-glucan synthase, thus damaging the fungal cell wall. Here, we demonstrate that caspofungin and other cell wall stressors can induce galactosaminogalactan (GAG)-dependent biofilm formation in the human pathogen Aspergillus fumigatus We further identified SomA as a master transcription factor playing a dual role in both biofilm formation and cell wall homeostasis. SomA plays this dual role by direct binding to a conserved motif upstream of GAG biosynthetic genes and genes involved in cell wall stress sensors, chitin synthases, and β-1,3-glucan synthase. Collectively, these findings reveal a transcriptional control pathway that integrates biofilm formation and cell wall homeostasis and suggest SomA as an attractive target for antifungal drug development.

Translation Regulation by eIF2α Phosphorylation and mTORC1 Signaling Pathways in Non-Communicable Diseases (NCDs).

Non-communicable diseases (NCDs) are medical conditions that, by definition, are non-infectious and non-transmissible among people. Much of current NCDs are generally due to genetic, behavioral, and metabolic risk factors that often include excessive alcohol consumption, smoking, obesity, and untreated elevated blood pressure, and share many common signal transduction pathways. Alterations in cell and physiological signaling and transcriptional control pathways have been well studied in several human NCDs, but these same pathways also regulate expression and function of the protein synthetic machinery and mRNA translation which have been less well investigated. Alterations in expression of specific translation factors, and disruption of canonical mRNA translational regulation, both contribute to the pathology of many NCDs. The two most common pathological alterations that contribute to NCDs discussed in this review will be the regulation of eukaryotic initiation factor 2 (eIF2) by the integrated stress response (ISR) and the mammalian target of rapamycin complex 1 (mTORC1) pathways. Both pathways integrally connect mRNA translation activity to external and internal physiological stimuli. Here, we review the role of ISR control of eIF2 activity and mTORC1 control of cap-mediated mRNA translation in some common NCDs, including Alzheimer’s disease, Parkinson’s disease, stroke, diabetes mellitus, liver cirrhosis, chronic obstructive pulmonary disease (COPD), and cardiac diseases. Our goal is to provide insights that further the understanding as to the important role of translational regulation in the pathogenesis of these diseases.

Relationship closeness buffers the effects of perceived stress on transcriptomic indicators of cellular stress and biological aging marker p16INK4a.

Chronic stress can accelerate biological aging, offering one mechanism through which stress may increase age-related disease risk. Chronic activation of the sympathoadrenal system increases cellular energy production, resulting in cell stress that can initiate cellular senescence, a permanent state of cell growth arrest. Our previous research linked psychosocial stress with increased expression of senescence marker p16INK4a; however, less is known about the role of protective psychosocial factors in biological aging. We examined relationship closeness (perceived interconnectedness with one’s spouse) as a protective buffer of the effects of stress on expression of the p16INK4a-encoding gene (CDKN2A) and transcription control pathways activated under cell stress. Seventy parents (Mage=43.2) completed interview-based and questionnaire measures of psychosocial stress and relationship closeness. Blood samples assessed CDKN2A expression and inferred activity of a priori-selected transcription factors Nrf2 and heat shock factors (HSFs) via genome-wide transcriptome profiling. Random intercept models adjusting for age, sex, and ethnicity/race revealed that perceived stress was associated with elevated CDKN2A expression for parents with low but not high closeness. Secondary bioinformatics analyses linked the interaction of perceived stress and relationship closeness to Nrf2 and HSF-1 activity. Findings identify relationship closeness as a protective factor that may buffer the impact of stress on cellular stress and senescence pathways.

Perioperative COX2 andβ-adrenergic blockade improves biomarkers of tumor metastasis, immunity, and inflammation in colorectal cancer: A randomized controlled trial.

