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Tumor Biology
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Introduction
Grainyhead is a transcription factor that is necessary for 
the correct morphology and characteristic of epithelial tis-
sues during Drosophila embryogenesis.1–3 It is also known 
as Brother-of-Mammalian grainyhead (BOM)4 and tran-
scription factor cellular promoter 2-like 3 (TFCP2L3).5 
Disruption of the gene function is embryonic lethal.3 This 
protein family of transcription factors is conserved from 
fly to human. Grainyhead-like 1 (GRHL1), grainyhead-
like 2 (GRHL2), and grainyhead-like 3 (GRHL3) are iden-
tified as homologs of grainyhead in human.4,6,7
The gene for GRHL2 is mapped to human chromosome 
8q22.5 GRHL2 expression can be detected in skin, lung, 
esophagus, and kidney during murine embryogenesis. In 
human adults, GRHL2 is expressed in placenta, brain, kid-
ney, prostate, thymus, lung, salivary gland, mammary 
gland, digestive tract, and pancreas.4,5
Three isoforms of GRHL2 have been identified so far. 
Isoform 1 represents the full length of GRHL2; the transla-
tion of isoform 2 starts at an alternative site which results 
in a protein that is five amino acids shorter than isoform 1 
in addition to six amino acids substitution at N-terminal. 
Compared with isoform 1, isoform 3 lacks 98 amino acids 
at N-terminal resulting in the loss of transcription activity. 
All three isoforms are expressed in pancreas and brain. In 
addition, isoform 1 is detected in testis, prostate, placenta, 
colon, kidney, and lymph node, while isoform 2 is detected 
in tonsil, thymus, placenta, and kidney, and isoform 3 is 
detected in testis.7
Grainyhead-like 2 in development 
and cancer
Lijun Ma1, Hongli Yan3, Hui Zhao4 and Jianmin Sun2,5,6
Abstract
Grainyhead-like 2 is a human homolog of Drosophila grainyhead. It inhibits epithelial-to-mesenchymal transition that is 
necessary for cell migration, and it is involved in neural tube closure, epithelial morphogenesis, and barrier formation 
during embryogenesis by regulation of the expression of cell junction proteins such as E-cadherin and vimentin. Cancer 
shares many common characters with development such as epithelial-to-mesenchymal transition. In addition to its 
important role in development, grainyhead-like 2 is implicated in carcinogenesis as well. However, the reports on 
grainyhead-like 2 in various cancers are controversial. Grainyhead-like 2 can act as either a tumor suppressor or an 
oncogene with the mechanisms not well elucidated. In this review, we summarized recent progress on grainyhead-like 
2 in development and cancer in order to get an insight into the regulation network of grainyhead-like 2 and understand 
the roles of grainyhead-like 2 in various cancers.
Keywords
E-cadherin, epithelial-to-mesenchymal transition, metastasis, oncogene, tumor suppressor
Date received: 17 August 2016; accepted: 24 December 2016
1 Department of Oncology, Tongren Hospital, School of Medicine, 
Shanghai Jiaotong University, Shanghai, P.R. China
2 Department of Pathogen Biology and Immunology, School of Basic 
Medical Sciences, Ningxia Medical University, Yinchuan, P.R. China
3 Department of Laboratory Medicine, Changhai Hospital, The Second 
Military Medical University, Shanghai, P.R. China
4 School of Biomedical Sciences, Faculty of Medicine, the Chinese 
University of Hong Kong, Hong Kong
5 Division of Translational Cancer Research, Department of Laboratory 
Medicine, Lund University, Lund, Sweden
6 Lund Stem Cell Center, Department of Laboratory Medicine, Lund 
University, Lund, Sweden
Corresponding author:
Jianmin Sun, Department of Pathogen Biology and Immunology, School 
of Basic Medical Sciences, Ningxia Medical University, No. 1160 Shengli 
Street, Yinchuan 750004, P.R. China. 
