The Role of Transcription Factor YY1 in the Biology of Cancer

Abstract

The transcription factor Yin Yang 1 (YY1) is a ubiquitously expressed protein involved in several biological functions, including embryogenesis, differentiation, replication, and cellular proliferation. YY1 can both activate and repress transcription depending on its interactions with other transcription factors and co-factors. It affects the transcription of a large number of mammalian and viral genes. In this review, we focus on the role of YY1 in cancer biology, including its expression, function, regulation by other upstream factors, and some of its more significant downstream effectors. We also discuss its context-dependent and tumor-specific role in human cancer progression, roles as a prognostic marker, and how its interactions with other cancer-associated factors can be exploited to develop novel targeted therapies.

I. INTRODUCTION

Yin Yang 1 (YY1) is a ubiquitously expressed protein originally identified as a zinc finger protein belonging to the GLI–Kruppel gene family. It is composed of 414 amino acids and has a molecular mass of 44 kDa. The C-terminal domain of YY1 has a nuclear localization signal leading to its primary localization in the nucleus.¹ YY1 can also undergo a DNA-replication-associated switch in its localization from the cytoplasm to the nucleus at the onset of the S phase.² YY1 has a strong affinity for the DNA sequence CGCCATNTT and interacts physically with a large number of cellular factors. Approximately 7% of vertebrate and 24% of viral promoters contain the YY1 consensus sites in their regulatory regions.³ It is involved in both transcriptional activation and repression^4–6 depending on the binding site, protein interactions, or levels within the cell. For example, as YY1 levels increase in 293T, the androgen receptor (AR)-mediated prostate specific antigen (PSA) transcription is enhanced while further increases begin to decrease PSA expression.^5,7–11 YY1 also interacts with a large number of proteins, including coactivators, corepressors, and other transcription factors. YY1 is implicated in chromatin structure, cytokinesis, differentiation, cell cycle control, oncogenesis, imprinting control, X-chromosome inactivation, and Polycomb group function.^12–16 YY1 also regulates the expression of retroviruses, herpes viruses, and adeno-related viruses^17,18 and plays a crucial role in development, as suggested by embryonic lethality in homozygous YY1-knockout mice. YY1 heterozygote embryos showed developmental growth retardation and neurulation defects of exencephaly with open brain, suggesting its role in later stages of mouse embryogenesis.¹⁹ In another study, constitutive ablation of YY1 in mice caused peri-implantation lethality and the investigators noted a dosage-dependent requirement of YY1 during late embryogenesis. YY1 reduces cell proliferation of mouse embryonic fibroblasts in a concentration-dependent manner, with complete ablation causing cytokinesis failure and cell cycle arrest.²⁰ Conditional knock-down of YY1 in mouse B cells revealed its role as a critical regulator of early B-cell development.^21,22 Oligodendrocyte-lineage-specific conditional ablation of YY1 in mouse causes defective myelination, ataxia, and tremor.²³ Melanocyte-specific YY1 knock-out in mice suggests that it is required for melanocyte development and post-developmental survival.²⁴

II. FUNCTIONS AND REGULATION

A. Regulation of YY1 Activity

Studies have shown YY1 activity is modulated by adenovirus-derived E1A protein, which activates the AAV P5 promoter.²⁵ The presence of E1A activates YY1-mediated transcription, whereas the absence of E1A suppresses YY1-mediated transcription.³ Many studies have shown increased activity at promoters for genes such as the histone deacetylase complex 1 (HDAC1) and 2 (HDAC2) with increased YY1 activity. HDACs may regulate YY1 in conjunction with cofactors such as mSin3A, nuclear receptor corepressor, and Sin3-associated polypeptide 18/30.²⁶ In an interesting observation, Hiromura et al.²⁷ showed that a subset of nuclear YY1 is O-GlcNAcylated regardless of the differentiation status of the cells. They found that glucose strongly stimulates O-linked N-acetylglucosaminylation on YY1. Glycosylation of YY1 promotes its activity by disrupting its interaction with the retinoblastoma protein (Rb), which allows it to bind DNA. Reports have also shown the direct modulation of YY1 transcriptional activity. Lee et al.⁸ have shown that the bone morphogenic protein (BMP) modulates the YY1 transcriptional activity by direct interaction of YY1 with BMP-activated SMADs. YY1 binds directly to the Rel-B component of the transcription factor nuclear factor kappa B (NF-κB), which in turn binds to the hs4 enhancer region of the B-cell lymphoma Igh gene, leading to an anti-apoptotic response and upregulation of proliferative potential of these lymphocytes in vivo.²⁸

