DEAR1 maps close to another tumorsuppressor, CHD5, which was recently mapped to Chromosome 1q by a chromo- some engineering approach . With the identification of DEAR1 we are beginning to develop a deeper understanding of the molecular basis for the loss of Chromosome 1q in human cancers. While CHD5 regulates cell proliferation pathways by increasing expression of the cell cycle inhibitor p16/ink4a locus, DEAR1 regu- lates cell architecture. It is possible that CHD5 and DEAR1 represent the two sides of the transformation process, where loss of CHD5 results in aberrant proliferation while loss of DEAR1 results in loss of tissue architecture, and the combination of the events can drive changes in the epithelial tissues that can progress towards cancerous growth. It would be interesting to deter- mine the effect of combined loss of CHD5 and DEAR1 both for clinical prognosis and during transformation of epithelial cells in culture and animal models.
To clarify the possible mechanism of VEZT’s tumor-inhibiting function, we screened the target genes regulated by VEZT. cDNA microarrays provide a powerful tool for exploring complex gene expression profiles. Microarray analysis of experimental samples, such as gene-transfected cells, has led to identification of valuable molecular markers involved in tumor proliferation, invasion, migration, prognosis and therapeutic response [44,45,46]. Thus, we used a global cDNA microarray to identify downstream target genes of VEZT. We compared changes in gene expression profiles in gastric cancer cells with or without VEZT gene transfection and Figure 5. Molecular mechanism of the inhibitory activity of VEZT on gastric cancer growth. (A) TCF19-transfected MKN-45 showed increased percentage of G2/M phase cells and decreased percentage of S and G0/ G1 phase cells. (B) Over-expression of TCF19 promoted cell growth by CCK-8 assay. (C) Schematic summary of the VEZT tumorsuppressor gene on carcinogenesis of gastric cancer. VEZT is inactivated via hypermethylation, which may be induced by H. pylori infection. Restoring VEZT expression inhibits cell proliferation, migration, invasion and tumorigenesis both in vitro and in vivo, which could be explained by the downregulation of specific target genes identified by global microarray analysis. *P < 0.05. Each bar represents the mean value ± standard deviation from three independent experiments.
In a previous study, we demonstrated that KLF4 is downreg- ulated during the development and progression of cervical carcinoma . The overexpression of exogenous KLF4 protein was found to inhibit cervical carcinoma cell growth and tumor formation both in vitro and in vivo by activating the cell cycle suppressor p27 Kip1 , suggesting that KLF4 works as a tumorsuppressor in cervical carcinoma. Promoter CpG island hyper- methylation is a common cause in many malignancies, resulting in transcriptional silencing of many tumor suppression genes. The methylation status of the KLF4 promoter was therefore examined in tissues from normal cervix and cervical carcinoma. We profiled two CpG islands upstream of the KLF4 transcriptional start site, from 25 to 2266 bp (BSQ1), containing 22 CpG sites, and from 21684 to 21878 bp (BSQ3), containing 18 CpG sites (Fig. 1A). Two pairs of primers were designed to amplify the KLF4 promoter BSQ1 and BSQ3 regions. In the BSQ3 region, we performed quantitative bisulfite sequencing (BSQ) analysis using genomic DNA templates isolated from 24 primary cervical cancer tissues and 12 normal cervix tissues (Fig. 1B). As shown in Fig. 1C, low methylation levels were detected at the KLF4 promoter BSQ3 region in normal cervix samples (average methylation level was 11.11%). However, in cervical cancer tissues, methylation levels in this region were significantly higher than in normal cervix tissues at each individual CpG site except CpG4 (41.9%, P ,0.05). In the BSQ1 region of the KLF4 promoter, low methylation levels were detected in both cervical cancer and normal cervix tissues (data not shown). Altogether, these results suggest that hypermethylation of the KLF4 promoter BSQ3 region, and not the BSQ1 region, is involved in cervical carcinogenesis.
