Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

GLI Family Zinc Finger 2

  • David A. Jackson
  • Jason A. Misurelli
  • Sherine F. Elsawa
Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_101917

Synonyms

Historical Background

GLI2 belongs to the GLI family of genes, encoding Kruppel-like zinc finger transcription factors. Evidence for GLI proteins and their function was first elucidated through the characterization of the Drosophila GLI homolog Cubitus interruptus (Ci), responsible for proper tissue patterning and segmentation (Goodrich et al. 1996). While highly conserved, vertebrates have three different forms of the transcription factor (GLI1, GLI2, and GLI3), each taking on redundant and unique roles in normal and pathologic biology. GLI1, the first of the GLI family characterized in humans, was identified in 1987 as a highly expressed gene in human glioma (Kinzler et al. 1987). Both GLI2 and GLI3 were discovered in 1988 through the use of cloned GLI1 cDNA as a probe targeting similar sequences (Ruppert et al. 1988). The expression of GLI transcription factors in embryonic tissue as well as their upregulation in malignant adult tissues suggested the importance of the GLI factors in development and their role in multiple forms of disease.

Structure and Function

In humans, the GLI2 gene is found on chromosome 2 at 2q14.2 and is 202,363 bp in length. The gene transcript is 6780 bp and contains 13 exons. The full length GLI2 protein (GLIFL) is 1586 amino acids in length; however partial proteasomal degradation results in a truncated form with transcriptional repressor activity (GLI2R). There is an overall 95% amino acid sequence similarity between GLI2 and GLI3, with an 86–87% similarity within the zinc finger region (Hui et al. 1994). Alternative splicing of GLI2 transcript by two independent mechanisms results in four distinct isoforms: GLI2 α, β, γ, and δ. Compared to the full length GLI2, which is 197 kDa, the isoforms are 133, 88, 86, and 131 kDa in size, respectively (Tojo et al. 2003).
GLI Family Zinc Finger 2, Fig. 1

Biochemical and regulatory domains of GLI2. GLI2 contains an N-terminal repressor domain (red), a zinc finger domain (blue), a processing determinant domain (PDD) (orange), and a C-terminal transcriptional activation domain (green). The SuFu-binding site (yellow) binds Suppressor of Fused, and Degrons (DC and DN) influence degradation properties of GLI2

Members of the GLI protein family have five consecutive C2-H2 zinc finger motifs (Fig. 1), which bind the DNA of target genes. These motifs consist of a X3-Cys-X4-Cys-X12-His-X4-His-X3 (X representing any amino acid) pattern that recognize a 5′-GACCACCCA-3′ target DNA sequence (Mccleary-Wheeler 2014). All GLI protein family members also possess a transactivation domain on the C-terminus (Fig. 1). The specialized function of the three GLI proteins arises primarily due to the presence or absence of an N-terminal transcriptional repression domain. GLI1 lacks a repression domain and therefore functions only as a transcriptional activator (GLIA). Both GLI2 and GLI3 possess an N-terminal transcriptional repressor domain, allowing them to be either transcriptional activators or repressors (GLIR). The fate of full length GLI2 is determined by sequential phosphorylation first by Protein Kinase A (PKA), Casein Kinase 1 (CK1), and finally Glycogen Synthase Kinase 3b (GSK3β) within its processing determinant domain (PDD). Hyperphosphorylated GLI2FL is ubiquitinated by a Cul1 E3 ligase, targeting it for partial proteolysis by proteasomes to remove the C-terminal activation domain, resulting in the repressor form which translocates to the nucleus and blocks transcription of target genes. However, complete degradation by proteasomes following ubiquitination by the SPOP-Cul3 E3 ligase is the more common route for GLI2, therefore it is observed to function primarily as an activator while GLI3 serves mostly as a transcriptional repressor (Pak and Segal 2016). Deletion of the PDD domain in GLI2 prevents the formation of GLI2R (Hui and Angers 2011). In its full length form, GLI2 is sequestered in the cytoplasm by Suppressor of Fused (SuFu). Sequestration of GLI2 by SuFu renders GLI2 inactive but does stabilize GLI2, increasing its half-life. For GLI2 to act as a transcriptional activator, it must be released from the SuFu-complex to translocate to the nucleus.

