Visual communication design: a neglected factor in nutrition promotion via packaged food labels.

Packaging design is a communication device and a critical component in branding strategy, and has relevance for food policy. Presently, packaging-related nutrition policy initiatives focus on the role of regulated claims, nutrition information panels and front-of-pack nutrition labels to help guide consumer food choices and address high prevalences of discretionary and ultra-processed food consumption in many countries. However, these nutrition labelling systems are not optimized as public health policy tools as many consumers do not use them to inform their food choices. Visual communication design theory posits that a designer orders the elements and principles of design into hierarchies that prioritize certain elements over others, and that some of these elements are more dominant and given more emphasis than others. The overall design of the package thereby directs consumer attention to some aspects of pack design (e.g., characters, contents of the package) and away from others (e.g., nutrition details). Dual processing frameworks propose that food decisions are made with the interplay between automatic and rational thinking processes. Packaging designs affect whether consumers rely predominantly on automatic or rational thinking to select a food. This narrative review outlines the role of food packaging design and how it impacts the clear communication of nutrition aspects of food products and how the use of nutrition information by consumers to make decisions may depend upon design structures in packaging. This article attests that nutrition scientists and policy makers should incorporate visual communication design into research on the food packaging as a public health promotion tool. A stronger focus on the communication of regulated front-of-pack nutrition information can be made with a re-evaluation of the hierarchy of elements in the front-of-pack design enabling consumers to make healthier decisions.

Australian Stakeholder Perspectives on Healthier Retail Food Environments for Toddlers-The Era of "Band Aids and Small Inroads".

Background: A healthy diet in young children is crucial for optimal growth and development. However, many toddlers (1-3 y) consume suboptimal diets, and as a result, are at a high risk of experiencing negative health outcomes. Moreover, minimal progress has been made to improve the healthiness of retail food environments for toddlers to date despite the potential and advocacy for the issue. Objectives: To gain insight into stakeholder perceptions and opinions on the healthiness of Australian retail food environments for toddlers, as well as perspectives on the options and barriers to improve their healthiness. Methods: Qualitative, online study utilizing semi-structured individual interviews with 27 key stakeholders from food industry, academia, nongovernment organizations, public health, and government in Australia. Results: Most stakeholders agreed that retail food environments for toddlers were not health promoting. Stakeholders identified that a multifaceted approach including nutrition education and strong government mandated regulation were essential to improve the healthiness of retail food environments for toddlers. Interviews also highlighted the main perceived barriers to progress, and reasons for policy inaction in this area are the food industry and government support. Many stakeholders were concerned that child health is being undermined due to the government favoring business needs over public health. Conclusions: Stakeholders in this study overwhelmingly agreed that there is more that can and should be done to create health promoting retail food environments for toddlers in Australia. Stakeholders identified a range of strategies that can be used to improve the healthiness of toddler food environments, but advocacy efforts are being undermined due to government inaction. Stakeholders believed that strong governance is required to create equitable, sustainable healthy retail food environments for young children. Improving the healthiness of retail food environments for young children will not only reduce diet related disease across the lifespan but will help to address financial and societal costs of a poor diet.

APOBEC3B regulates R-loops and promotes transcription-associated mutagenesis in cancer.

The single-stranded DNA cytosine-to-uracil deaminase APOBEC3B is an antiviral protein implicated in cancer. However, its substrates in cells are not fully delineated. Here APOBEC3B proteomics reveal interactions with a surprising number of R-loop factors. Biochemical experiments show APOBEC3B binding to R-loops in cells and in vitro. Genetic experiments demonstrate R-loop increases in cells lacking APOBEC3B and decreases in cells overexpressing APOBEC3B. Genome-wide analyses show major changes in the overall landscape of physiological and stimulus-induced R-loops with thousands of differentially altered regions, as well as binding of APOBEC3B to many of these sites. APOBEC3 mutagenesis impacts genes overexpressed in tumors and splice factor mutant tumors preferentially, and APOBEC3-attributed kataegis are enriched in RTCW motifs consistent with APOBEC3B deamination. Taken together with the fact that APOBEC3B binds single-stranded DNA and RNA and preferentially deaminates DNA, these results support a mechanism in which APOBEC3B regulates R-loops and contributes to R-loop mutagenesis in cancer.