To investigate the molecular mechanism of the tumorsuppressor role of miR-217 in GC, we used luciferase reporter assay and western blot to confirm that GPC5 was a target of miR-217 in GC cells. To confirm the direct regulation of GPC5 by miR-217, we used GPC 3’ UTR reporter vector bearing the potential miR-217 binding site in the fluorescent reporter. Further- more, qRT-PCR and western blot assay showed that overexpression of miR-217 inhibited GPC expression. Glypicans are a family of proteoglycans that are linked to the exocytoplasmic sur- face of the plasma membrane via a glycosyl phosphatidylinositol anchor[30, 31]. Six glypicans have been identified in mammals (GPC1 to GPC6)[32, 33]. GPC5 is mainly expressed in devel- oping central nervous system, limbs, kidney, lung and liver[34, 35]. Recent studies have indi- cated that some GPCs, especially GPC3 andGPC5, might play an important role in regulating cancer progression[35, 36]. For example, Zhang et al. showed that GPC5 was highly expressed in SACC-M(high lung-metastatic cell line) and in clinical samples of salivary adenoid cystic carcinoma (SACC) cases with lung metastasis. The overall expression level of GPC5 in clinical cases of SACC with lung metastasis was higher. Williamson et al. showed that the gene encoding GPC5was amplified in 20% of patients with alveolar rhabdomyosarcoma (RMS) and that this glypican was overexpressed in RMS patients. Moreover, down-regulation of GPC5 expression by siRNA inhibited the proliferation rate of RMS cells. Another study found that high levels of GPC5 expression predicted poor postsurgical survival times for curatively respected NSCLC patients, suggesting the value of GPC5 as a molecular prognostic indicator [35, 37]. In our study, the expression of GPC5 was higher in GC cell lines and the protein levels
In normal neural stem cells, the H3K27 specific demethylase JMJD3 was shown to modulate differentiation by affecting p53 nuclear distribution . The implications of these findings in any cancer model, however, remains unclear. Here, we reveal that the H3K27-specific demethylase, JMJD3, promotes differentiation and suppresses proliferation of GSCs, not only through its well- known chromatin-dependent activation of the INK4A/ARF locus, but also through p53 protein nuclear stabilization. Based on previous studies demonstrating global lysine demethylation on p53 following interaction with JMJD3 in differentiating neural stem cells , our results suggest that lysine 372 modifications may represent one of many putative mechanisms resulting in p53 protein stabilization following interaction with JMJD3 resulting in differentiation and suppression of proliferation in glioblastoma stem cells. We also demonstrate that primary-patient-derived GSCs and primary human GBMs counter the tumorsuppressor effects of JMJD3 through hypermethylation of an evolutionarily conserved intragenic and enhancer-like DNA regulatory element within the JMJD3 locus or direct somatic mutations of the JMJD3 gene. Pharmacological demethylation of GSC with hypermethyla- tion-silenced JMJD3 or overexpression of a wildtype JMJD3 in JMJD3 mutated GSCs results in p53 pathway activation and GSC differentiation in vitro and hugely extended survival of animals with intracranial GSC xenografts. The clinical relevance of these findings is reflected by our demonstration through genome wide transcriptional motif analyses in a large number of GBM patients that GBMs with high levels of JMJD3 expression and wildtype TP53 show significantly enhanced p53 pathway activation.
