Abstract A19 RARB2 is a master tumor suppressor that mediates the growth-inhibitory action of retinoic acid (RA). Homozygosis for epigenetically silent RARB2 alleles, which results into loss of RARB2 tumor suppressor activity, leads to RA-resistance, and apparently precedes the acquisition of morphological transformation of breast epithelial cells (Bistulfi et al., Cancer Research, 2006). RARB2 epigenetic silencing is marked by chromatin repressive changes, including DNA methylation of the RARB2 CpG island (Sirchia et al., Oncogene, 2000). In a mechanistic study, we demonstrated that methylation arises at a specific epicenter in the RARB2 CpG island, which we call the RARB2 methylation epicenter (RME) (Ren et al., MCB, 2005). Here we show that RME methylation is detectable in both benign and ductal carcinoma in situ within the same patient tissue sample. This finding implies sequential epigenetic silencing of RARB2 alleles. Timely identification of the first epigenetic hit in breast tissue of women at risk of breast cancer could prevent progression to RME methylation homozygosity, the consequent loss of RARB2-mediated tumor suppressor function, RA-resistance, and morphological epithelial cell transformation. This work was supported by the Roswell Park Alliance Foundation Award (NS), the Breast Cancer Coalition of Rochester (NS), the Susan Komen Foundation (SR). Citation Information: Cancer Prev Res 2008;1(7 Suppl):A19.
Core-binding factor leukemia (CBFL) is a subgroup of acutemyeloid leukemia (AML) characterized by genetic mutations involving the subunits of the core-binding factor (CBF). The leukemogenesis model for CBFL posits that one, or more, gene mutations inducing increased cell proliferation and/or inhibition of apoptosis cooperate with CBF mutations for leukemia development. One of the most commonmutations associated with CBF mutations involves the KIT receptor. A high expression of KIT is a hallmark of a high proportion of CBFL. Previous studies indicate that microRNA (MIR) 222/221 targets the 3′ untranslated region of the KIT messenger RNA and our observation that AML1 can bind the MIR-222/221 promoter, we hypothesized that MIR-222/221 represents the link between CBF and KIT. Here, we show that MIR-222/221 expression is upregulated after myeloid differentiation of normal bone marrow AC133+ stem progenitor cells. CBFL blasts with either t(8;21) or inv(16) CBF rearrangements with high expression levels of KIT (CD117) display a significantly lower level of MIR-222/221 expression than non-CBFL blasts. Consistently, we found that the t(8;21) AML1-MTG8 fusion protein binds the MIR-222/221 promoter and induces transcriptional repression of a MIR-222/221-LUC reporter. Because of the highly conserved sequence homology, we demonstrated concomitant MIR-222/221 down-regulation and KIT up-regulation in the 32D/WT1 mouse cell model carrying the AML1-MTG16 fusion protein. This study provides the first hint that CBFL-associated fusion proteins may lead to up-regulation of the KIT receptor by down-regulating MIR-222/221, thus explaining the concomitant occurrence of CBF genetic rearrangements and overexpression of wild type or mutant KIT in AML.
Abstract All-trans retinoic acid (RA), the bioactive derivative of vitamin A, can paradoxically exert both anti-cancer and cancer-promoting actions. According to our studies, RA promotes breast cancer cell growth and invasion whenever it fails to exert its genome-wide epigenetic control of transcription via the RA receptor alpha (RARA). Factors that negatively affect RARA genomic function lead to genome-wide deregulation of the transcriptome epigenetically regulated by RA, including RARA-target genes controlling critical sphingolipid functions. We show here that functional inhibition of genomic RARA concomitantly leads to epigenetic repression of neutral sphingomyelinase 2 (nSMase2/SMDP3), a critical enzyme involved in the synthesis of pro-apoptotic ceramide, and upregulation of S1PR1, a key receptor of sphingosine-1-phosphate (S1P), a sphingolipid with pro-proliferative and pro-invasive activity. Selective activation of S1PR1 recapitulates RA pro-proliferative/pro-invasive action, while selective inhibition of S1PR1 counteracts it. Apparently, activation of the S1P-S1PR1 axis, combined with lack of RARA-mediated epigenetic control of SMPD3-ceramide axis, contributes to determine RA tumorigenic action. This study was supported by the NCI R01 CA127614 grant (NS). Citation Format: Stefano Rossetti, Vincenzo Gagliostro, Nicoletta Sacchi. Deregulation of retinoic acid (RA) epigenetic control of sphingolipid signaling. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4532.