Email: jianmin.sun@med.lu.se; jianmin.sun@nxmu.edu.cn
698375 TUB0010.1177/1010428317698375Tumor BiologyMa et al.
research-article2017
Review
2 Tumor Biology 
Studies in Drosophila indicated that the grainyhead pro-
teins can form dimers through its c-terminal and the dimer 
can stabilize its interaction with DNA. The DNA binding 
domain localizes in the middle of grainyhead, and the 
domain with transcription activity is close to the 
N-terminal.7–9 The alignment of GRHL2 with grainyhead 
suggests similar distribution of the three function domains 
in GRHL27 (Figure 1).
GRHL2 is necessary for neural tube 
closure during embryogenesis
Neural tube closure is a crucial step in the development of 
central nervous system during embryogenesis. The closure 
starts at the caudal hindbrain/rostral spinal cord and con-
tinues through the head and goes down along the spinal 
cord. Failure of neural tube closure leads to neural tube 
defects10 which is one of the most common serious birth 
defects.
The non-neural ectoderm is crucial for full neural tube 
closure.11,12 It can influence the patterning of the dorsal 
neural tube13 and induces neural crest cells.14 GRHL2 is 
expressed in the non-neural ectoderm. It promotes the 
expression of key epithelial genes and suppresses epithe-
lial-to-mesenchymal transition (EMT) to keep the epithe-
lial nature of the non-neural ectoderm during the formation 
of neural tube. Mutation of GRHL2 leads to the loss of 
non-neural ectoderm integrity and abnormal mesenchymal 
phenotype in non-neural ectoderm by downregulation of 
EMT suppressors Esrp1, Sostdc1, Fermt1, Tmprss2, and 
Lamc2.15 In addition, loss of GRHL2 function inhibits the 
expression of E-Cadherin16 that is important to keep the 
epithelial phenotype of cells and enhances vimentin 
expression, leading to impaired cellular dynamics that 
affects the non-neural ectoderm and neural crest cells.15 
Knockout of GRHL2 is embryonic lethal and the embryos 
die at E11.5. In the GRHL2 knockout murine embryos, the 
posterior neuropore keeps open and a fully penetrant split-
face malformation and cranioschisis is observed.17
Expression of GRHL2 during neural tube closure is 
regulated by tet methylcytosine dioxygenase 1 (TET1).18 
TET1 catalyzes the conversion of 5-methylcytosine (5 mC) 
to 5-hydroxymethylcytosine (5 hmC) and regulates gene 
expression.19 Mutation of TET1 leads to enzyme activity 
loss in tuft embryo heads which results in reduced expres-
sion of multiple genes including GRHL2 that are associ-
ated with neural tube closure.18
GRHL2 regulates junction formation 
of epithelial cells
The formation of an apical junctional complex including 
adherens junctions and tight junctions is necessary for the 
definition of cell polarity, junction formation, and differen-
tiation of epithelial cells during embryogenesis. The regu-
lation of junction formation by GRHL2 is widely studied 
in various epithelial cells in addition to its necessary role 
in neural tube closure.
GRHL2 can activate the transcription and regulate the 
expression of the adherens junction gene E-cadherin and 
the tight junction genes claudin 3 (Cldn3) and claudin 4 
(Cldn4) in several types of epithelia, and it is crucial for 
epithelial differentiation.20 GRHL2 also upregulates 
Rab25 that enhances the localization of Cldn4 in the tight 
junction.21
GRHL2 is involved in the regeneration of a polarized 
mucociliary epithelium from basal stemcells in pseu-
dostratified airway epithelium of the lung. It upregulates the 
expression of both notch and ciliogenesis genes (Mcidas, 
Rfx2, and Myb), and promotes organoid morphogenesis and 
the differentiation of ciliated cells.22 Furthermore, GRHL2 
is the key regulator of cell morphogenesis, adhesion, and 
motility of human airway epithelium in adults.23 GRHL2 
and NK2-homeobox 1 transcription factor (Nkx2-1) can 
enhance the expression of each other and they form a posi-
tive feedback loop in lung epithelial cells. Downregulation 
of GRHL2 and Nkx2-1 results in morphological changes of 
flattening lung alveolar type II cells. GRHL2 upregulates 
the expression of semaphorins and their receptors in addi-
tion to E-Cadherin and Cldn4, these genes contribute to 
impaired collective cell migration induced by GRHL2 
mutation.24
Expression of GRHL2 is high in chorionic trophoblast 
cells of placenta. Loss of GRHL2 leads to deficiency of 
basal chorionic trophoblast cell polarity, basement mem-
brane integrity at the chorioallantoic interface, and labyrinth 
branching morphogenesis. GRHL2 can activate the tran-
scription of genes that regulate placental development and 
epithelial morphogenesis, including serine peptidase inhibi-
tor Kunitz type 1 (Spint1) that mediates basal chorionic 
trophoblast (BCT) cell integrity and labyrinth formation.25
Kidney is another organ that has high expression of 
GRHL2. High expression of GRHL2 is detected in nephric 
duct, ureteric bud, and collecting duct epithelia in kidney. 