Preclinical studies have implicated excess release of catecholamines and prostaglandins in the mediation of prometastatic processes during surgical treatment of cancer. In this study, we tested the combined perioperative blockade of these pathways in patients with colorectal cancer (CRC). In a randomized, double-blind, placebo-controlled biomarker trial involving 34 patients, the β-blocker propranolol and the COX2-inhibitor etodolac were administered for 20 perioperative days, starting 5days before surgery. Excised tumors were subjected to whole genome messenger RNA profiling and transcriptional control pathway analyses. Drugs were well-tolerated, with minor complications in both the treatment group and the placebo group. Treatment resulted in a significant improvement (P<.05) of tumor molecular markers of malignant and metastatic potential, including 1) reduced epithelial-to-mesenchymal transition, 2) reduced tumor infiltrating CD14+ monocytes and CD19+ B cells, and 3) increased tumor infiltrating CD56+ natural killer cells. Transcriptional activity analyses indicated a favorable drug impact on 12 of 19 a priori hypothesized CRC-related transcription factors, including the GATA, STAT, and EGR families as well as the CREB family that mediates the gene regulatory impact of β-adrenergic- and prostaglandin-signaling. Alterations observed in these transcriptional activities were previously associated with improved long-term clinical outcomes. Three-year recurrence rates were assessed for long-term safety analyses. An intent-to-treat analysis revealed that recurrence rates were 12.5% (2/16) in the treatment group and 33.3% (6/18) in the placebo group (P=.239), and in protocol-compliant patients, recurrence rates were 0% (0/11) in the treatment group and 29.4% (5/17) in the placebo group (P=.054). The favorable biomarker impacts and clinical outcomes provide a rationale for future randomized placebo-controlled trials in larger samples to assess the effects of perioperative propranolol/etodolac treatment on oncological clinical outcomes.

Profiles of gene expression in maternal blood predict offspring birth weight in normal pregnancy.

The association between lower birth weight and increased disease risk in adulthood has drawn attention to the physiological processes that shape the gestational environment. We implement genome-wide transcriptional profiling of maternal blood samples to identify subsets of genes and associated transcription control pathways that predict offspring birth weight. Female participants (N = 178, mean = 27.0 years) in a prospective observational birth cohort study were contacted between 2009 and 2014 to identify new pregnancies. An in-home interview was scheduled for early in the third trimester (mean = 30.3 weeks) to collect pregnancy-related information and a blood sample, and birth weight was measured shortly after delivery. Transcriptional activity in white blood cells was determined with a whole-genome gene expression direct hybridization assay. Fifty transcripts were differentially expressed in association with offspring birth weight, with 18 up-regulated in relation to lower birth weight, and 32 down-regulated. Examination of transcription control pathways identified increased activity of NF-κB, AP-1, EGR1, EGR4, and Gfi families, and reduced the activity of CEBP, in association with lower birth weight. Transcript origin analyses identified non-classical CD16+ monocytes, CD1c+ myeloid dendritic cells, and neutrophils as the primary cellular mediators of differential gene expression. These results point toward a systematic regulatory shift in maternal white blood cell activity in association with lower offspring birth weight, and they suggest that analyses of gene expression during gestation may provide insight into regulatory and cellular mechanisms that influence birth outcomes.