The big squeeze: a product content and labelling analysis of ready-to-use complementary infant food pouches in Australia.

BACKGROUND: Encouraging the early development of healthy eating habits prevents diet-related chronic disease. It is well understood that highly processed foods with high amounts of sugars, salt and fats are a risk factor for non-communicable diseases. Commercial baby foods in ready-to-use squeeze pouches emerged in the global food market around 2012. The long-term effects of this now ubiquitous packaging on the quality of infant diets, baby food consumption and marketing are unknown. This study aimed to conduct a rigorous mixed-methods audit of squeeze pouches in Australia to inform product regulation and policy. METHODS: Nutritional and marketing data were sourced from products available in Australian retailers. Analysis of nutritional content, texture and packaging labelling and serving size was conducted. Pouches were given a Nutrition Profile Index (NPI) score and compared with the Australian Infant Feeding Guidelines. Marketing text was thematically analysed and compared to existing infant nutrition policy around regulation of marketing claims. RESULTS: 276 products from 15 manufacturers were analysed, targeting infants from 4 + to 12 + months. Total sugar content ranged 0.8-17.5 g/100 g, 20% (n = 56) of products had added sugars, 17% (n = 46) had added fruit juice, 71% (n = 196) had added fruit puree. Saturated fat content ranged from 0.0 to 5.0 g/100 g, sodium 0.0-69 mg/100 g and dietary fibre 0.0-4.3 g/100 g. Only two products were nutritionally adequate according to a nutrient profiling tool. Marketing messages included ingredient premiumisation, nutrient absence claims, claims about infant development and health, good parenting, and convenience. Claims of 'no added sugar' were made for 59% of pouches, despite the addition of free sugars. CONCLUSIONS: Squeeze pouch products available in Australia are nutritionally poor, high in sugars, not fortified with iron, and there is a clear risk of harm tothe health of infant and young children if these products are fed regularly. The marketing messages and labelling on squeeze pouches are misleading and do not support WHO or Australian NHMRC recommendations for breastfeeding or appropriate introduction of complementary foods and labelling of products. There is an urgent need for improved regulation of product composition, serving sizes and labelling to protect infants and young children aged 0-36 months and better inform parents.

PARP inhibition and pharmacological ascorbate demonstrate synergy in castration-resistant prostate cancer.

Prostate cancer (PCa) is the second leading cause of cancer death for men in the United States. While organ-confined disease has reasonable expectation of cure, metastatic PCa is universally fatal upon recurrence during hormone therapy, a stage termed castration-resistant prostate cancer (CRPC). Until such time as molecularly defined subtypes can be identified and targeted using precision medicine, it is necessary to investigate new therapies that may apply to the CRPC population as a whole. The administration of ascorbate, more commonly known as ascorbic acid or Vitamin C, has proved lethal to and highly selective for a variety of cancer cell types. There are several mechanisms currently under investigation to explain how ascorbate exerts anti-cancer effects. A simplified model depicts ascorbate as a pro-drug for reactive oxygen species (ROS), which accumulate intracellularly and generate DNA damage. It was therefore hypothesized that poly(ADP-ribose) polymerase (PARP) inhibitors, by inhibiting DNA damage repair, would augment the toxicity of ascorbate. Results: Two distinct CRPC models were found to be sensitive to physiologically relevant doses of ascorbate. Moreover, additional studies indicate that ascorbate inhibits CRPC growth in vitro via multiple mechanisms including disruption of cellular energy dynamics and accumulation of DNA damage. Combination studies were performed in CRPC models with ascorbate in conjunction with escalating doses of three different PARP inhibitors (niraparib, olaparib, and talazoparib). The addition of ascorbate augmented the toxicity of all three PARP inhibitors and proved synergistic with olaparib in both CRPC models. Finally, the combination of olaparib and ascorbate was tested in vivo in both castrated and non-castrated models. In both cohorts, the combination treatment significantly delayed tumor growth compared to monotherapy or untreated control. Conclusions: These data indicate that pharmacological ascorbate is an effective monotherapy at physiological concentrations and kills CRPC cells. Ascorbate-induced tumor cell death was associated with disruption of cellular energy dynamics and accumulation of DNA damage. The addition of PARP inhibition increased the extent of DNA damage and proved effective at slowing CRPC growth both in vitro and in vivo. These findings nominate ascorbate and PARPi as a novel therapeutic regimen that has the potential to improve CRPC patient outcomes.