Loss of GRHL2 function results in deficient nephric duct 
lumen expansion and impaired epithelial barrier formation 
and inhibited lumen expansion in collecting duct epithelia 
in mice. GRHL2 activates the expression of the ovo-like 2 
Figure 1. Schematic structure of human GRHL2 and 
Drosophila grainyhead.
AD: activation domain; DBD: DNA binding domain; DD: Dimerization 
domain.
Ma et al. 3
zinc finger transcription factor (Ovol2), which controls 
Cldn4 and Rab25 expression. The two genes promote 
lumen expansion and barrier formation in subtypes of epi-
thelia in kidney.26
In keratinocytes, GRHL2 directly binds to the promotor 
of p63 and promotes expression of p63. The p63 keeps the 
epithelial phenotype and inhibits the EMT of keratinocytes.27 
Expression of GRHL2 is inhibited by miR-217 in keratino-
cytes, and miR-217 promotes the differentiation of keratino-
cytes by downregulating GRHL2.28 Germline GRHL2 
mutation leads to autosomal-recessive ectodermal dysplasia 
syndrome characterized by nail dystrophy or nail loss, mar-
ginal palmoplantar keratoderma, hypodontia, enamel hypo-
plasia, oral hyperpigmentation, and dysphagia. The mutation 
alters cell morphology and attenuates tight junctions, and 
induces adhesion defects and cytoplasmic translocation of 
GRHL2 from nucleus in keratinocytes.29
GRHL2 in hearing loss
Correlation between GRHL2 and hearing loss was also 
studied. Mutations in GRHL2 can cause progressive auto-
somal dominant hearing loss,5 and that can be recapitu-
lated in zebrafish carrying GRHL2B (homolog of human 
GRHL2 in zebrafish) mutation.30 Single nucleotide poly-
morphisms of GRHL2 are associated with noise-induced 
hearing loss31,32 and age-related hearing loss;33 however, 
another study showed no correlation between single nucle-
otide polymorphisms of GRHL2 and age-related hearing 
loss.34 The association between single nucleotide polymor-
phisms of GRHL2 and noise-induced hearing loss needs to 
be further confirmed.
GRHL2 inhibits metastasis of cancer
EMT is a process that epithelial cells lose cell–cell adhe-
sion and cell polarity, and gain migratory and invasive abil-
ities. During embryogenesis, cells such as neural crest cells 
undergo EMT and migrate to various sites and contribute to 
the formation of various organs. EMT occurs under patho-
logical situations as well; cancer cells migrate from their 
primary sites and colonize at other sites after EMT, leading 
to the recurrence and treatment failure.35 Due to its regula-
tion on cell junction proteins and EMT in development, the 
role of GRHL2 in cancer metastasis was studied.