B. YY1-Mediated Transcriptional Repression

Two studies have suggested the presence of two transcriptional domains in YY1, a repression domain at the C-terminal region coinciding with the zinc finger DNA-binding motif, and an activation domain at the N-terminus either within the first 90 or 69 amino acids of YY1.³ Three models have been proposed for YY1-mediated repression. According to first model, YY1 hinders the binding of trans-activators to DNA sterically by binding competitively to promoter elements with overlapping DNA recognition sites for transcriptional activators. The second model proposes that YY1 inhibits the interaction of activators with the general transcriptional machinery or masks the general factors from an activator. In the third model, YY1 recruits a corepressor or complex to a promoter, which in turn affects negatively other factors that are present or alters the local chromatin structure for repression. These proposed models are not mutually exclusive and it is likely that a combination of any two or all three operate at a single promoter.

C. YY1-Mediated Transcriptional Activation

The first indication that YY1 could function as a transcriptional activator came from the observation that the adenovirus E1A protein can relieve YY1-mediated repression and further activates transcription through a YY1-binding site.⁴ Soon after the cloning of YY1, it was shown that YY1 activates transcription by binding to the AAV P5 initiator element.⁷ Mutation of YY1 binding site on mouse ribosomal protein gene promoter down-regulates transcription.⁶ Based on results by different groups of investigators, it has been suggested that YY1 activates transcription by interacting directly with general transcription factors such as TBP, TAFII55, TFIIB, and AR^29–32; by inhibiting repressor activity and/or unmasking activation domain; or by recruiting co-activators that elicit a response at the target promoter. YY1 also interacts with well-known coactivators CBP and p300, both of which contain histone acetyltransferase activity.^10,33–37 Our recent study suggested that YY1 interacts with GON4L, which acts as a coactivator and enhances the binding of YY1 to the AR, another transcription factor, to activate transcription of CD24.³⁸

III. ROLE IN CANCER

A. YY1 Expression in Cancer

Over the last 12 years, a number of studies reported increased expression of YY1 at the mRNA, copy number variation, and protein levels in patient samples compared with normal samples in different types of cancers, including bladder cancer,⁴ brain cancer,⁶ breast cancer,^8,9,10,14 cervical cancer,¹⁶ colon cancer, endometrioid endometrial cancer,²⁴ esophageal squamous cell carcinoma,²⁶ follicular lymphoma,^27,28 gastric adenocarcinoma (GAC),²⁹ hematological malignancies,³¹ melanoma,^39,40 multiple myeloma (MM),^41–43 osteosarcoma,^44,45 ovarian cancer,^46–49 pancreatic ductal adenocarcinoma (PDAC),^50–52 prostate cancer,^53–56 and renal cell carcinoma (RCC).⁵⁷ Similar results have also been reported in cell lines representing some of the above cancer types.^39–45 However, increased YY1 expression is not always correlated with poor prognosis in patients^46–48 or leads to enhancement of in vitro tumor phenotypes.^46,49–55 In conclusion, it appears that high levels of YY1 can drive more aggressive phenotypes, but, in many cases, this is dependent on the cellular/tumor context.

B. YY1 Mutation in Cancer

YY1 is rarely mutated in human cancer (Fig. 1A); the most commonly mutated tumor type is in insulinomas (Fig. 1B).⁵⁶ Three different studies have reported the recurrent somatic point mutation at Thr372Arg of the DNA-binding zinc finger domain of YY1 in insulinoma patient samples (Fig. 1C).⁵⁶ Cromer et al. reported that this mutation results in the constitutive activation of cAMP and Ca(2+) signaling pathways involved in insulin secretion.⁵⁷ Lichtenauer et al. reported this mutation in 13% of patient samples.⁵⁸ In that study, the mutations were associated with a trend toward higher age and all affected patients were females. Cao et al. reported this mutation in 30% (34/113) of insulinoma tumors. They also observed that this mutation was associated with the later onset of tumors.⁵⁹

FIG. 1.

Genomic alterations in YY1 across different cancer types; (A) Bar graph showing the frequency of various alterations in YY1; (B) Occurrence of mutations in the YY1 gene as reported from different cancer type datasets; (C) Three missense mutations at T372R at the C-terminal end in insulinoma patient samples. (Data adapted from cBioPortal, http://www.cbioportal.org⁵⁶).