Participation of ATM, SMG1, and DDX5 in a DNA Damage-Induced Alternative Splicing Pathway.

Altered cellular responses to DNA damage can contribute to cancer development, progression, and therapeutic resistance. Mutations in key DNA damage response factors occur across many cancer types, and the DNA damage-responsive gene, TP53, is frequently mutated in a high percentage of cancers. We recently reported that an alternative splicing pathway induced by DNA damage regulates alternative splicing of TP53 RNA and further modulates cellular stress responses. Through damage-induced inhibition of the SMG1 kinase, TP53 pre-mRNA is alternatively spliced to generate TP53b mRNA and p53b protein is required for optimal induction of cellular senescence after ionizing radiation-induced DNA damage. Herein, we confirmed and extended these observations by demonstrating that the ATM protein kinase is required for repression of SMG1 kinase activity after ionizing radiation. We found that the RNA helicase and splicing factor, DDX5, interacts with SMG1, is required for alternative splicing of TP53 pre-mRNA to TP53b and TP53c mRNAs after DNA damage, and contributes to radiation-induced cellular senescence. Interestingly, the role of SMG1 in alternative splicing of p53 appears to be distinguishable from its role in regulating nonsense-mediated RNA decay. Thus, ATM, SMG1, and DDX5 participate in a DNA damage-induced alternative splicing pathway that regulates TP53 splicing and modulates radiation-induced cellular senescence.

Regulated nutrition claims increase perceived healthiness of an ultra-processed, discretionary toddler snack food and ultra-processed toddler milks: A discrete choice experiment.

There has been a prolonged increase in the sale and consumption of ultra-processed, discretionary foods and ultra-processed milks for toddlers, which display numerous on-pack claims that influence health perceptions. This study investigated the relative impact of different regulated and unregulated claims on parent perceptions of the healthiness of a toddler snack food and milk in Australia. Participants aged 18+ years completed an online survey, including discrete choice experiments for an ultra-processed, discretionary toddler snack food and an ultra-processed toddler milk, which displayed combinations of claims across nutrition, health, and other domains. Participants were asked to choose the 'most and least healthy' products between three alternatives over seven choice sets. Data were analysed using an ordinal logistic regression model. Likelihood-ratio tests revealed the most important contribution was variation in regulated nutrition-content claims. For the toddler snack, participants were nearly 14 times more likely to perceive a product with the regulated nutrition-content claim "no added sugar, no added salt" as most healthy (OR 13.71, p < 0.001), compared to when no regulated nutrition-content claim was present. For the toddler milk, participants were more than two and a half times more likely to choose a product that contained the regulated nutrition-content claim "2 serves = up to 50% of RDI recommended dietary intake of 14 vitamins and minerals" as most healthy (OR 2.65, p < 0.001) compared to when no regulated nutrition-content claim was present. In Australia, regulated nutrition-content claims can be displayed on packaged foods regardless of healthiness. These results indicate that such claims increase perceptions of healthiness of ultra-processed, discretionary toddler snack foods and ultra-processed toddler milks. Further controls are required to regulate the use of nutrition-content and health claims to facilitate informed consumer choice.

Toddler foods and milks don't stack up against regular foods and milks.