Zinc finger E-box binding homeobox 1 (ZEB1) is a tran-
scription factor that can downregulate or upregulate the 
expression of its target genes by recruiting co-suppressors 
or co-activators.36,37 ZEB1 promotes EMT and cancer 
metastasis by inhibiting the expression of E-Cadherin that 
is important to keep cell adhesion.38,39 High expression of 
ZEB1 is associated with low expression of E-Cadherin and 
advanced diseases of many tumors.40–43 GRHL2 can sup-
press EMT by inhibition of ZEB1 expression via direct 
repression of the ZEB1 promoter or by impairment of the 
Six1 and DNA complex in breast cancer. Over expression of 
GRHL2 induces mesenchymal-to-epithelial transition 
(MET) of breast cancer cells.44,45 GRHL2 expression is lost 
in the invasive front of primary tumors of breast cancer, and 
the loss of GRHL2 expression is associated with lymph 
node metastasis of breast cancer.46 Loss of GRHL2 expres-
sion promotes the migration of epithelial ovarian cancer 
cells, and it correlates with the mesenchymal molecular 
subtype and a poor overall survival of epithelial ovarian 
cancer patients. GRHL2 directly activates the transcription 
of microRNA-200b/a which regulates the epithelial status 
of epithelial ovarian cancer through inhibition of the 
expression of ZEB1 and E-cadherin.47 ZEB1 can inhibit 
expression of GRHL2 as well;45 GRHL2 and ZEB1 form a 
negative feedback loop.
Partial EMT leads to a hybrid of epithelial/mesenchy-
mal cells which migrate collectively during gastrulation 
and wound healing. In addition to the direct regulation on 
EMT, GRHL2 can stabilize hybrid epithelial/mesenchy-
mal phenotype of lung cancer cells which is associated 
with aggressive tumor progression.48
Transforming growth factor-β (TGF-β) is a cytokine that 
can act as both oncogene and tumor suppressor. It inhibits 
tumor growth by inducing cell cycle arrest and it induces 
EMT to promote tumor metastasis.49,50 TGF-β mediated 
EMT can be suppressed by GRHL2 by inhibition of 
Smad2/3 activation, downregulation of ZEB1 expression, 
and upregulation of mir-200b/c and bone morphogenetic 
protein 2 (BMP2) expression.44 GRHL2 expression is regu-
lated by TGF-β as well. TGF-β and Wnt can downregulate 
GRHL2 expression through ZEB1.45
GRHL2 enhances carcinogenesis
As a tumor suppressor, GRHL2 can inhibit EMT and can-
cer metastasis, and it is downregulated in many cancers. 
However, many studies showed that GRHL2 acts as an 
oncogene and it can enhance the proliferation of tumor 
cells. Overexpression of GRHL2 has been observed in 
several cancers such as oral squamous cell carcinoma,51 
colorectal cancer,52,53 and hepatocellular carcinoma.54 
High expression of GRHL2 enhances proliferation of can-
cer cells, leading to bad prognosis.
The mechanism that GRHL2 promotes cell prolifera-
tion has been studied. Knockdown of GRHL2 in oral squa-
mous cell carcinoma cells results in reduced expression of 
human telomerase reverse transcriptase (hTERT), telomer-
ase activity, and cell survival.55 GRHL2 can inhibit the 
DNA methylation at the 5′-CpG Island in normal keratino-
cytes, resulting in the increase of hTERT expression and 
enhanced cell proliferation.56 This finding proves the 
important role of hTERT in the regulation of GRHL2-
mediated cell proliferation. Furthermore, GRHL2 sup-
presses death-receptor fas cell surface death receptor 
(FAS) expression and FAS induced apoptosis of cancer 
4 Tumor Biology 
cells,57 and it can promote the proliferation of cancer cells 
by upregulation of other oncogenes such as erb-b2 recep-
tor tyrosine kinase 3 (ERBB3).46
GRHL2 regulates the expression of cell junction pro-
teins and inhibits EMT and metastasis of cancer cellswhen 
it acts as a tumor suppressor. It still shows the inhibitory 
role in EMT in oral squamous cell carcinoma and breast 
cancer when it acts as an oncogene that can enhance can-
cer cell proliferation and survival,51,58 suggesting that the 
inhibition of EMT by GRHL2 and its contribution to cell 
proliferation might go through two independent pathways. 