Experienced discrimination and racial differences in leukocyte gene expression

Racial disparities in health outcomes between African Americans and European Americans have been well-documented, but not fully understood. Chronic inflammation contributes to several of the diseases showing racial disparities (e.g., Human Immunodeficiency Virus [HIV]), and racial differences in stress exposure (e.g., experiences of racial discrimination) that stimulate pro-inflammatory processes that may contribute to differential health outcomes.We performed a cross-sectional bioinformatic analyses relating perceived discrimination (as measured by the Perceived Ethnic Discrimination Questionnaire [PED-Q]) to the activity of pro-inflammatory, neuroendocrine, and antiviral transcription control pathways relevant to the conserved transcriptional response to adversity (CTRA) in peripheral blood leukocytes. Subjects were 71 individuals (37 HIV-seropositive (HIV+); 34 HIV-seronegative (HIV-)) (mean age = 53 years, range 27–63), who self-identified either as African American/Black (n = 48) or European American/White (n = 23). This provided the opportunity to examine the independent effects of race and HIV, as well as the modifying role of perceived discrimination on pathways involved in CTRA. Exploratory analysis examined the interactive effects of HIV and race on pathways involved in CTRA. Relative to European Americans, African Americans showed increased activity of two key pro-inflammatory transcription control pathways (NF- кB and AP-1) and two stress-responsive signaling pathways (CREB and glucocorticoid receptor); these effects did not differ significantly as a function of HIV infection (HIV x Race interaction, all p > .10). Results suggested that differences in experiences of racial discrimination could potentially account for more than 50% of the total race-related difference in pro-inflammatory transcription factor activity. In sum, differential exposure to racial discrimination may contribute to racial disparities in health outcomes in part by activating threat-related molecular programs that stimulate inflammation and contribute to increased risk of chronic illnesses.

Moderators for depressed mood and systemic and transcriptional inflammatory responses: a randomized controlled trial of endotoxin.

Activation of the innate immune system is thought to contribute to depression. Multiple social and behavioral factors are also known to instigate depression. Whether these socio-behavioral factors interact with inflammatory stimuli to alter proinflammatory responses and depressed mood is not known. In 115 healthy adults, social, emotional, and behavioral factors were assessed at baseline. A single infusion of endotoxin (Escherichia coli; 0.8 ng/kg of body weight) or placebo (0.9% saline) was administered with hourly assessment of depressed mood and proinflammatory cytokines (interleukin-6 (IL-6); tumor necrosis factor-α (TNF)). Inflammatory gene expression was examined at 30 min after infusion, prior to increase of inflammatory cytokines. As compared to placebo, endotoxin-induced increases of depressed mood were moderated by baseline levels of perceived stress, trait sensitivity to social disconnection, and severity of symptoms of anxiety and depression (all Ps < 0.05) but not early life stress, social status, social support, neuroticism, or sleep disturbance. Anxiety symptoms remained significant in multivariable analyses (P < 0.01). None of these socio-behavioral factors were related to increases in proinflammatory cytokines. Transcriptome profiling analyses indicated that perceived stress, sensitivity to social disconnection, and depressive symptoms were associated with increased activation of pro-inflammatory transcription control pathways (i.e., activator protein-1, nuclear factor-κB) in response to endotoxin (all Ps < 0.05). These results indicate that an array of socio-behavioral factors, which are associated with depression risk, modify vulnerability to inflammation-induced depressed mood. Together, these observations may be used to help target therapeutic interventions to mitigate occurrence of the inflammatory biotype of depression.

Differential regulation of NF-kB and IRF target genes as they relate to fatigue in patients with head and neck cancer

Previous studies have linked plasma inflammatory markers to elevated fatigue in patients with head and neck cancer (HNC). To identify the molecular mechanisms underlying this association, we conducted promoter-based bioinformatics analyses to determine the relationship between fatigue and specific gene expression profiles associated with inflammation in human papillomavirus (HPV)-related and -unrelated HNC patients undergoing treatment. Patients with newly diagnosed HNC without distant metastasis were assessed at baseline (pre-radiotherapy) and one-month post-radiotherapy. Fatigue was measured by the Multidimensional Fatigue Inventory. Genome-wide gene expression profiles were collected from peripheral blood mononuclear cells (PBMC). Promoter-based bioinformatics analyses were employed to identify transcription control pathways underlying transcriptomic correlates of fatigue in the sample as a whole and in HPV-related and HPV-unrelated HNC patients separately. In transcriptome profiling analyses of PBMC from 44 patients, TELiS bioinformatics analyses linked fatigue to increased nuclear factor-kappa B (NF-kB) transcriptional activity and decreased interferon regulatory factor family (IRF) transcription factor activity. Patients with HPV-related HNC showed lower levels of fatigue-related gene expression profile compared to HPV-unrelated HNC. Fatigue in HNC patients undergoing treatment is associated with gene expression profiles consistent with the conserved transcriptional response to adversity (CTRA) characterized by increased proinflammatory and decreased anti-antiviral transcriptional activity. Interestingly, this CTRA response was mitigated in patients with HPV-related HNC and may explain the lower level of fatigue they experience relative to HPV-unrelated HNC.