cascade via both classical and alternative pathways. No activated complement C3 fragments were shown to deposit on cell surface. Human tumorsuppressor gene WWOX encodes the mRNA for translating into WWOX/WOX1, WOX2 and other isoforms (if present) [20,21,43,44]. In the absence of C1q or C6, expression of WOX2 is downregulated, whereas WOX1 expression is not affected. These observations indicate that C1q and C6 are likely to support alternatively splicing of WWOX mRNA, thus leading to the generation of WOX2 protein. Whether WOX2 acts as a tumorsuppressor is unknown. Presumably, C1q and C6 are needed for serum-dependent cell survival by maintaining ERK activation and WOX2 expression. WOX2 may act as a dominant negative and counteracts the tumorsuppressor function of wild type WWOX/WOX1 in vivo, thereby supporting cancer growth. Both WOX1 and WOX2 have been shown to be downregulated in the neurons of Alzheimer’s disease . Indeed, C1q has been implicated in the pathogenesis of neuronal death in the neurodegenerative diseases such as in Alzheimer’s disease [45,46]. There is a strong possibility that C1q activates WOX1 in neurons, which ultimately leads to cell death. Conceivably, the Figure 8. Complement C1q and C6 are essential for maintaining ERK and WOX1 activation and expression of isoform WOX2. (A,B) DU145 cells were cultured overnight under serum-free (SF) conditions, or in 1% normal human serum (NHS) or NHS depleted with C1q (DC1q), C6 (DC6), C7 (DC7), C8 (DC8), or C9 (DC9). Significant reduction of isoform WOX2 expression was observed in DU145 cells when cultured in DC1q or DC6 serum (versus SF controls; p,0.001; Student’s t test), whereas DC7, DC8, or DC9 serum was less effective. Activation or phosphorylation of ERK (p-ERK) was dependent upon the presence of C1q or C6 in serum (versus SF controls; p,0.001; Student’s t test). A representative set of data from 3 experiments is shown. The bar graphs show the average of 3 experiments. (C) The above cells were also grown on cover glass under identical serum conditions. By immunofluorescence microscopy, serum without C1q or C6 could not support the constitutive expression of WOX2 (p,0.0001, as versus SF or NHS; Student’s t test). Approximately 200 cells were quantified individually under fluorescence microscopy, and the extent of fluorescence was subtracted (or normalized) from negative controls (cells stained with secondary antibody only). Nuclei were stained with DAPI. (D,E) The same cells, as indicated in (A), were stained with p-WOX1 antibody and the extent of protein expression was quantified. (F) Again, identical experiments were carried out for Western blotting. Under C1q-free conditions (DC1q serum), the basal activation of WOX1 was significantly reduced (,50% reduction; p,0.001 from versus SF and NHS; Student’s t test; n = 3).
Over-expression of Notch1 leads to induction of the EMT phenotype and increased expression of miR-21 . Genistein has been shown to inactivate Notch and hedgehog signaling [31,32] and we previously reported that genistein inhibited tumor cell growth by reducing miR-21 expression in renal cell carcinoma . Thus genistein has a tumorsuppressor function, regulating ‘Notch signaling’ by down-regulation of miR-21. Genistein can also reduce cell proliferation by regulating ‘Wnt signaling’ [33,34,35] through miR-574-3p in cancer. In this study EGFR, a putative target gene for miR-574-3p, was up-regulated in PCa and increased in advanced cancer [36,37]. EGFR expression was correlated with a high Gleason score, disease relapse and hormone-refractory status [37,38]. Researchers have reported that EGFR is regulated by several miRNAs such as miR-7, miR- 128b, miR-133, miR-145, miR146a, miR-146b-5p, miR-331-3p, miR-542-5p [39–47]. Genistein up-regulated miR-146a expres- sion in pancreatic cancer cells and functions as a tumorsuppressor in castration-resistant PCa [44,45]. Genistein might down- regulated EGFR levels by up-regulating miR-574-3p.