Further studies are necessary to demonstrate the interac-
tion between the regulation of EMT and cell proliferation 
by GRHL2 in order to well understand the opposite roles 
of GRHL2 in different cancers. The regulation network of 
GRHL2 is summarized in Figure 2.
Molecules that are upstream or downstream of GRHL2 
in various cells are summarized together: red color, the 
molecule is an antagonist of EMT; blue color, the molecule 
is an agonist of EMT; green color, the molecule enhances 
cell proliferation; purple color, the molecule is related to 
cell survival.
Conclusion
GRHL2 is a key transcription factor in development. By 
regulation on the expression of cell junction proteins and 
inhibition of EMT, GRHL2 plays a crucial role in neural 
tube closure and epithelial cell morphology. In line to its 
inhibitory role in EMT in development, GRHL2 can 
inhibit EMT of cancer cells and cancer metastasis as well, 
and it is considered as a tumor suppressor in this context. 
However, GRHL2 can also act as an oncogene by promot-
ing cell proliferation and survival. In agreement with the 
opposite roles of GRHL2 in various cancers, the expres-
sion of GRHL2 can be upregulated or downregulated in 
various tumors compared with normal tissues.59 The con-
troversial roles of GRHL2 in different cancers could be 
explained by the different cellular context of host cells. It 
is possible that the cellular context of host cells decides 
the availability of the signaling pathways that regulate 
EMT and cell proliferation by GRHL2, making GRHL2 
behave as tumor suppressor or oncogene in different can-
cers. As an oncogene, GRHL2 could be a potential treat-
ment target by inhibition of its activity. However, the 
tumor suppressing function of GRHL2 in certain types of 
cancers makes it complicated to target GRHL2 in cancer. 
GRHL2 is not widely studied in cancer so far, more stud-
ies are needed to well elucidate the role of GRHL2 in can-
cer and the knowledge could contribute to a better therapy 
of cancer.
Declaration of conflicting interests
The author(s) declared no potential conflicts of interest with 
respect to the research, authorship, and/or publication of this 
article.
Figure 2. The regulation network of GRHL2.
Ma et al. 5
Funding
The author(s) disclosed receipt of the following financial support 
for the research, authorship, and/or publication of this article: The 
work is supported by Ningxia Medical University Foundation (No. 
XZ2016001), Ningxia High Education Scientific Research Program 
(No. NGY2016120), National Natural Science Foundation of 
China (No. 81472770), and Science and Technology Commision of 
Shanghai Changning District (No. CNKW 2014102).
References
 1. Hemphala J, Uv A, Cantera R, et al. Grainy head controls 
apical membrane growth and tube elongation in response 
to branchless/FGF signalling. Development 2003; 130(2): 
249–258.
 2. Mace KA, Pearson JC and McGinnis W. An epidermal bar-
rier wound repair pathway in Drosophila is mediated by 
grainy head. Science 2005; 308(5720): 381–385.
 3. Bray SJ and Kafatos FC. Developmental function of Elf-
1: an essential transcription factor during embryogenesis in 
Drosophila. Genes Dev 1991; 5(9): 1672–1683.
 4. Wilanowski T, Tuckfield A, Cerruti L, et al. A highly con-
served novel family of mammalian developmental tran-
scription factors related to Drosophila grainyhead. Mech 
Dev 2002; 114(1–2): 37–50.
 5. Peters LM, Anderson DW, Griffith AJ, et al. Mutation of 
a transcription factor, TFCP2L3, causes progressive auto-
somal dominant hearing loss, DFNA28. Hum Mol Genet 
2002; 11(23): 2877–2885.
 6. Kudryavtseva EI, Sugihara TM, Wang N, et al. Identification 
and characterization of grainyhead-like epithelial transacti-
vator (GET-1), a novel mammalian grainyhead-like factor. 
Dev Dyn 2003; 226(4): 604–617.
 7. Ting SB, Wilanowski T, Cerruti L, et al. The identification and 
characterization of human sister-of-mammalian grainyhead 
(SOM) expands the grainyhead-like family of developmental 
transcription factors. Biochem J 2003; 370(Pt 3): 953–962.