PO-024 Perioperative blockade of stress-inflammatory responses improves biomarkers of metastasis in excised tumours and blood samples: phase ii clinical trials in colorectal and breast cancer patients

IntroductionExcess release of catecholamines and prostaglandins has been shown to mediate pro-metastatic processes of stress and surgery, specifically during the critical perioperative period.Material and methodsHere, in two randomised placebo controlled clinical trials in colorectal (CRC, n=34) and in breast cancer patients (BC, n=38), we tested the combined 20 day perioperative use of the b-blocker, propranolol, and the COX2-inhibitor, etodolac, initiated 5 days before surgery. Tumour samples were collected during surgery, and were subjected to histological analyses, whole genome mRNA profiling, and transcriptional control pathways analyses. In BC patients, four blood samples were also collected perioperatively.Results and discussionsDrugs were well tolerated, with adverse effects equivalent to placebo. In blood samples, treatment reduced serum IL-6 and CRP levels even before surgery, improved markers of NK cytotoxicity, and enhanced induced production of IFNg and IL-12, without affecting anti-inflammatory soluble factors (cortisol and IL-10). In both studies, whole genome mRNA profiling of excised tumours showed decreased epithelial-to-mesenchymal transition (EMT); down-regulation of the transcriptional activity of CREB, NFkB, GATA family, and STAT3; reduced presence of tumor-associated monocytes; and increased presence of NK cells in CRC and B cells in BC tissue. The tumour proliferation marker Ki67 was tested in BC patients, and was significantly reduced by drug treatment. In CRC patients, three-year follow-up showed large but statistically insignificant improvement in disease free survival (DFS) in the treatment group (1/15 vs 5/19), further suggesting the safety of the paradigm.ConclusionThese findings suggest a critical impact to the short pre-operative period, clearly indicate the efficacy of this combined drug regimen, and suggest its metastatic-reducing impact, which should be tested in larger clinical trials. Such a stress-inflammatory-reducing approach can be exploited during the critical perioperative period, potentially improving long-term survival rates.

Dark chocolate (70% cacao) effects human gene expression: Cacao regulates cellular immune response, neural signaling, and sensory perception