Abstract Background Human myelogenous leukemia characterized by either the non random t(8; 21)(q22; q22) or t(16; 21)(q24; q22) chromosome translocations differ for both their biological and clinical features. Some of these features could be consequent to differential epigenetic transcriptional deregulation at AML1 targets imposed by AML1-MTG8 and AML1-MTG16, the fusion proteins deriving from the two translocations. Preliminary findings showing that these fusion proteins lead to transcriptional downregulation of AML1 targets, marked by repressive chromatin changes, would support this hypothesis. Here we show that combining conventional global gene expression arrays with the power of bioinformatic genomic survey of AML1-consensus sequences is an effective strategy to identify AML1 targets whose transcription is epigenetically downregulated by the leukemia-associated AML1-MTG16 protein. Results We interrogated mouse gene expression microarrays with probes generated either from 32D cells infected with a retroviral vector carrying AML1-MTG16 and unable of granulocyte differentiation and proliferation in response to the granulocyte colony stimulating factor (G-CSF), or from 32D cells infected with the cognate empty vector. From the analysis of differential gene expression alone (using as criteria a p value < 0.01 and an absolute fold change > 3), we were unable to conclude which of the 37 genes downregulated by AML1-MTG16 were, or not, direct AML1 targets. However, when we applied a bioinformatic approach to search for AML1-consensus sequences in the 10 Kb around the gene transcription start sites, we closed on 17 potential direct AML1 targets. By focusing on the most significantly downregulated genes, we found that both the AML1-consensus and the transcription start site chromatin regions were significantly marked by aberrant repressive histone tail changes. Further, the promoter of one of these genes, containing a CpG island, was aberrantly methylated. Conclusion This study shows that a leukemia-associated fusion protein can impose a distinct epigenetic repressive signature at specific sites in the genome. These findings strengthen the conclusion that leukemia-specific oncoproteins can induce non-random epigenetic changes.
An entry from the Cambridge Structural Database, the world’s repository for small molecule crystal structures. The entry contains experimental data from a crystal diffraction study. The deposited dataset for this entry is freely available from the CCDC and typically includes 3D coordinates, cell parameters, space group, experimental conditions and quality measures.
Abstract Upregulation of RNA Polymerase (Pol I)-mediated transcription of ribosomal RNA (rRNA) and increased ribogenesis, which are necessary to sustain the increased metabolic demand of highly proliferating cancer cells, are hallmarks of cancer. Increased rRNA transcription can be due deregulation of tumor suppressors and oncogenes that affect Pol I activity. Overexpression of the MYC oncogene, a potent Pol I activators, is particularly frequent in cancer. In addition, based on our analysis of the Cancer Genome Atlas (TCGA), amplification/upregulation of genes encoding for basal components of the Pol I transcriptional machinery is also frequent in cancers of various histotype (Rossetti et al., Cell Cycle, 2016). By using breast and ovarian epithelial cells, we mechanistically found that: 1) ectopic expression of either MYC or RRN3 (TIF-IA), a key Pol I basal component, by increasing rRNA synthesis, is sufficient to induce in vitro transformation phenotypes and to promote cell proliferation; 2) there is a causal link between MYC overexpression and RRN3 upregulation; 3) MYC- or RRN3-induced rRNA upregulation sensitizes cells to the anti-proliferative action of drugs inhibiting rRNA transcription. Our findings provide a rationale for using drugs targeting rRNA transcription to curb proliferation of cancers cells due to rRNA upregulation by MYC. Funding for this study was provided by an RPCI-UPCI Ovarian Cancer Spore DRP award (NS), the NCI R01 CA127614 grant (NS), the Terri Brodeur Breast Cancer Foundation (SR), the Susan Komen Foundation (SR), and the NCI P30 CA016056 institutional grant. Citation Format: Stefano Rossetti, Andrzej J. Wierzbicki, Nicoletta Sacchi. Cancer cell ribogenesis: MYC and the integrity of the RNA polymerase I-rRNA machinery [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4479. doi:10.1158/1538-7445.AM2017-4479
Most physiological and biological processes are regulated by endogenous circadian rhythms under the control of both a master clock, which acts systemically and individual cellular clocks, which act at the single cell level. The cellular clock is based on a network of core clock genes, which drive the circadian expression of non-clock genes involved in many cellular processes. Circadian deregulation of gene expression has emerged to be as important as deregulation of estrogen signaling in breast tumorigenesis. Whether there is a mutual deregulation of circadian and hormone signaling is the question that we address in this study. Here we show that, upon entrainment by serum shock, cultured human mammary epithelial cells maintain an inner circadian oscillator, with key clock genes oscillating in a circadian fashion. In the same cells, the expression of the estrogen receptor α (ER A) gene also oscillates in a circadian fashion. In contrast, ER A-positive and -negative breast cancer epithelial cells show disruption of the inner clock. Further, ER A-positive breast cancer cells do not display circadian oscillation of ER A expression. Our findings suggest that estrogen signaling could be affected not only in ER A-negative breast cancer, but also in ER A-positive breast cancer due to lack of circadian availability of ER A. Entrainment of the inner clock of breast epithelial cells, by taking into consideration the biological time component, provides a novel tool to test mechanistically whether defective circadian mechanisms can affect hormone signaling relevant to breast cancer.
