T-lymphokine-activated killer cell-originated protein kinase (TOPK), also known as PDZ-binding kinase (PBK), was a member of the MEK3/6-related MAPKK family. As a mitotic serine/threonine protein kinase, accumulating evidence supported its role in mitosis and cell-cycle progression of mitotically active cells, especially proliferative malignant cells. PBK/TOPK was confirmed to be associated with the development, progression, and metastasis of malignancies, which made it a potential therapeutic target in cancer therapy. Further, it was also demonstrated to play crucial roles in ischemic injury and was involved in protection against ischemia. This protective effect of PBK/TOPK in the context of ischemia challeged the development of PBK/TOPK inhibitors in anti-tumor therapy, and more research was required to further explore its role and underlying mechanisms to translate its application to clinical astudies.
T-lymphokine-activated killer-cell-originated protein kinase (TOPK), also known as PDZ-binding kinase (PBK), is a novel mitotic serine/threonine protein kinase [1,2]. PBK/TOPK is overexpressed in various actively proliferative cells, including malignant tumor cells, as well as normal cells, such as sperm cells. The transcription, activation, and degradation of PBK/TOPK is regulated by mitosis progression, which is mediated by a series of proteins. As a mitotic kinase, PBK/TOPK is important for the proliferation, progression, and metastasis of many cancers, including leukemia and myeloma, among others [3,4]. Upregulation of PBK/TOPK has also been proven to be associated with cancer diagnosis and prognosis, and, thus, it may be a potential therapeutic target in various malignant tumors. Thus, various potent PBK/TOPK inhibitors have been developed. The function of PBK/TOPK as an emerging target for cancer-specific therapeutics has been reviewed previously by Herbert et al. [5]. Furthermore, ours and other research teams also demonstrated that PBK/TOPK plays crucial roles in ischemic injury and is involved in protection against ischemia and ischemic postconditioning [6,7,8]. This protective effect of PBK/TOPK in the context of ischemia challenged the development of PBK/TOPK inhibitors in antitumor therapy, and more research is required to further explore its role and underlying mechanisms, to translate its applicability to clinical studies.
The human homologue of the Drosophila tumor suppressor Discs-large (hDlg) is a membrane-associated guanylate kinase homologue (MAGUK) [9]. Several proteins that bind to hDlg have been demonstrated to be involved in cell growth control, including in the pathogenesis of adenomatous polyposis coli [10,11,12,13]. In 2000, Gaudet et al. sought to elucidate the signal transduction pathway through which hDlg regulates cellular proliferation, and they first identified a 322 amino acid serine/threonine kinase, using a two-hybrid screen and named it PDZ-binding kinase (PBK) [2]. At the same time, Abe et al. cloned a novel protein kinase from a lymphokine-activated killer T (T-LAK) cell subtraction cDNA fragment library and named it T-LAK cell-originated protein kinase (TOPK) [1]. This gene was later confirmed to be similar to that found by Gaudet et al., and this novel gene, or its protein product, was called PBK/TOPK.
Unlike hDlg, which is ubiquitously expressed, the expression and distribution of PBK/TOPK varies. It is abundant in highly proliferative tissues, such as the placenta, testis, T-LAK cells, activated lymphoid cells and lymphoid tumors, but it is expressed at a very low level or is even absent in non-proliferative normal tissues, such as normal adult brain tissue (with the exception of the subependymal zone and early postnatal cerebellar external layer, which is enriched with rapidly proliferating progenitor cells) [14,15]. Regarding its intracellular distribution, PBK/TOPK is expressed both in the cytoplasm and the nucleus; however, it may be expressed exclusively in the nucleus of tumor cells or in mitosis, particularly around chromosomal surfaces during prophase and metaphase [16,17]. Moreover, both PBK/TOPK protein expression and its activation are subject to cell-cycle regulation. Using the thymidine double-block method, PBK/TOPK protein expression was demonstrated to increase and peak at 8 h after being released from the cell-cycle block during G2 to M phase in HeLa cells, and this was accompanied by the upregulation of its phosphorylation activity [18].
As a member of the MEK3/6-related MAPKK family, PBK/TOPK shares the characteristic serine/threonine protein kinase subdomains and a C-terminal PDZ-binding T/SXV motif, which enables it to bind specifically to the PDZ2 domain of hDlg or other PDZ-containing proteins. Activated PBK/TOPK is also able to phosphorylate P38 MAPK and histone H3, among other proteins [2,19,20]. However, the phosphotransferase activity of PBK/TOPK appears to be regulated in a cell-cycle-dependent manner; that is, it is activated following its phosphorylation by cyclin-dependent kinase 1 (CDK1)/cyclin B1 exclusively at mitosis (Figure 1). Notably, this process depends on the binding of PBK/TOPK to the CDK1/cyclin B1 complex at the mitotic spindle, which in turn induces its phosphorylation and thereby increases its binding ability to the CDK1/cyclin B1 complex through phosphorylation and inactivation of protein phosphatase 1 alpha (PP1α) [2,21,22]. These results all suggest that PBK/TOPK may be involved in the regulation of cellular proliferation and cell-cycle progression.