AIM: To compare the cost and nutritional profiles of toddler-specific foods and milks to 'regular' foods and milks. METHODS: Cross-sectional audit of non-toddler specific ('regular') foods and milks and secondary analysis of existing audit data of toddler specific (12-36 months) foods and milks in Australia. MAIN FINDINGS: The cost of all toddler-specific foods and milks was higher than the regular non-toddler foods. Foods varied in nutritional content, but toddler foods were mostly of poorer nutritional profile than regular foods. Fresh milk cost, on average, $0.22 less per 100 mL than toddler milk. Toddler milks had higher mean sugar and carbohydrate levels and lower mean protein, fat, saturated fat, sodium and calcium levels per 100 mL, when compared to fresh full fat cow's milk. CONCLUSIONS: Toddler specific foods and milks cost more and do not represent value for money or good nutrition for young children.

A Novel Role for DNA-PK in Metabolism by Regulating Glycolysis in Castration-Resistant Prostate Cancer.

PURPOSE: DNA-dependent protein kinase catalytic subunit (DNA-PKcs, herein referred as DNA-PK) is a multifunctional kinase of high cancer relevance. DNA-PK is deregulated in multiple tumor types, including prostate cancer, and is associated with poor outcomes. DNA-PK was previously nominated as a therapeutic target and DNA-PK inhibitors are currently undergoing clinical investigation. Although DNA-PK is well studied in DNA repair and transcriptional regulation, much remains to be understood about the way by which DNA-PK drives aggressive disease phenotypes. EXPERIMENTAL DESIGN: Here, unbiased proteomic and metabolomic approaches in clinically relevant tumor models uncovered a novel role of DNA-PK in metabolic regulation of cancer progression. DNA-PK regulation of metabolism was interrogated using pharmacologic and genetic perturbation using in vitro cell models, in vivo xenografts, and ex vivo in patient-derived explants (PDE). RESULTS: Key findings reveal: (i) the first-in-field DNA-PK protein interactome; (ii) numerous DNA-PK novel partners involved in glycolysis; (iii) DNA-PK interacts with, phosphorylates (in vitro), and increases the enzymatic activity of glycolytic enzymes ALDOA and PKM2; (iv) DNA-PK drives synthesis of glucose-derived pyruvate and lactate; (v) DNA-PK regulates glycolysis in vitro, in vivo, and ex vivo; and (vi) combination of DNA-PK inhibitor with glycolytic inhibitor 2-deoxyglucose leads to additive anti-proliferative effects in aggressive disease. CONCLUSIONS: Findings herein unveil novel DNA-PK partners, substrates, and function in prostate cancer. DNA-PK impacts glycolysis through direct interaction with glycolytic enzymes and modulation of enzymatic activity. These events support energy production that may contribute to generation and/or maintenance of DNA-PK-mediated aggressive disease phenotypes.

Mutant p53 elicits context-dependent pro-tumorigenic phenotypes.

The tumor suppressor gene TP53 is the most frequently mutated gene in numerous cancer types, including prostate cancer (PCa). Specifically, missense mutations in TP53 are selectively enriched in PCa, and cluster to particular "hot spots" in the p53 DNA binding domain with mutation at the R273 residue occurring most frequently. While this residue is similarly mutated to R273C-p53 or R273H-p53 in all cancer types examined, in PCa selective enrichment of R273C-p53 is observed. Importantly, examination of clinical datasets indicated that TP53 heterozygosity can either be maintained or loss of heterozygosity (LOH) occurs. Thus, to mimic tumor-associated mutant p53, R273C-p53 and R273H-p53 isogenic PCa models were developed in the presence or absence of wild-type p53. In the absence of wild-type p53, both R273C-p53 and R273H-p53 exhibited similar loss of DNA binding, transcriptional profiles, and loss of canonical tumor suppressor functions associated with wild-type p53. In the presence of wild-type p53 expression, both R273C-p53 and R273H-p53 supported canonical p53 target gene expression yet elicited distinct cistromic and transcriptional profiles when compared to each other. Moreover, heterozygous modeling of R273C-p53 or R273H-p53 expression resulted in distinct phenotypic outcomes in vitro and in vivo. Thus, mutant p53 acts in a context-dependent manner to elicit pro-tumorigenic transcriptional profiles, providing critical insight into mutant p53-mediated prostate cancer progression.