C. Functional Association of YY1 Expression and Tumor Phenotype

Manipulating YY1 expression in cancer cell lines can have either positive or negative effects on their anchorage-dependent and -independent growth, migration, invasion, xenograft tumor growth, drug sensitivity, and apoptosis. We reported recently that YY1 depletion retards growth in monolayers and in soft agar in vitro of bladder, prostate, lung, and breast cancer cell lines. Furthermore, YY1 knockdown completely abolishes the tumor-forming ability of the high tumorigenic bladder cancer cell line UMUC3.³⁸ In breast cancer, overexpression of YY1 results in downregulation of FEN1 and sensitization of cancer cells to mitomycin C or Taxol.⁶⁰ YY1 depletion inhibits clonogenicity, migration, invasion, and tumor formation of breast cancer cells, but has no effect on clonogenicity of immortal non-tumorigenic cells. Ectopic expression of YY1 in nontumorigenic MCF-10A cells enhanced migration and invasion.⁶¹ Conversely, Lee et al.⁴⁹ reported that YY1 overexpression in breast cancer cells inhibited cell proliferation, foci formation, and tumor growth in nude mice. Ishii et al.⁵⁴ reported that YY1 can suppress the growth of different tumor cell types, including breast and glioblastoma cancer cells.

Knocking down YY1 in colon cancer cell lines suppresses cell proliferation and induced apoptosis.⁶² Similarly, YY1 depletion inhibits endometrioid endometrial cancer cell proliferation and migration both in vitro and in vivo, whereas its overexpression promotes cell growth.⁴⁰ In gastric cancer, YY1 expression increases cell proliferation, migration, invasion, and tumor sphere formation.^41,63 YY1 overexpression promotes the growth of immortalized nontumorigenic normal human hepatocyte cells, whereas its knock-down in hepatocellular carcinoma (HCC) cells inhibited growth and was accompanied by distinct morphological changes.⁴³ In melanoma and MM cells, YY1 knock-down inhibited growth properties, migration, and invasion.^64,65 YY1 knock-down in ovarian cancer cells inhibits growth and motility, but also increases resistance to taxanes.⁴⁵ YY1 knock-down also inhibits the growth, migration, and invasion of renal cell carcinoma cells.⁶⁶ YY1 overexpression in esophageal squamous cell carcinoma TE-1 cells confers radio-resistance and reduces cell proliferation.⁵¹ Wang et al. reported that YY1 overexpression in CL1-10 lung cancer cells reduces cell-invasive capability by up-regulation of E-cadherin.⁵² YY1 overexpression in pancreatic cancer cell lines promotes apoptosis and increases the pro-apoptotic Bax protein.⁵³

D. YY1 Regulation in Cancer

YY1 expression is regulated by several cancer-associated factors. The E3 ubiquitin ligase Smurf2, the deficiency of which in mice leads to B-cell lymphomas, mediates ubiquitination and degradation of YY1. Smurf2 deficiency enhances YY1-mediated transactivation of c-Myc and B-cell proliferation.⁶⁷ Down-regulation of miR-193a-5p in endometrioid endometrial cancer results in overexpression of YY1, which in turn causes epigenetic silencing of APC through recruitment of EZH2 and trimethylation of histone 3 lysine 27 on its promoter region.⁴⁰ KRAS, the activation of which occurs in more than 90% of pancreatic ductal adenocarcinoma, acts through inflammatory NF-κB signaling to activate YY1, which in turn represses the expression of the tumor suppressor miR-489.⁶⁸ Petrella and Brincherhoff⁶⁹ reported that PTEN inhibits the expression of YY1, thereby inducing HIF-2α transcriptional activity and promoting early renal tumorigenesis resulting from VHL inactivation. The carboxyl-terminal Src kinase homologous kinase down-regulates CXCR4 through YY1, leading to decreased CXCR4-mediated breast cancer cell motility and migration.⁷⁰ The miR-7 and miR-34 family can bind to the 3′ untranslated region of the YY1 mRNA and cause translational repression.^62,63,71