GLI2 activity can be regulated by different posttranslational modifications as well as epigenetically through histone-modifying enzymes. Acetylation of GLI2 on K757 by the histone acetyltransferase CREB-binding protein (CBP) decreases its transcriptional activity by interfering with target gene promoter interaction. Upregulation of Histone Deacetylase 1 (HDAC1) increases GLI1 and GLI2 activity. This pathway is antagonized by an E3 ubiquitin ligase that targets HDAC1 for degradation (Hui and Angers 2011). Sumoylation of GLI2 by PKA at K630 and K716 results in the recruitment of Histone Deacetylase 5 (HDAC5) which in turn inhibits GLI2 transcriptional activity as GLI2 is prevented from localizing to the nucleus (Mccleary-Wheeler 2014). Two dual specificity tyrosine-regulated kinases, DYRK1B and DYRK2, also inhibit GLI2 by targeting it for proteosomal degradation (Hui and Angers 2011).

GLI2 Signaling Pathways

The Hedgehog (HH) signaling pathway has been considered the canonical mechanism which activates GLI proteins. The pathway derives its name from the HH ligand of which three mammalian homologues exist: Sonic (SHH), Desert (DHH), and Indian (IHH). HH binds to Patched (PTCH1), a 12-pass transmembrane protein that inhibits another 7-pass transmembrane protein, Smoothened (SMO). Upon HH binding to PTCH1, both molecules are degraded, releasing the inhibitory effect on SMO. Following phosphorylation by CK1 and G-protein-coupled kinase 2 (GRK2), SMO migrates to the base of the primary cilium via Kif3a, a kinesin motor protein (Pak and Segal 2016). HH binding also initiates the degradation of Gpr161, an activator of PKA. Active SMO further inhibits PKA, preventing the initial phosphorylation of GLI2. Without first being primed by PKA, GSK3β is unable to phosphorylate GLI2, while CK1 phosphorylates GLI2 at multiple alternative sites. SuFu is destabilized and the complex dissociates, releasing GLI2 in its activator form (GLI2A), now able to translocate to the nucleus to initiate transcription of target genes (Gorojankina 2016).

While canonical activation of GLI2 is certainly important, particularly in normal development, GLI2 has been shown to be activated by a handful of noncanonical pathways including KRAS, PI3K/Akt, and TGF-β (Fig. 2). An understanding of additional pathways (other than HH) that regulate GLI2 is important as noncanonical activation is often associated with GLI2 involvement in malignancy.

While activation and interaction of GLI1 with KRAS signaling has been established, little work has shown a relationship between KRAS and GLI2 (Mills et al. 2013). Recently, Wu et al. demonstrated GLI2 being elevated in tumor samples utilizing combined KRAS and androgen receptor (AR) signaling to maintain cell proliferation (Wu et al. 2016). Early studies showed that crosstalk between canonical HH signaling and PI3K/Akt was needed for stabilization and activation of GLI2 (Riobo et al. 2006). More recent evidence suggests GLI2 activation occurring via PI3K/Akt signaling independent of canonical HH signaling (Elsawa et al. 2011). GLI2 was found to be activated in bone marrow (BM) stromal cells upon ligation of CCL5 to its receptor, CCR3. Activation of GLI2 through CCR3 was found to go through PI3K/Akt and NF-κB signaling (Elsawa et al. 2011). Activation of GLI2 by PI3K/Akt signaling independent of HH signaling has been further supported by work done using renal cell carcinoma (RCC) cell lines (Zhou et al. 2016). Overexpression of Akt in RCC cells resulted in increased GLI2 mRNA expression as well as increased GLI activity, and pharmacological inhibition of Akt with perifosine resulted in decreased GLI2 mRNA expression (Zhou et al. 2016). In both of these studies treatment with the SMO inhibitor cyclopamine had no effect on GLI2 expression or GLI-mediated activity (Elsawa et al. 2011; Zhou et al. 2016).
GLI Family Zinc Finger 2, Fig. 2