Epigenetic mechanisms are frequently deregulated in cancer cells and can lead to the silencing of genes with tumorsuppressor activities. The isoform A of the Ras-association domain family member 1 (RASSF1A) gene is one of the most frequently silenced transcripts in human tumors; however, few studies have simultaneously investigated epigenetic abnormalities associated with the 3p21.3 tumorsuppressor gene cluster lanking RASSF1 (i.e., SEMA3B, HYAL3, HYAL2, HYAL1, TUSC2, RASSF1, ZMYND10, NPRL2, TMEM115 and CACNA2D2). This study aimed to investigate the role of epigenetic changes to these genes in 17 breast cancer cell lines and in three non-tumorigenic epithelial breast cell lines (184A1, 184B5 and MCF 10A) and to evaluate the efect on gene expression of treatment with the demethylating agent 5-Aza -2’-deoxycytidine and/or Trichostatin A (TSA), a histone deacetylase inhibitor. We report that, although the RASSF1A isoform was determined to be epigenetically silenced in 15 of the 17 breast cancer cell lines, all the cell lines expressed the RASSF1C isoform. Five breast cancer cell lines overexpressed RASSF1C when compared with the normal epithelial cell line 184A1. Furthermore, the genes HYAL1 and CACNA2D2 were signiicantly overexpressed after the treatments. After the combined treatment, RASSF1A re-expression was accompanied by an increase in expression levels of the lanking genes. The Spearman’s correlation coeicient indicated a positive co-regulation of the following gene pairs: RASSF1 and TUSC2 (r = 0.64, p = 0.002), RASSF1 and ZMYND10 (r = 0.58, p = 0.07), RASSF1 and NPRL2 (r = 0.48, p = 0.03), ZMYND10 and NPRL2 (r = 0.71; p = 0.0004) and NPRL2 and TMEM115 (r = 0.66, p = 0.001). Interestingly, the genes TUSC2, NPRL2 and TMEM115 were found to be unmethylated in each of the untreated cell lines. Chromatin immunoprecipitation using antibodies against the acetylated and trimethylated lysine 9 of histone H3 demonstrated low levels of histone methylation in these genes, which are located closest to RASSF1. These results provide evidence that epigenetic repression is involved in the downregulation of multiple genes at 3p21.3 in breast cancer cells.
WWOX , the gene that spans the second most common human chromosomal fragile site, FRA16D, is inactivated in multiple human cancers and behaves as a suppressor of tumor growth. Since we are interested in understanding WWOX function in both normal and cancer tissues we generated mice harboring a conditional Wwox allele by flanking Exon 1 of the Wwox gene with LoxP sites. Wwox knockout (KO) mice were developed by breeding with transgenic mice carrying the Cre- recombinase gene under the control of the adenovirus EIIA promoter. We found that Wwox KO mice suffered from severe metabolic defect(s) resulting in growth retardation and all mice died by 3 wk of age. All Wwox KO mice displayed significant hypocapnia suggesting a state of metabolic acidosis. This finding and the known high expression of Wwox in kidney tubules suggest a role for Wwox in acid/base balance. Importantly, Wwox KO mice displayed histopathological and hematological signs of impaired hematopoeisis, leukopenia, and splenic atrophy. Impaired hematopoeisis can also be a contributing factor to metabolic acidosis and death. Hypoglycemia and hypocalcemia was also observed affecting the KO mice. In addition, bone metabolic defects were evident in Wwox KO mice. Bones were smaller and thinner having reduced bone volume as a consequence of a defect in mineralization. No evidence of spontaneous neoplasia was observed in Wwox KO mice. We have generated a new mouse model to inactivate the Wwox tumorsuppressor gene conditionally. This will greatly facilitate the functional analysis of Wwox in adult mice and will allow investigating neoplastic transformation in specific target tissues.
The Drosophila and mammalian forms of TSG101 are quite similar at a primary sequence level (46% identical/61% similar), share the same domain structure , and are predicted to have very similar molecular properties. Each has also been shown to function as part of the same conserved complex, ESCRT-I, and to be involved in the same biological process: endocytic trafficking of internalized receptors and membrane proteins. Thus the observed differences in the phenotypes elicited by loss of vertebrate and invertebrate TSG101 are likely to arise either due to 1) differences in the strength of alleles used in each system, 2) differences in the spectrum of proteins routed into the ESCRT pathway in flies and mammals, or 3) functional redundancy for TSG101 within the vertebrate genome. With regard to the former possibility, the genomic alleles used for analysis in both organisms are loss of function embryonic lethal [5,16,55] that can be rescued by reintroduction of a wild-type version of the gene [5,15]. In addition, TSG101 does not appear to be a member of a multi-gene family in vertebrates. Differences in cellular phenotypes produced by invertebrate and vertebrate ept/TSG101 alleles thus may reflect stage- or tissue-specific differences in the spectrum of proteins routed into the ESCRT pathway in each type of organism. In addition to Crb and Notch, Drosophila ept has now been shown to affect localization and levels of the Dome receptor (see model in Fig. 7). Loss of the ESCRT-II subunit and tumorsuppressor gene Figure 5. Stat92E sensor activity in ept mutant eye-antennal