Novel Oncogenic Transcription Factor Cooperation in RB-Deficient Cancer.

The retinoblastoma tumor suppressor (RB) is a critical regulator of E2F-dependent transcription, controlling a multitude of protumorigenic networks including but not limited to cell-cycle control. Here, genome-wide assessment of E2F1 function after RB loss in isogenic models of prostate cancer revealed unexpected repositioning and cooperation with oncogenic transcription factors, including the major driver of disease progression, the androgen receptor (AR). Further investigation revealed that observed AR/E2F1 cooperation elicited novel transcriptional networks that promote cancer phenotypes, especially as related to evasion of cell death. These observations were reflected in assessment of human disease, indicating the clinical relevance of the AR/E2F1 cooperome in prostate cancer. Together, these studies reveal new mechanisms by which RB loss induces cancer progression and highlight the importance of understanding the targets of E2F1 function. SIGNIFICANCE: This study identifies that RB loss in prostate cancer drives cooperation between AR and E2F1 as coregulators of transcription, which is linked to the progression of advanced disease.

RB/E2F1 as a Master Regulator of Cancer Cell Metabolism in Advanced Disease.

Loss of the retinoblastoma (RB) tumor suppressor protein is a critical step in reprogramming biological networks that drive cancer progression, although mechanistic insight has been largely limited to the impact of RB loss on cell-cycle regulation. Here, isogenic modeling of RB loss identified disease stage-specific rewiring of E2F1 function, providing the first-in-field mapping of the E2F1 cistrome and transcriptome after RB loss across disease progression. Biochemical and functional assessment using both in vitro and in vivo models identified an unexpected, prominent role for E2F1 in regulation of redox metabolism after RB loss, driving an increase in the synthesis of the antioxidant glutathione, specific to advanced disease. These E2F1-dependent events resulted in protection from reactive oxygen species in response to therapeutic intervention. On balance, these findings reveal novel pathways through which RB loss promotes cancer progression and highlight potentially new nodes of intervention for treating RB-deficient cancers. SIGNIFICANCE: This study identifies stage-specific consequences of RB loss across cancer progression that have a direct impact on tumor response to clinically utilized therapeutics. The study herein is the first to investigate the effect of RB loss on global metabolic regulation and link RB/E2F1 to redox control in multiple advanced diseases.This article is highlighted in the In This Issue feature, p. 2113.

Structural basis for recognition of distinct deaminated DNA lesions by endonuclease Q.

Spontaneous deamination of DNA cytosine and adenine into uracil and hypoxanthine, respectively, causes C to T and A to G transition mutations if left unrepaired. Endonuclease Q (EndoQ) initiates the repair of these premutagenic DNA lesions in prokaryotes by cleaving the phosphodiester backbone 5' of either uracil or hypoxanthine bases or an apurinic/apyrimidinic (AP) lesion generated by the excision of these damaged bases. To understand how EndoQ achieves selectivity toward these structurally diverse substrates without cleaving undamaged DNA, we determined the crystal structures of Pyrococcus furiosus EndoQ bound to DNA substrates containing uracil, hypoxanthine, or an AP lesion. The structures show that substrate engagement by EndoQ depends both on a highly distorted conformation of the DNA backbone, in which the target nucleotide is extruded out of the helix, and direct hydrogen bonds with the deaminated bases. A concerted swing motion of the zinc-binding and C-terminal helical domains of EndoQ toward its catalytic domain allows the enzyme to clamp down on a sharply bent DNA substrate, shaping a deep active-site pocket that accommodates the extruded deaminated base. Within this pocket, uracil and hypoxanthine bases interact with distinct sets of amino acid residues, with positioning mediated by an essential magnesium ion. The EndoQ-DNA complex structures reveal a unique mode of damaged DNA recognition and provide mechanistic insights into the initial step of DNA damage repair by the alternative excision repair pathway. Furthermore, we demonstrate that the unique activity of EndoQ is useful for studying DNA deamination and repair in mammalian systems.