The major regulatory pathways of GLI2. (a) CCL5/CCR3: Binding of cytokine CCL5 to the CCR3 receptor signals through the PI3K-AKT-NFκ-B pathway, resulting in the transcription of GLI2. (b) TGF-β: Transforming growth factor beta (TGF-β) binds to the TGF-β receptor, initiating the phosphorylation of receptor-regulated SMAD3 and complexes with SMAD4. The SMAD complex translocates to the nucleus where it is bound by β-catenin and initiates transcription of GLI2. (c) Hedgehog Signaling Pathway: The Hedgehog ligand (HH) binds to Patched (PTCH), removing the inhibitory effect on Smoothened (SMO). Active SMO blocks GLI2 degradation and GLI2 repressor (GLI2R) formation by inhibiting phosphorylation by Protein Kinase A (PKA). In its active state, SMO also destabilizes Suppressor of Fused (SuFu), releasing full length GLI2 (GLI2FL) to be converted to its transcriptional activator form (GLI2A). (d) PI3K/AKT: Signaling through PI3K/AKT stabilizes GLI2, enhancing its transcriptional activity. (e) RAS: Epidermal growth factor (EGF) binds to the EGF receptor, initiating a signaling cascade through KRAS-RAF-MEK-ERK, stabilizing GLI2, and enhancing its transcriptional activity

Over the last 10 years, evidence has increased suggesting signaling through TGF-β and SMAD regulates the expression and activity of GLI2 in a number of different malignancies. Treatment of multiple cell lines of different malignant origins with TGF-β resulted in increased GLI2 mRNA and protein expression. Elevated GLI2 expression as a result of TGF-β treatment was found to be independent of HH signaling, and genetic inhibition of the transcription factor SMAD3 resulted in decreased TGF-β-induced expression of GLI2 (Dennler et al. 2007). This mechanism appears to be a result of direct GLI2 promoter manipulation by SMAD3 as there is a SMAD3 binding site located in the GLI2 promoter, and TGF-β-induced GLI2 expression is enhanced by overexpressing SMAD3 (Dennler et al. 2009). This evidence is supported by in vivo data in which mice expressing TGF-β were found to have higher GLI2 protein levels in epidermal samples. However, this effect was abolished in TGF-β-expressing mice with heterozygous SMAD 3 knockout (Dennler et al. 2007). Functionally, regulation of GLI2 expression via TGF-β may aid in melanoma cells becoming resistant to SMO inhibitors such as vemurafenib and result in a more aggressive phenotype. Vemurafenib-resistant cells express higher levels of TGF-β receptors and SMAD2/3. Furthermore, treatment of these vemurafenib-resistant cells with recombinant TGF-β results in both increased GLI2 expression and increased invasiveness (Alexaki et al. 2010; Faiao-Flores et al. 2016).

Roles in Development and Disease

The GLI family of proteins was originally identified due to the importance of the HH signaling pathway in embryonic development. The crucial role played by the GLI genes is evident based on knockout experiments in mice. These models have demonstrated both GLI2 and GLI3 are necessary for proper embryogenesis, while GLI1 is not required. Homozygous deletion of the GLI2 zinc finger domains results in a lethal phenotype in late embryonic stages. These embryos can, however, be rescued with the expression of GLI1 (Mccleary-Wheeler 2014). This illustrates some overlap in GLI protein function, but also complexity in GLI interaction. For example, GLI2 is required for initial HH signaling as GLI1 is a target gene of GLI2 (Hui and Angers 2011). GLI2 serves as the major transcriptional activator in HH signaling, while GLI3 serves as the major repressor. A careful balance of the temporal and spatial expression of GLI2 and GLI3 is necessary to create a gradient, allowing proper anterior/posterior limb patterning and neural development to occur (Hui and Angers 2011). GLI2 is particularly important for dorsal-ventral neural tube patterning in the midbrain and spinal cord, where it regulates the expression of Foxa2 and Nkx-2.2 (Jacob and Briscoe 2003). GLI2 deficiency impairs the development of the floorplate and interneurons, resulting in partial loss of the anterior pituitary and complete loss of the posterior pituitary. GLI2 mutations are associated with brain abnormalities such as holoprosencephaly (HPE), where complete separation of the forebrain fails to occur. Multiple organ systems are affected by loss of GLI2, resulting in defects in the esophagus, lungs, and trachea. GLI2 regulates Runx2, a transcription factor necessary for osteoblastogenesis in bone formation. Physical abnormalities such as cleft palate and polydactyly are associated with GLI2 mutations (Mccleary-Wheeler 2014).