We show that inhibition of erbB2 signaling, either genetically or by small chemical inhibitors, leads to suppression of EMT and a neoplastic phenotype in pen/lgl2 mutant larvae. Intriguingly, E-cad membrane localization is restored in these larvae, indicating that the loss of E-cad is a consequence of activation of ErbB signaling rather than a cause of it. Thus, our analyses presented here suggest that Lgl2 acts as a tumorsuppressor by regulating the amplitude of ErbB signaling in the epidermis. Since we did not observe snail- mediated down-regulation of E-cad, we propose that an erbB2 dependent pathway in pen/lgl2 mutants leads to the destabilization of E-cad at adherens junctions. Indeed in pen/lgl2 mutant epidermal cells, known modifiers of E-cad function, such as mmp9 and sgk1 are up-regulated (Figure 3). While Mmps are known to be involved in ecto-domain shedding of E-cad [57,58], Sgk1 functions in the phosphorylation of Ndrg1, a protein involved in vesicular recycling of E-cad [59,60]. Additionally, recently published data from cell culture suggests an involvement of RTK signaling in the destabilization of adherens junctions via Numb . Here, Numb functions as an adapter protein coupling Figure 5. Phylogenetic analysis of erbB family members and zebrafish mutants of erbB paralogs. (A) Phylogenetic analysis of erbB family members in the zebrafish genome (zv7) with the human orthologs (Minimum evolution algorithm, 100 replicates). With the exception of erbB2, all other erbB family members are duplicated in zebrafish. (B–D) DIC Images of wild-type (B), erbB2 2/2 (C) and erbB3b 2/2 (D) larvae at 132hpf. (E) In-situ
Epithelial malignancies such as colorectal and breast cancer are thought to arise and progress towards malignancy due to alterations in signal transduction pathways that regulate the balance between self-renewal and differentiation in adult stem cell compartments . The canonical Wnt/b-catenin signal transduc- tion pathway plays a rate-limiting role in embryonic and adult stem cell renewal, and its aberrant activation is among the most common signaling defect in human cancers . Activation of the canonical Wnt pathway leads to intracellular b-catenin stabiliza- tion and its translocation to the nucleus where it interacts with members of the Tcf/Lef family of transcription factors to modulate the expression of specific Wnt target genes (http:// www.stanford.edu/˜rnusse/pathways/targets.html). In the gastro- intestinal tract, Wnt/b-catenin signaling regulates stemness and differentiation of epithelial cells along the crypt-villus axis [3,4]. Accordingly, truncating mutations in the APC tumorsuppressor gene, the main negative regulator of the Wnt/b-catenin pathway,
WITH TUMORSUPPRESSOR GENE p53 IN BREAST CARCINOMA Correlation of standard pathomorphological prognostic parameters, primary tumor size and axillary nodal status with new prognostic factor in breast carcinoma: tu- mor suppressor gene p53 was analyzed. The studied sample included 65 women who underwent surgery for breast carcinoma at the Surgical Clinic of Clinical Center Banja Luka, from January 1st 1997 till January 1st 1999. Statistical data analysis was performed and correlation of prognostic factors was determined. The majority of authors in this field agree that the primary tumor size and axillary nodal status are the two most important prognostic factors. These factors are the best predictors of prognosis and survival of women who had the tumor and were oper- ated on. Tumor markers were immunohistochemicaly determined in the last ten years and, according to the majority of authors, are still considered the additional or relative prognostic factors in breast carcinoma. Their prognostic value and sig- nificance increase almost daily. Most frequently determined tumor markers are bcl-2, pS2, Ki-67 and p53. There was a positive, directlly proportional relationship between primary tumor size and tumorsuppressor gene p53, but there was no positive correlation between the axillary nodal status and tumorsuppressor gene p53. Significance of determination of new tumor markers as the prognostic factors was emphasized. These markers represent a powerful tool in the early detection and prevention of breast carcinoma.