 8. Uv AE, Thompson CR and Bray SJ. The Drosophila tissue-
specific factor grainyhead contains novel DNA-binding and 
dimerization domains which are conserved in the human 
protein CP2. Mol Cell Biol 1994; 14(6): 4020–4031.
 9. Attardi LD and Tjian R. Drosophila tissue-specific tran-
scription factor NTF-1 contains a novel isoleucine-rich acti-
vation motif. Genes Dev 1993; 7(7B): 1341–1353.
 10. Wilde JJ, Petersen JR and Niswander L. Genetic, epige-
netic, and environmental contributions to neural tube clo-
sure. Annu Rev Genet 2014; 48: 583–611.
 11. Hackett DA, Smith JL and Schoenwolf GC. Epidermal 
ectoderm is required for full elevation and for convergence 
during bending of the avian neural plate. Dev Dyn 1997; 
210(4): 397–406.
 12. Ybot-Gonzalez P, Cogram P, Gerrelli D, et al. Sonic hedge-
hog and the molecular regulation of mouse neural tube clo-
sure. Development 2002; 129(10): 2507–2517.
 13. Dickinson ME, Selleck MA, McMahon AP, et al. 
Dorsalization of the neural tube by the non-neural ectoderm. 
Development 1995; 121(7): 2099–2106.
 14. Selleck MA and Bronner-Fraser M. Origins of the avian 
neural crest: the role of neural plate-epidermal interactions. 
Development 1995; 121(2): 525–538.
 15. Ray HJ and Niswander LA. Grainyhead-like 2 downstream 
targets act to suppress epithelial-to-mesenchymal transition 
during neural tube closure. Development 2016; 143(7): 
1192–1204.
 16. Pyrgaki C, Liu A and Niswander L. Grainyhead-like 2 regu-
lates neural tube closure and adhesion molecule expression 
during neural fold fusion. Dev Biol 2011; 353(1): 38–49.
 17. Rifat Y, Parekh V, Wilanowski T, et al. Regional neural 
tube closure defined by the Grainy head-like transcription 
factors. Dev Biol 2010; 345(2): 237–245.
 18. Fong KS, Hufnagel RB, Khadka VS, et al. A mutation in the 
tuft mouse disrupts TET1 activity and alters the expression 
of genes that are crucial for neural tube closure. Dis Model 
Mech 2016; 9: 585–596.
 19. Tahiliani M, Koh KP, Shen Y, et al. Conversion of 5-methyl-
cytosine to 5-hydroxymethylcytosine in mammalian DNA 
by MLL partner TET1. Science 2009; 324(5929): 930–935.
 20. Werth M, Walentin K, Aue A, et al. The transcription factor 
grainyhead-like 2 regulates the molecular composition of 
the epithelial apical junctional complex. Development 2010; 
137(22): 3835–3845.
 21. Senga K, Mostov KE, Mitaka T, et al. Grainyhead-like 2 
regulates epithelial morphogenesis by establishing func-
tional tight junctions through the organization of a molecu-
lar network among claudin3, claudin4, and Rab25. Mol Biol 
Cell 2012; 23(15): 2845–2855.
 22. Gao X, Bali AS, Randell SH, et al. GRHL2 coordinates 
regeneration of a polarized mucociliary epithelium from 
basal stem cells. J Cell Biol 2015; 211(3): 669–682.
 23. Gao X, Vockley CM, Pauli F, et al. Evidence for multiple 
roles for grainyhead-like 2 in the establishment and mainte-
nance of human mucociliary airway epithelium. Proc Natl 
Acad Sci U S A 2013; 110(23): 9356–9361.
 24. Varma S, Cao Y, Tagne JB, et al. The transcription factors 
grainyhead-like 2 and NK2-homeobox 1 form a regulatory 
loop that coordinates lung epithelial cell morphogenesis and 
differentiation. J Biol Chem 2012; 287(44): 37282–37295.
 25. Walentin K, Hinze C, Werth M, et al. A Grhl2-dependent 
gene network controls trophoblast branching morphogen-
esis. Development 2015; 142(6):1125–1136.