Major target genes of GLI2 are related to cell proliferation, survival, and cell cycle regulation. Direct targets include cell cycle progression genes CDC45L, CDC2, CCNA2, and CCNB1, which are upregulated, while cell cycle inhibition genes, such as CDKN1A, are downregulated (Mccleary-Wheeler 2014). Antiapoptotic genes such as CFLAR and BCL2 have also been shown to be elevated with GLI2 overexpression (Mccleary-Wheeler 2014). GLI2 has less of a developmental role in adult tissues outside the maintenance of certain adult stem cells (Gorojankina 2016). However, GLI2 is still expressed in all adult tissues with the exception of the placenta and most highly expressed in the testes, myometrium, and kidneys (Ruppert et al. 1988). Generally, two major categories of diseases related to GLI2 can be distinguished. Mutations which result in loss of function of GLI2 severely affect embryonic development, leading to congenital disorders as previously highlighted. In adult tissues, gain of function mutations are more relevant as the upregulation of GLI2 has been associated with various malignancies (Mccleary-Wheeler 2014).

Roles in Cancer

As has been discussed above, GLI2 is essential in normal development, and GLI2 knockout in mice results in a lethal phenotype. Not surprisingly, aberrant activation of GLI2 is important in the pathogenesis and progression of a number of different cancers. The list of different malignancies involving GLI2 continues to grow each year. Perhaps more surprising than GLI2 involvement in the biology of these diseases are the different mechanisms by which GLI2 becomes active and the diverse downstream effects of GLI2 activation. In cancer, GLI2 can be activated via canonical HH signaling, noncanonical signaling, or a combination of both.

One cancer in which GLI2 plays a role is basal cell carcinoma (BCC) (Atwood et al. 2013; Atwood et al. 2015; Luongo et al. 2014; Pantazi et al. 2014). BCC is the most common cancer in the United States (Atwood et al. 2015). While often times treatable as what is known as syndromic BCC, on rare occasions it becomes more aggressive and metastatic (Atwood et al. 2015). The SMO inhibitor vismodegib was recently approved by the FDA for treatment of BCC, and all patients with syndromic BCC respond to vismodegib whereas patients with advanced or metastatic BCC only have a response rate of 48% and of those 20% develop resistance to this treatment. This appears to be related to the ability of atypical protein kinase C ι/λ (aPKC- ι/λ) to activate HH effectors GLI1 and GLI2 in the absence of signaling through SMO (Atwood et al. 2013; Atwood et al. 2015). Inhibition of aPKC- ι/λ resulted in decreased viability of BCC cells which had developed SMO inhibitor resistance (Atwood et al. 2013). Furthermore, it was found that mRNA and protein expression of GLI2 were increased in SMO inhibitor resistant BCC cells, and inhibition of GLI2 with arsenic trioxide (ATO) resulted in decreased GLI1 expression (Atwood et al. 2015). Other studies have suggested the importance of GLI2 in BCC and partially elucidated the mechanism by which it is stabilized. One such mechanism is due to a feedback loop in which GLI2 positively regulates the transcription of Type 3 deiodinase (D3), an enzyme which inhibits the activity of the thyroid hormone triiodothyronine (T3) (Luongo et al. 2014). Essential to embryonic development, D3 is not expressed in normal tissues but is found to be elevated in a number of different cancers (Dentice et al. 2009; Dentice et al. 2012; Huang et al. 2000; Kester et al. 2006). In BCC cells, activation of HH signaling by SHH results in increased D3 expression. Subsequently, D3 inactivates T3 which induces PKA which is responsible for marking GLI2 for degradation (Luongo et al. 2014). Treatment of BCC cells with T3 results in decreased GLI activity measured using an 8X GLI luciferase reporter, decreased GLI2 protein levels, and decreased GLI1 and PTCH1 mRNA levels. Interestingly, treatment with T3 does not result in decreased GLI2 mRNA levels, supporting the hypothesis of activation of PKA resulting in increased degradation of GLI2 protein. In further support of this, T3 does not affect GLI2 activity when the degron Dc region, need for signaling leading to degradation, is cloned out. Furthermore, when PKA is inhibited pharmacologically, treatment with T3 does not result in decreased GLI2 activity (Luongo et al. 2014). Finally, not only does GLI2 appear to be involved in the maintenance of BCC but also appears to contribute to the initiation of BCC (Pantazi et al. 2014; Epstein 2008; Grachtchouk et al. 2000). Early experiments showed that overexpressing GLI2 in mice resulted in the development of BCC (Grachtchouk et al. 2000). Furthermore, it was established that GLI2 activation occurred through canonical HH signaling (Epstein 2008). Genomic instability is highly present in BCC, and stable expression of GLI2 without an N-terminal repressor domain (ΔN GLI2) in immortalized newborn keratinocytes results in tetraploidy, polyploidy, and aneuploidy as well as decreased p21 and 14-3-3σ expression and increased Bcl-2 protein levels. Apoptosis is diminished in cells exposed to UVB expressing ΔN GLI2. However, pharmacological inhibition of Bcl-2 in cells expressing ΔN GLI2 sensitizes cells to UVB exposure and reduces tetraploidy and aneuploidy when compared to cells without Bcl-2 inhibition (Pantazi et al. 2014).