LKB1 (also known as STK11) has emerged as a major tumorsuppressor in diverse malignancies, particularly melanoma, cervical cancer, and lung cancer [1–5]. The LKB1 gene encodes a serine/threonine kinase that acts through a multitude of targets to control diverse aspects of cell polarity, metabolism, and cell growth [6,7]. Among these diverse substrates, the a catalytic subunit of AMP-activated protein kinase (AMPK) is the best established both in normal physiologic states and cancer. The LKB1 protein, in association with the accessory proteins STRAD and MO25, phosphorylates AMPKa at Thr172 in its activation loop, leading to AMPK activation when AMPK is in the AMP- bound state. AMPK directly phosphorylates TSC2 and raptor to suppress signaling through mTOR pathway, and mTOR pathway hyperactivity in the LKB1-deficient state is believed to account for some, but not all of LKB1 tumorsuppressor functions [8,9].
At PND84, we show that absolute tumor volume was not statistically different between all three genotypes (total tumor volume alone or as a percentage of the retinal volume). However, since these tumors arise from fewer tumor-initiating cells, we conclude that tumor growth per initiating cell was greater in mice with mutant Cdh11 alleles (Figure 7B and 7C, Figure 8A). We conclude that Cdh11 functions as a tumorsuppressor. Since tumor ‘‘growth’’ results from an imbalance between cell death and proliferation, we examined cell proliferation (Figure 8B) and cell death (Figure 8C) in TAg-RB tumors of mice with normal Cdh11 alleles versus mutated Cdh11 alleles. Our data indicate that when Cdh11 is lost, cell death is deficient while proliferation remains unchanged, suggesting that the tumorsuppressor function of Cdh11 is mediated through promotion of apoptosis rather than inhibition of cell proliferation. This is further supported by our in vitro data showing significant decrease in caspase-3 and increase in b-catenin expression in Cdh11 knockdown experiments using siRNA (Figure 8D and Figure S2A, S2B), while proliferation markers PCNA and Ki67 remain unchanged (Figure S3).
We have also experimentally tested whether RASSF1A (genomic DNA, exons1 and 2) harbored mutations in normal tissues and found one mutated clone out of 14 in normal kidney (normal control to T356, see section ‘‘Frequent mutations in RASSF1A in human carcinomas’’). Important to note that so called ‘‘normal’’ kidney could be already partially transformed despite of normal phenotype because it was obtained from tissues adjacent to the tumor. We also sequenced complete RASSF1A cDNA from normal heart and detected six mutated clones out of 15 tested. All six heart mutated clones contained the same two mutations: L214L with codon changed from CTA to CTG and V236V with codon changed from GTA to GTG. Mutations in heart RASSF1 cDNA were most likely SNP as they could be also found in other RASSF1 clones in public databases (e.g. AC002481, NM_170713.2, NM_170714.1). In any case it is clear that mutations in RASSF1 in normal cells are more rare than in cancer cells.
methylation contributes to carcinogenesis and is responsi- ble for the chromosomal instability, reactivation of transposable elements, and loss of imprinting (Esteller and Herman, 2002). Therefore, the profile of hypermethylation promoters differs according to each cancer type, because for each tumor type specific genes are methylated. In addi- tion to that, the epigenetic inactivation may affect all mo- lecular mechanisms involved in cell immortalization and transformation. Last but not least, it seems that epigenetic changes are among the several early steps in carcinogenesis (Mukai and Sekiguchi, 2002).
We now document a physiological role of Arf in mouse male germ cell development that is distinct from its tumor suppressive functions in key respects. First, Arf is expressed in spermatogonia, but not in the primary spermatocytes that arise from them. Expression of p19 Arf neither arrests spermatogonial mitotic progression nor triggers their p53-dependent apoptosis. However, the absence of Arf expression in spermatogonia leads to p53- dependent apoptosis of spermatocytes before they exit meiosis-I. The defect in spermatogenesis is germ cell autonomous and results in a significant reduction in sperm counts by the time Arf-null mice are two months old, although residual sperm production maintains fertility in young males. Thus, expression of Arf in mitotic progenitor cells enhances the survival of their meiotic progeny in which Arf expression is normally extinguished. These features indicate that Arf expression initiates a salutary, feed-forward program that facilitates meiotic progression. Indeed, although Arf and Ink4a are widely viewed to convey tumor suppressive functions that coordinate the activities of the p53 and Rb signaling ‘‘pathways,’’ inactivation of Arf and Ink4a in the testes leads to opposing outcomes. Disruption of Ink4a increases the mitotic activity of spermatogonial progenitors to enhance sperm output and, in this respect, compensates for Arf loss of function without eliminating the cellular defects that arise in the Arf-null setting. In short, loss of Ink4a increases the spermatogonial pool size, but without Arf expression, spermatocytes undergo increased apopto- sis, returning the number of mature sperm to normal levels.