 26. Aue A, Hinze C, Walentin K, et al. A grainyhead-like 2/ovo-
like 2 pathway regulates renal epithelial barrier function and 
lumen expansion. J Am Soc Nephrol 2015; 26(11): 2704–2715.
 27. Mehrazarin S, Chen W, Oh JE, et al. The p63 gene is regu-
lated by grainyhead-like 2 (GRHL2) through reciprocal 
feedback and determines the epithelial phenotype in human 
keratinocytes. J Biol Chem 2015; 290(32): 19999–20008.
 28. Zhu H, Hou L, Liu J, et al. MiR-217 is down-regulated 
in psoriasis and promotes keratinocyte differentiation via 
targeting GRHL2. Biochem Biophys Res Commun 2016; 
471(1): 169–176.
 29. Petrof G, Nanda A, Howden J, et al. Mutations in GRHL2 
result in an autosomal-recessive ectodermal Dysplasia syn-
drome. Am J Hum Genet 2014; 95(3): 308–314.
 30. Han Y, Mu Y, Li X, et al. Grhl2 deficiency impairs otic 
development and hearing ability in a zebrafish model of 
the progressive dominant hearing loss DFNA28. Hum Mol 
Genet 2011; 20(16): 3213–3226.
 31. Zhang X, Liu Y, Zhang L, et al. Associations of genetic var-
iations in EYA4, GRHL2 and DFNA5 with noise-induced 
6 Tumor Biology 
hearing loss in Chinese population: a case-control study. 
Environ Health 2015; 14: 77.
 32. Li X, Huo X, Liu K, et al. Association between genetic vari-
ations in GRHL2 and noise-induced hearing loss in Chinese 
high intensity noise exposed workers: a case-control analy-
sis. Ind Health 2013; 51(6): 612–621.
 33. Van Laer L, Van Eyken E, Fransen E, et al. The grainy-
head like 2 gene (GRHL2), alias TFCP2L3, is associated 
with age-related hearing impairment. Hum Mol Genet 2008; 
17(2): 159–169.
 34. Lin YH, Wu CC, Hsu CJ, et al. The grainyhead-like 2 gene 
(GRHL2) single nucleotide polymorphism is not associ-
ated with age-related hearing impairment in Han Chinese. 
Laryngoscope 2011; 121(6): 1303–1307.
 35. Acloque H, Adams MS, Fishwick K, et al. Epithelial-
mesenchymal transitions: the importance of changing 
cell state in development and disease. J Clin Invest 2009; 
119(6): 1438–1449.
 36. Postigo AA, Depp JL, Taylor JJ, et al. Regulation of Smad 
signaling through a differential recruitment of coactivators 
and corepressors by ZEB proteins. EMBO J 2003; 22(10): 
2453–2462.
 37. Eger A, Aigner K, Sonderegger S, et al. DeltaEF1 is a tran-
scriptional repressor of E-cadherin and regulates epithelial 
plasticity in breast cancer cells. Oncogene 2005; 24(14): 
2375–2385.
 38. Remacle JE, Kraft H, Lerchner W, et al. New mode of DNA 
binding of multi-zinc finger transcription factors: deltaEF1 
family members bind with two hands to two target sites. 
EMBO J 1999; 18(18): 5073–5084.
 39. Peinado H, Olmeda D and Cano A. Snail, Zeb and bHLH 
factors in tumour progression: an alliance against the epi-
thelial phenotype? Nat Rev Cancer 2007; 7(6): 415–428.
 40. Bronsert P, Kohler I, Timme S, et al. Prognostic signifi-
cance of zinc finger E-box binding homeobox 1 (ZEB1) 
expression in cancer cells and cancer-associated fibroblasts 
in pancreatic head cancer. Surgery 2014; 156(1): 97–108.
 41. Sahay D, Leblanc R, Grunewald TG, et al. The LPA1/
ZEB1/miR-21-activation pathway regulates metastasis 
in basal breast cancer. Oncotarget 2015; 6(24): 20604–
20620.