The circadian cryptochrome, CRY1, is a pro-tumorigenic factor that rhythmically modulates DNA repair.

Mechanisms regulating DNA repair processes remain incompletely defined. Here, the circadian factor CRY1, an evolutionally conserved transcriptional coregulator, is identified as a tumor specific regulator of DNA repair. Key findings demonstrate that CRY1 expression is androgen-responsive and associates with poor outcome in prostate cancer. Functional studies and first-in-field mapping of the CRY1 cistrome and transcriptome reveal that CRY1 regulates DNA repair and the G2/M transition. DNA damage stabilizes CRY1 in cancer (in vitro, in vivo, and human tumors ex vivo), which proves critical for efficient DNA repair. Further mechanistic investigation shows that stabilized CRY1 temporally regulates expression of genes required for homologous recombination. Collectively, these findings reveal that CRY1 is hormone-induced in tumors, is further stabilized by genomic insult, and promotes DNA repair and cell survival through temporal transcriptional regulation. These studies identify the circadian factor CRY1 as pro-tumorigenic and nominate CRY1 as a new therapeutic target.

DNA-Damage-Induced Alternative Splicing of p53.

Cellular responses to DNA damage and other stresses are important determinants of mutagenesis and impact the development of a wide range of human diseases. TP53 is highly mutated in human cancers and plays an essential role in stress responses and cell fate determination. A central dogma of p53 induction after DNA damage has been that the induction results from a transient increase in the half-life of the p53 protein. Our laboratory recently demonstrated that this long-standing paradigm is an incomplete picture of p53 regulation by uncovering a critical role for protein translational regulation in p53 induction after DNA damage. These investigations led to the discovery of a DNA-damage-induced alternative splicing (AS) pathway that affects p53 and other gene products. The damage-induced AS of p53 pre-mRNA generates the beta isoform of p53 (p53ß) RNA and protein, which is specifically required for the induction of cellular senescence markers after ionizing irradiation (IR). In an attempt to elucidate the mechanisms behind the differential regulation and apparent functional divergence between full-length (FL) p53 and the p53ß isoform (apoptosis versus senescence, respectively), we identified the differential transcriptome and protein interactome between these two proteins that may result from the unique 10-amino-acid tail in p53ß protein.

The Nutritional Profile and On-Pack Marketing of Toddler-Specific Food Products Launched in Australia between 1996 and 2020.

With the food system evolving, it is not clear how the nutrition and on-pack claims of toddler foods have been impacted. Data on the trends in Australia are lacking, so we sought to determine the changes in the nutrition and on-pack claims of toddler-specific packaged foods over time. A retrospective cross-sectional analysis was conducted using the Mintel Global New Products Database. The number of toddler-specific foods increased from 1996 to 2020. Over time, a lower proportion of meals and snacks were classified as "ultra-processed", but a higher proportion of snacks were classified as "discretionary". Meals launched after 2014 had higher median values for energy, saturated fat, and sugar than those in earlier years. Toddler snacks launched after 2014 had lower median values for sodium, and higher median values for fat, saturated fat, and sugar than those in earlier years. The mean number of total claims per package increased over time for snacks, with an increase in unregulated claims for both meals and snacks. Public health action is needed to ensure that the retail food environment for young children is health-promoting, including stringent and clear regulations for on-pack claims, and compositional guidelines and guidance on how to reduce the number of ultra-processed foods for toddlers.

Nutrition and packaging characteristics of toddler foods and milks in Australia.