p53 protects us from cancer by transcriptionally regulating tumor suppressive programs designed to either prevent the development or clonal expansion of malignant cells. How p53 selects target genes in the genome in a context- and tissue-specific manner remains largely obscure. There is growing evidence that the ability of p53 to bind DNA in a cooperative manner prominently influences target gene selection with activation of the apoptosis program being completely dependent on DNA binding cooperativity. Here, we used ChIP-seq to comprehensively profile the cistrome of p53 mutants with reduced or increased cooperativity. The analysis highlighted a particular relevance of cooperativity for extending the p53 cistrome to non-canonical binding sequences characterized by deletions, spacer insertions and base mismatches. Furthermore, it revealed a striking functional separation of the cistrome on the basis of cooperativity; with low cooperativity genes being significantly enriched for cell cycle and high cooperativity genes for apoptotic functions. Importantly, expression of high but not low cooperativity genes was correlated with superior survival in breast cancer patients. Interestingly, in contrast to most p53-activated genes, p53-repressed genes did not commonly contain p53 binding elements. Nevertheless, both the degree of gene activation and repression were cooperativity- dependent, suggesting that p53-mediated gene repression is largely indirect and mediated by cooperativity- dependently transactivated gene products such as CDKN1A, E2F7 and non-coding RNAs. Since both activation of apoptosis genes with non-canonical response elements and repression of pro-survival genes are crucial for p53’s apoptotic activity, the cistrome analysis comprehensively explains why p53-induced apoptosis, but not cell cycle arrest, strongly depends on the intermolecular cooperation of p53 molecules as a possible safeguard mechanism protecting from accidental cell killing.
Malignant diseases are caused by defects in cell differentiation, cell cycle progression, DNA repair mechanisms, or apoptotic pathways [55–57]. Although previous studies have revealed the genetic backgrounds of certain types of cancer—for example, the MSH2 gene in familial nonpolyposis colon cancer , the BRCA1 gene in familial breast cancer [59–61], the APC gene in hereditary adenomatous polyposis [62,63], and the RB gene in retinoblastoma [64,65]—the functional mechanisms leading to cancer initiation, progression and development remain to be elucidated. BCL7B, located on chromosome 7, is a member of the BCL7 family of genes and is thought to be a tumor-associated gene [7–9]. In this study, we investigated the functional roles of the bcl- 7 and BCL7B genes in the Wnt signaling pathway and apoptosis in C. elegans and in human gastric cancer cells, respectively. Our results revealed that BCL-7 regulates terminal cell differentiation in somatic ‘‘stem-like’’ seam cells and SGPs via negative regulation of the Wnt pathway in C. elegans. BCL-7 also functions as a positive regulator of apoptosis by inhibiting the expression of an anti-apoptotic factor. Additionally, similar to its role in C. elegans, human BCL7B functions as a negative regulator of Wnt signaling, presumably upstream of ß-catenin, and induces apoptosis in gastric cancer cells. Furthermore, this study revealed that BCL-7 and BCL7B are also involved in the mechanisms of nuclear enlargement, which is an important signature of malignancy. Collectively, our data suggest that the members of the BCL7 family and their homolog proteins may function as tumor suppressors by affecting multiple pathways (S14 Fig.). Therefore, patients with WBS who have a heterozygous deletion in BCL7B may be at risk of malignancy. This study may also be clinically significant in terms of the long-term medical care of WBS patients.