In addition, the transcription of PBK/TOPK and its degradation varies with cell-cycle progression (Figure 1). The binding of the cell-cycle-specific transcription factors E2F and cyclic AMP-responsive element-binding protein/activating transcription factor (CREB/ATF) to the −146 and −312 bp binding sites within the PBK/TOPK promoter directly leads to its transcriptional upregulation [17,23]. C-Myc is functionally linked with E2F1 in controlling cell-cycle progression, and it activates PBK/TOPK transcription by indirectly enhancing E2F1 activity and cooperatively binding with E2F1 [24]. Checkpoint protein with FHA and RING domains (CHFR) is an E3 ubiquitin ligase that was demonstrated to ubiquitinate and mediate PBK/TOPK degradation, which is essential for its mitotic checkpoint function [25]. Src, a transforming protein with tyrosine kinase activity, was also identified as an upstream regulator of PBK/TOPK. It can directly bind and phosphorylate PBK/TOPK at Y74 and Y272, thereby enhancing its activity and enhancing PBK/TOPK stability, which allows it to avoid degradation after ubiquitination [26].
Mitotic progression is regulated by the co-ordination of several proteins and is crucial for the maintenance of genomic stability. PBK/TOPK is known to play pivotal roles in cellular mitosis; its main mitotic substrates and detailed functions in mitosis have been studied in depth (Figure 2). PBK/TOPK is of crucial importance in CHFR-mediated G2/M progression via phosphorylation and inactivation of PTEN, which results in activation of Akt [25]. Activated TOPK was also confirmed as the principal C2H2 zinc finger protein (ZFP) linker kinase that phosphorylates C2H2 ZFPs linkers within minutes in the prophase stage. This is a unique mechanism for reducing the DNA-binding activity of C2H2 ZFPs, which mediates the dissociation of hundreds of proteins from condensed chromatin during transition from prophase to telophase. PBK/TOPK was the first kinase to be identified as a master regulator of an entire family of transcription factors, based on their conserved motif [27]. At the late stage of mitosis, upregulated PBK/TOPK was shown to mediate the phosphorylation of LGN/GPSM2 (Leu-Gly-Asn repeat-enriched protein/G-protein signaling modulator 2) at Thr450, which might play a critical role in the segregation of sister chromatids and the formation of F-actin polymerization at the contractile ring [28]. In addition, activated PBK/TOPK was demonstrated to enhance the phosphorylation of the microtubule-binding protein PRC1 and ATPase protein p97 with p47 as an adaptor protein, which is indispensable for mitotic spindle formation and cytokinesis during mitosis [29].
PBK/TOPK overexpression in oncogenic pathways has been implicated in the proliferation, metastasis, and anti-apoptosis of tumors (Figure 3). Its overexpression allows the tumor cells to bypass the natural surveillance mechanism associated with the G2/M checkpoint and leads to aberrant entry into the mitotic phase by phosphorylating histone H3 at Ser10, as well as down-modulating the tumor suppressor p53, upregulating the cyclin-dependent kinase inhibitor p21, and thereby contributing to tumorigenesis [30,31,32]. Conversely, suppression of PBK/TOPK impaired its tumorigenic properties by reducing the phosphorylation and activation of mitogen-activated protein (MAP) kinase, including extracellular signal-regulated kinase (ERK) 2 and P38, inhibiting the expression of mutant p53, and inhibiting Akt activation [33,34,35,36,37,38]. Notably, phosphorylated ERK2 could in turn phosphorylate PBK/TOPK and increase its kinase activity, resulting in a positive feedback loop between PBK/TOPK and ERK2. PBK/TOPK also possesses the capacity to activate the interaction of transcriptional factor β-catenin, with its transcriptional coactivators T-cell factor/lymphoid enhancer-binding factor (TCF/LEF), which subsequently upregulates the transcription of matrix metalloproteinase MMP-2 and MMP-9, thereby facilitating the invasiveness and metastasis of tumor cells [16].
Aside from promoting proliferation and metastasis, activated PBK/TOPK could also alleviate As3+ treatment–induced apoptosis by binding with and phosphorylating histone H2AX at Ser139 in the nucleus. This may be responsible for the resistance of some melanoma cells to As3+ treatment [39]. Moreover, PBK/TOPK could inhibit UVB-induced apoptosis as well, by phosphorylating c-Jun-NH2-Kinase 1 (JNK1) at Thr183/Tyr185, activating it, and increasing the peroxidase activity of peroxiredoxin1 (Prx1) and decreasing the intracellular accumulation of H2O2 via phosphorylation Prx1 at Ser32 [40,41]. Moreover, TOPK directly phosphorylates inhibitor-κBα (IκBα) at Ser 32, which induces P65 nuclear translocation, nuclear factor-kappa B (NF-κB) activation, and subsequently leads to responsive transcription of anti-apoptotic genes [42,43,44]. These findings enrich our knowledge of the regulatory functions and underlying mechanisms of PBK/TOPK in mitotic progression of actively proliferating cells, especially tumors, and have led to a better understanding of its role in tumorigenesis. These studies all suggest that PBK/TOPK might serve as a diagnostic/prognostic indicator and therapeutic target in tumors.