GLI2 has also been suggested to play a role in the maintenance and progression of chronic lymphocytic leukemia (CLL). GLI2 mRNA expression was found to be elevated in samples from patients with poor clinical outcomes compared to samples of patients with good clinical outcomes (Hegde et al. 2008). In vitro pharmacological and genetic inhibition of GLI2 results in decreased viability of the CLL cell line MEC1 (Desch et al. 2010). Furthermore, the effect of GLI2 inhibition on survival is enhanced when cells are treated with PI3K/Akt inhibitors dactolisib and pictilisib (Desch et al. 2010). However, GLI2 activity was not involved in the initiation of CLL (Desch et al. 2010).

In colon cancer cell lines inhibition of GLI1/GLI2 using a pharmacological inhibitor, GANT61, resulted in DNA damage and 75% cell death (Agyeman et al. 2014; Agyeman et al. 2012). GLI2 was found to regulate expression of hTERT which functions to prevent telomere shortening in malignant cells, thus increasing replication potential. Overexpression of ΔN GLI2 resulted in increased hTERT mRNA expression and hTERT promoter activity while treatment of cells with GANT61 had an opposite effect on hTERT regulation (Mazumdar et al. 2013). Furthermore, higher expression of GLI2 in patient samples positively correlated with shorter survival times (Singovski et al. 2016). Together these studies suggest a role for GLI2 in the progression of colon cancer.

Recently, GLI2 involvement in osteosarcoma has been fairly extensively characterized. Osteosarcoma results in 60% of all cancer-related deaths in children and has a 5-year survival rate of 60–70% (Sun et al. 2014). It has been shown that GLI2 is expressed, to some degree, in all samples taken from patients with osteosarcoma, GLI2 expression is positively correlated with poor prognosis, and that genetic inhibition of GLI2 results in decreased proliferation and viability of osteosarcoma cells (Nagao et al. 2011; Nagao-Kitamoto et al. 2015a; Nagao-Kitamoto et al. 2015b; Yang et al. 2013). Further studies have uncovered some of the mechanisms by which GLI2 operates in osteosarcoma. GLI2 appears to be negatively regulated by the microRNA miR-202 as overexpression of miR-202 results in decreased proliferation in vitro and less tumor growth of in vivo xenografts (Sun et al. 2014). Furthermore, overexpression of miR-202 results in decreased GLI2 protein expression, and expression of GLI2 without a 3′ UTR which possesses the binding site for miR-202 in a miR-202-expressing background does not result in a decrease in cell proliferation (Sun et al. 2014). Ribosomal protein S3 (RPS3) has been identified as being upregulated in osteosarcoma cell lines compared to normal osteoblasts cells as well as being upregulated in patient samples with lung metastasis in comparison to samples from patients without lung metastasis. It appears GLI2 is, in part, responsible for this upregulation of RPS3 (Nagao-Kitamoto et al. 2015b). Expression of GLI2 as well as expression of RPS3 resulted in increased invasiveness of osteosarcoma cell lines (Nagao-Kitamoto et al. 2015b). Candidate GLI binding sites were identified on the RPS3 promoter, overexpression and knockdown of GLI2 resulted in increased and decreased expression of RPS3, respectively, and expression of RPS3 was able to rescue the effect of genetic inhibition of GLI2 on cell invasion (Nagao-Kitamoto et al. 2015b). Finally, the long coding RNA BCAR4 is evidenced to be a cooperative activator of transcription targeting GLI2 target genes in osteosarcoma (Chen et al. 2016). BCAR4 was found to associate with the promoter of GLI2 target genes RPS3, IL-6, MUC5AC, and TGF-β1. Furthermore, genetic inhibition of either GLI2 or BCAR4 resulted in decreased expression of these GLI2 target genes, and in tissue samples, BCAR4 expression positively correlated with GLI2 expression (Chen et al. 2016). Together, these studies provide strong evidence for a role of GLI2 in the maintenance, progression, and invasive potential of osteosarcoma.