Dark chocolate (70% Cacao) is a major source of flavonoid antioxidants, which appear to support cognitive, endocrine, and cardiovascular health benefits. However, the in vivo effects of cacao on human genome function are unknown. In a pilot feasibility experimental trial, we examined the effects acute and chronic cacao consumption on human gene expression in circulating blood cells.PurposeMap the impact of 70% cacao consumption on human immune and dendritic cell gene expression, with focus on pro/anti‐inflammatory cytokines.MethodsThis 8‐day (d) pilot study enrolled 4 healthy experimental subjects (2 female and 2 male, age 25–50) and one control (female, 28), who were not taking prescribed or OTC medications. Subjects abstained from all other high‐antioxidant foods or supplements 48 h prior to and during the study period. After a fasting baseline blood sample, experimental subjects consumed 48 g 70% cacao (Parliament Chocolates, Redlands, CA) over 10 min (antioxidant activity 46,000 μmoles TE/100g). Follow‐up blood samples were obtained 2 h later (acute effects) and 1 wk later (chronic effects). During waking hours (14 h/d) experimental subjects consumed 48 g 70% cacao per day (2 pieces/hr; antioxidant activity 3834 μmoles TE/100g). Total RNA was assayed by RNAseq, with data analyzed by linear models to identify differentially expressed genes, followed by bioinformatic analyses of transcription control pathways and cellular origins.ResultsLeukocyte transcriptome profiles showed little systematic change in the initial 2 h of Cacao exposure, although pre‐specified bioinformatics analyses indicated small increases in activity of genes regulated by NF‐kB and AP1 transcription factors and CD4+ and CD8+ T cells (all, p &lt; .05). By contrast, analyses of change from baseline to 1 wk identified 177 genes showing significant differential expression using a priori‐specified False Discovery Rate cut‐point of .20 (140 up‐regulated, 37 down‐regulated), with 116 achieving q &lt; .05. Up‐regulated genes were involved in leukocyte activation and motility (e.g., CD40, TICAM2, AZU1, HRAS and RASD2, HSPA1L, FGF22); derived from CD4+ and CD8+ T lymphocytes (p &lt; .001 and .0148, respectively); and showed over‐representation of promoter elements for NF‐kB, SP‐1, MZF‐1, and HIF‐1 transcription factors (all p &lt; .01) and under‐representation of AP‐1 (p = .018). Up‐regulated transcripts also included genes involved in neural signaling and sensory perception (e.g., OR2J3, OR52A5, OR6C3, OR9I1, NR4A2, and NRXN2).ConclusionCacao consumption up‐regulates multiple intracellular signaling pathways involved in T cell activation and cellular immune response. These effects emerged weakly within 2 h but there was greater gene expression by the end of 1 wk intervention. Results are consistent with previous indications that cacao components inhibit MAPK activation, but suggest T cell function may be preserved and enhanced via other compensatory molecular pathways. It should be noted that the NF‐kB up‐regulation source was not from up‐regulated monocytes suggesting a role other than cascading inflammation. It is of interest, to note, that genes involved in neural signaling and sensory processes were up‐regulated. The functional significance of these effects for immune cells remains to be defined by future expansive research, as do the neurobiological mechanisms involved.Support or Funding InformationDepartmental Cacao and gene expressionThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Perioperative beta-adrenergic blockade and COX2 inhibition in colorectal cancer patients improves pro-metastatic indices in the excised tumor: EMT, tumor infiltrating lymphocytes (TILs), and gene regulatory pathways

Excess release of catecholamines and prostaglandins was shown to mediate pro-metastatic processes during the perioperative period. Here, in a randomized controlled biomarker clinical trial in colorectal cancer patients (CRCP) (n = 34), we tested the combined perioperative use of the β-blocker, propranolol, and the COX2-inhibitor, etodolac, scheduled for 20 perioperative days, starting 5 days before surgery. Tumor samples were subjected to whole genome mRNA profiling and transcriptional control pathways analyses. Recurrence-free survival (RFS) is currently monitored. Drugs were well tolerated. Whole genome mRNA profiling indicated that drug treatment significantly reduced (i) markers of epithelial-to-mesenchymal- transition (EMT), (ii) tumor infiltrating CD14 + monocytes, and (iii) tumor infiltrating CD19 + B-cells, which are associated with tumor progression and metastatic disease in CRCP. Drug treatment increased tumor infiltrating CD56 + NK-cells, potentially improving anti-metastatic immunity, as the majority of colorectal tumors are susceptible to NK cytotoxicity. Lastly, drug treatment reduced the indicated activity of several pro-metastatic transcription factors (GATA1/GATA2, IRF1/ISRE, OCT1, c-MYB, HSF1, CREB), but increased the potentially tumor-promoting STAT1 activity. Two-year recurrence rates are 7% (1/15 patients) in the drug-treated group and 27% (5/19 patients) in the placebo group (p=0.35 – underpowered study for this index). Overall, CRCP undergoing curative surgery may benefit from the combined drug treatment through reduction of stress-mediated pro-metastatic pathways in the primary tumor and potentially residual disease. Long-term cancer outcomes should be studied to test clinical ramifications.