E. YY1 Target Genes in Cancer

YY1 also regulates the expression and protein stability of many cancer-associated genes. YY1 promotes p27 ubiquitination and interacts physically with p27 in breast cancer cell lines.⁶¹ Somewhat in contrast to its tumor-promoting role, YY1 binds to the BRCA1 promoter and increases its expression and that of other downstream genes in breast cancer.⁴⁹ YY1 enhances AP-2α transcriptional activation of the ERBB2 promoter through an AP-2 site.⁷² YY1 interacts physically with the suppressor of zeste 12 (SUZ12) and acts as a mediator to recruit the polycomb proteins and DNA methyltransferases to participate in gene silencing of the tumor suppressor CEBPD.⁷³ One study suggests that the oncogenic effects of YY1 are linked to p53 inhibition and modulation of its downstream effectors such as p15, caspase cascades, c-Jun, and the Wnt signaling pathway.⁶² Silencing YY1 in esophageal squamous cell carcinoma TE-1 cells enhances binding of p21 to cyclin D1 and CDK4, whereas overexpressing YY1 stimulates HO-1 expression in esophageal cancer cells.⁵¹ Knocking down YY1 inhibits, whereas overexpression activates, the Wnt/β-catenin pathway in GAC.⁴¹ YY1 upregulates the levels of the pluripotency genes CD44, Oct4, SOX-2, and Nanog in SC-M1 cells.⁶³ Gene expression analysis of human HBV-associated HCC specimens showed concordant overexpression of YY1 and EZH2, which was correlated with poor patient survival.

Knock-down of YY1 reduces global H3K27me3 levels, leading to upregulation of tumor-suppressive miR-9 isoforms in HCC cells.⁷⁴ The CCAAT/enhancer-binding protein alpha (CEBPA), an important regulator of differentiation in hepatocytes, is a direct target downregulated by YY1.⁴³ YY1 regulates miR-9 transcription negatively in melanoma cells.⁶⁴ YY1 complexes with RelA, which is essential to repress a pro-apoptotic gene, Bim, transcriptionally. Depletion of YY1 or RelA impairs the colony-forming ability of MM progenitor cells.⁶⁵ Silencing YY1 in osteosarcoma cells reduces the expression of RIZ1, which is usually high in osteosarcoma.⁷⁵ Interestingly, in PDAC patients, YY1 tumor expression is correlated positively with better outcome. Experimentally, driving this YY1-mediated suppression of pancreatic cancer cell invasion and metastasis is by down-regulation of MMP10 via a MUC4/ErbB2/p38/MEF2C-dependent mechanism.⁴⁶ In AR-negative prostate cancer cells, YY1 interacts with HDAC4 and gets it recruited to the HOXB13 promoter, causing HOXB13 repression through an epigenetic mechanism.⁷⁶ Overexpression of YY1 causes repression of C/EBPα and the inhibition of C/EBPα leads to the suppression of miR-34a in RCC.⁶⁶ Ishii et al. reported that YY1 can regulate the growth of multiple malignant cell types negatively. YY1 overexpression in MCF7 cells reduces the protein levels of PCNA, a marker of cell growth. In tumors from 13762 MAT rat mammary adenocarcinoma cells, YY1 overexpression reduces pRb^{Ser249/Thr252} expression with a concomitant reduction in PCNA.⁵⁴ YY1 silencing in osteosarcoma cells alters the expression pattern of VEGF isoforms, resulting in an inactive cascade on their receptor, VEGFR2. Moreover, YY1 forms a complex with HIF-1α at the regulatory regions of VEGF-A, VEGF-B, and VEGF-C, acting as an activator in all of them.⁷⁷ We found recently that YY1 interacts with GON4L in bladder cancer cell lines. GON4L-YY1 interaction directs YY1 to bind with the AR and activates the transcriptional activity of AR, which in turn activates the oncogenic protein CD24.³⁸

IV. CONCLUSION AND FUTURE DIRECTIONS

Since its discovery in 1991, YY1 has been progressively implicated in cancer biology either as a biomarker, regulator, or effector of the malignant phenotype. It is a ubiquitous transcription factor that serves a dual role as both an activator and a repressor. Many studies suggest an oncogenic role of YY1 in various cancer types. It is found to be overexpressed in patient tumor samples and cancer cell lines, although its expression does not always correlate with worse prognosis; a few studies showed that elevated levels are correlated with better survival. Despite its context-dependent role in cancer progression, YY1 may be a good prognostic marker and its interactions with other cancer-associated factors can be exploited to develop novel targeted therapies.

ABBREVIATIONS

BMP

bone morphogenic protein

GAC

gastric adenocarcinoma

HCC

hepatocellular carcinoma

HDAC

histone deacetylase complex

MM

multiple myeloma

NF-κB

nuclear factor kappa B

PDAC

pancreatic ductal adenocarcinoma

PSA

prostate specific antigen

Rb

retinoblastoma protein

RCC

renal cell carcinoma