In melanoma, GLI2 mRNA and protein expression are elevated in melanoma cell lines that exhibit an invasive phenotype compared to less aggressive cell lines (Alexaki et al. 2010; Javelaud et al. 2011). Furthermore, GLI2 expression was inversely correlated with Melanogenesis Associated Transcription Factor (MITF) expression, and GLI2 was found to bind to the MITF promoter and negatively regulate MITF expression (Faiao-Flores et al. 2016; Javelaud et al. 2011; Pierrat et al. 2012). Vemurafenib is a BRAF inhibitor used clinically to treat melanoma, and GLI2 is found to be elevated in samples from patients with vemurafenib-resistant melanoma (Faiao-Flores et al. 2016; Sandri et al. 2016). Further, these studies suggest GLI2 is regulated by TGF-β and SMAD signaling and that GLI2 may function as a transcriptional activator of MMP2 and MMP9, two metalloproteinases thought to be involved in invasion and metastasis (Alexaki et al. 2010; Faiao-Flores et al. 2016; Pierrat et al. 2012; Kessenbrock et al. 2010).

GLI2 has been suggested to play a role in a number of other malignancies. In diffuse large B cell lymphoma (DLBCL), GLI2 appears to be active as 73% percent of patient samples were positive for GLI2 expression as evidenced by immunohistochemistry staining (Kim et al. 2009a). Further studies found that genetic inhibition of GLI2 as well as pharmacological inhibition of SMO resulted in decreased cell viability of DLBCL cell lines (Singh et al. 2010). These studies suggest a role for GLI2 in the biology of DLBCL and that GLI2 is activated via canonical HH signaling in this disease. Immunohistochemical analysis of endometrial samples (normal, hyperplastic, neoplastic) showed progressively increasing GLI2 protein expression. Moreover malignant samples had a significantly higher amount of GLI2 localized to the nucleus (Kim et al. 2009b). Hepatocellular carcinoma (HCC) Huh7 cells were sorted and selected using markers of invasive potential (CD133-/EpCAM-) (Fan et al. 2016). Cells with the highest invasive behavior and increased metastatic markers were found to have the highest levels of GLI2 nuclear protein and mRNA expression (Fan et al. 2016). GLI2 was also found to be expressed at high levels in samples from patients with the classical subtype of lung squamous cell carcinoma (LSCC) (Huang et al. 2014). Furthermore, genetic and pharmacological inhibition of GLI2 resulted in decreased LSCC cell survival in vitro and decreased tumor growth in vivo (Huang et al. 2014). GLI2 has been shown to be expressed in prostate cancer tissue samples as well as representative prostate cancer cell lines (Narita et al. 2008). Using a GLI2 antisense oligonucleotide, Narita et al. reported decreased cell viability in vitro, decreased tumor growth in vivo, and decreased protein expression of Bcl2, Bcl-XL, cyclin D1, p-Rb, and PKC-β (Narita et al. 2008). However, it was more recently shown that genetic inhibition of GLI2 in prostate cancer cells did not result in decreased proliferation but rather resulted in decreased expression of p63 which in turn regulates the renewal potential of progenitor cells (Wu et al. 2016). Furthermore, Wu et al. show that KRAS signaling in association with androgen receptor signaling results in activation of GLI1 and GLI2 in prostate cancer cells (Wu et al. 2016). In pancreatic cancer, selective inhibitors of BET bromodomain proteins are currently being used for treatment resulting in repression of MYC activity (Kumar et al. 2015). In cells resistant to the BET bromodomain protein inhibitor JQ1, MYC activation continues via GLI2, and genetic inhibition of GLI2 restores sensitivity to JQ1 (Kumar et al. 2015). In renal cell carcinoma (RCC), increased expression of GLI2 is found in RCC patient samples compared to healthy kidney samples, and high GLI2 expression negatively correlates with overall survival (Zhou et al. 2016). Further, transfection of a normal kidney cell line with a GLI2 expression construct resulted in increased proliferation and tumor formation in vivo, and it appears that GLI2 activation occurs via Akt signaling rather than canonical HH signaling (Zhou et al. 2016).