OBJECTIVE: To analyse nutritional and packaging characteristics of toddler-specific foods and milks in the Australian retail food environment to identify how such products fit within the Australian Dietary Guidelines (ADG) and the NOVA classification. DESIGN: Cross-sectional retail audit of toddler foods and milks. On-pack product attributes were recorded. Products were categorised as (1) food or milk; (2) snack food or meal and (3) snacks sub-categorised depending on main ingredients. Products were classified as a discretionary or core food as per the ADG and level of processing according to NOVA classification. SETTING: Supermarkets and pharmacies in Australia. RESULTS: A total of 154 foods and thirty-two milks were identified. Eighty percentage of foods were snacks, and 60 % of foods were classified as core foods, while 85 % were ultraprocessed (UP). Per 100 g, discretionary foods provided significantly more energy, protein, total and saturated fat, carbohydrate, total sugar and Na (P < 0·001) than core foods. Total sugars were significantly higher (P < 0·001) and Na significantly lower (P < 0·001) in minimally processed foods than in UP foods. All toddler milks (n 32) were found to have higher energy, carbohydrate and total sugar levels than full-fat cow's milk per 100 ml. Claims and messages were present on 99 % of foods and all milks. CONCLUSIONS: The majority of toddler foods available in Australia are UP snack foods and do not align with the ADG. Toddler milks, despite being UP, do align with the ADG. A strengthened regulatory approach may address this issue.

The ARF tumor suppressor targets PPM1G/PP2Cγ to counteract NF-κB transcription tuning cell survival and the inflammatory response.

Inducible transcriptional programs mediate the regulation of key biological processes and organismal functions. Despite their complexity, cells have evolved mechanisms to precisely control gene programs in response to environmental cues to regulate cell fate and maintain normal homeostasis. Upon stimulation with proinflammatory cytokines such as tumor necrosis factor-α (TNF), the master transcriptional regulator nuclear factor (NF)-κB utilizes the PPM1G/PP2Cγ phosphatase as a coactivator to normally induce inflammatory and cell survival programs. However, how PPM1G activity is precisely regulated to control NF-κB transcription magnitude and kinetics remains unknown. Here, we describe a mechanism by which the ARF tumor suppressor binds PPM1G to negatively regulate its coactivator function in the NF-κB circuit thereby promoting insult resolution. ARF becomes stabilized upon binding to PPM1G and forms a ternary protein complex with PPM1G and NF-κB at target gene promoters in a stimuli-dependent manner to provide tunable control of the NF-κB transcriptional program. Consistently, loss of ARF in colon epithelial cells leads to up-regulation of NF-κB antiapoptotic genes upon TNF stimulation and renders cells partially resistant to TNF-induced apoptosis in the presence of agents blocking the antiapoptotic program. Notably, patient tumor data analysis validates these findings by revealing that loss of ARF strongly correlates with sustained expression of inflammatory and cell survival programs. Collectively, we propose that PPM1G emerges as a therapeutic target in a variety of cancers arising from ARF epigenetic silencing, to loss of ARF function, as well as tumors bearing oncogenic NF-κB activation.

Characterization of the mechanism by which the RB/E2F pathway controls expression of the cancer genomic DNA deaminase APOBEC3B.

APOBEC3B (A3B)-catalyzed DNA cytosine deamination contributes to the overall mutational landscape in breast cancer. Molecular mechanisms responsible for A3B upregulation in cancer are poorly understood. Here we show that a single E2F cis-element mediates repression in normal cells and that expression is activated by its mutational disruption in a reporter construct or the endogenous A3B gene. The same E2F site is required for A3B induction by polyomavirus T antigen indicating a shared molecular mechanism. Proteomic and biochemical experiments demonstrate the binding of wildtype but not mutant E2F promoters by repressive PRC1.6/E2F6 and DREAM/E2F4 complexes. Knockdown and overexpression studies confirm the involvement of these repressive complexes in regulating A3B expression. Altogether, these studies demonstrate that A3B expression is suppressed in normal cells by repressive E2F complexes and that viral or mutational disruption of this regulatory network triggers overexpression in breast cancer and provides fuel for tumor evolution.