 42. Zhou YM, Cao L, Li B, et al. Clinicopathological signifi-
cance of ZEB1 protein in patients with hepatocellular carci-
noma. Ann Surg Oncol 2012; 19(5): 1700–1706.
 43. Zhang GJ, Zhou T, Tian HP, et al. High expression of ZEB1 
correlates with liver metastasis and poor prognosis in colo-
rectal cancer. Oncol Lett 2013; 5(2): 564–568.
 44. Cieply B, Riley P, Pifer PM, et al. Suppression of the epithe-
lial-mesenchymal transition by grainyhead-like-2. Cancer 
Res 2012; 72(9): 2440–2453.
 45. Cieply B, Farris J, Denvir J, et al. Epithelial-mesenchymal 
transition and tumor suppression are controlled by a 
 reciprocal feedback loop between ZEB1 and grainyhead-
like-2. Cancer Res 2013; 73(20): 6299–6309.
 46. Werner S, Frey S, Riethdorf S, et al. Dual roles of the tran-
scription factor grainyhead-like 2 (GRHL2) in breast can-
cer. J Biol Chem 2013; 288(32): 22993–23008.
 47. Chung VY, Tan TZ, Tan M, et al. GRHL2-miR-200-ZEB1 main-
tains the epithelial status of ovarian cancer through transcriptional 
regulation and histone modification. Sci Rep 2016; 6: 19943.
 48. Jolly MK, Tripathi SC, Jia D, et al. Stability of the hybrid 
epithelial/mesenchymal phenotype. Oncotarget 2016; 
7(19): 27067–27084.
 49. Massague J. TGFbeta in cancer. Cell 2008; 134(2): 215–230.
 50. Taylor MA, Parvani JG and Schiemann WP. The patho-
physiology of epithelial-mesenchymal transition induced 
by transforming growth factor-beta in normal and malig-
nant mammary epithelial cells. J Mammary Gland Biol 
Neoplasia 2010; 15(2): 169–190.
 51. Chen W, Yi JK, Shimane T, et al. Grainyhead-like 2 regu-
lates epithelial plasticity and stemness in oral cancer cells. 
Carcinogenesis 2016; 37: 500–10.
 52. Quan Y, Xu M, Cui P, et al. Grainyhead-like 2 promotes 
tumor growth and is associated with poor prognosis in colo-
rectal cancer. J Cancer 2015; 6(4): 342–350.
 53. Quan Y, Jin R, Huang A, et al. Downregulation of GRHL2 
inhibits the proliferation of colorectal cancer cells by target-
ing ZEB1. Cancer Biol Ther 2014; 15(7): 878–887.
 54. Tanaka Y, Kanai F, Tada M, et al. Gain of GRHL2 is asso-
ciated with early recurrence of hepatocellular carcinoma. 
J Hepatol 2008; 49(5): 746–757.
 55. Kang X, Chen W, Kim RH, et al. Regulation of the hTERT 
promoter activity by MSH2, the hnRNPs K and D, and 
GRHL2 in human oral squamous cell carcinoma cells. 
Oncogene 2009; 28(4): 565–574.
 56. Chen W, Dong Q, Shin KH, et al. Grainyhead-like 2 
enhances the human telomerase reverse transcriptase gene 
expression by inhibiting DNA methylation at the 5′-CpG 
island in normal human keratinocytes. J Biol Chem 2010; 
285(52): 40852–40863.
 57. Dompe N, Rivers CS, Li L, et al. A whole-genome RNAi 
screen identifies an 8q22 gene cluster that inhibits death 
receptor-mediated apoptosis. Proc Natl Acad Sci U S A 
2011; 108(43): E943–E951.
 58. Xiang X, Deng Z, Zhuang X, et al. Grhl2 determines the 
epithelial phenotype of breast cancers and promotes tumor 
progression. PLoS ONE 2012; 7(12): e50781.
 59. Riethdorf S, Frey S, Santjer S, et al. Diverse expression patterns 
of the EMT suppressor grainyhead-like 2 (GRHL2) in normal 
and tumour tissues. Int J Cancer 2016; 138(4): 949–963.