Recently, a role for GLI2 was demonstrated in the biology of the IgM secreting hematological malignancy Waldenström macroglobulinemia (WM) (Jackson et al. 2015). Prior to this study a role for GLI had not been determined in WM; however, GLI2 had been shown to regulate the WM tumor microenvironment (Elsawa et al. 2011). Overproduction of IgM is a hallmark of WM and results in the majority of symptoms caused by the disease. GLI2 was found to regulate the expression of IL-6 in BM stromal cells, which in turn resulted in higher levels of IgM secretion by WM cells. Upon investigating a role for GLI in WM cells, a complementary role for GLI2 was found where GLI2 directly regulates the expression of the IL-6 receptor α subunit (IL-6Rα) which also results in increased IgM secretion by WM cells (Jackson et al. 2015). Thus GLI2 plays a twofold role in the biology of WM both in the stromal and malignant compartments.

Due to their implication in tumorigenesis, the GLI transcription factors have been considered targets for cancer therapies. While many upstream inhibitors have been discovered, GLI-specific pharmacological agents have also been identified. Cyclohexyl-methyl aminopyrimidines (CMAPs) have been found to be indirect inhibitors of GLI1 and GLI2 by activating the mitogen-activated protein kinase (MAPK) pathway. JQ1 and I-BET151 interfere with GLI target gene recruitment by antagonizing BDR4, a bromo and extra C-terminal (BET) family protein able to modulate GLI1- and GLI2-mediated transcription (Infante et al. 2015). Pyrvinium can indirectly regulate GLI2 by activating CK1, promoting complete proteasomal degradation of GLI2 (Infante et al. 2015). Arsenic Trioxide (ATO) binds directly to GLI2, destabilizing the protein and preventing its accumulation in the cytoplasm. GLI antagonists (GANTs) are also direct inhibitors of GLI activity. GANT61 and GANT58 bind the zinc finger domain in a groove opposite of DNA binding in GLI1 and GLI2, blocking their transcriptional activity (Rimkus et al. 2016).

Roles in Inflammation

Recently, roles for GLI2 in the modulation of inflammatory processes have been uncovered. GLI2 was found to regulate IL-6 expression in BM stromal cells (Elsawa et al. 2011). Furthermore, GLI2 was found to regulate the expression of the IL-6 receptor α subunit in not only malignant B cells but also in normal mouse B-1 primary cells (Jackson et al. 2015). IL-6 is a well-established important inflammatory cytokine (Tanaka et al. 2014). These recent findings suggest a role for GLI2 in not only pathology, such as cancer and developmental disease, but also in normal immunology, particularly the inflammatory response.

Summary

Over the last four decades, ample evidence has been produced illuminating the role of GLI2 not only in normal development but also in pathologic processes leading to developmental diseases and the formation and maintenance of numerous malignancies. More recent evidence suggests a role for GLI2 in inflammation and perhaps normal immunological function. Further studies are certain to uncover many new roles for GLI2 and continue to highlight its importance in multiple aspects of human biology.

Related Molecules

References

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Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  • David A. Jackson
    • 1
  • Jason A. Misurelli
    • 1
  • Sherine F. Elsawa
    • 1
  1. 1.Department of Biological SciencesNorthern Illinois UniversityDeKalbUSA