PBK/TOPK, a Proliferating Neural Progenitor-Specific Mitogen-Activated Protein Kinase Kinase

“This approach was based on the observation that genes that are involved in the same biological process are often tightly coregulated (Eisen et al., 1998; Ren et al., 2000; Miki et al., 2001).”

“DAVID (database for annotation, visualization, and integrated discovery) was used to classify correlated and anticorrelated genes into level 5 biological processes gene ontologies (Dennis et al., 2003).”

“To provide additional functional annotation that would aid in identifying genes involved in NSC proliferation or self-renewal, we identified genes expressed in multiple CNS germinal zones and non-neural stem cell populations (Terskikh et al., 2001; Easterday et al., 2003).”

“…throughout the germinal zones of the developing embryonic brain and persist into adulthood, providing for ongoing neurogenesis in select regions of the mammalian brain, offering hope for neural repair strategies (Palmer et al., 1999; Gage, 2000; Lie et al., 2004; Morshead and van der Kooy, 2004).”

“Cytological investigations have identified three major morphologically defined populations of cells in the SEZ: ultrastructurally migrating neurons, GFAP-positive glial cells, and highly proliferative ultrastructurally immature cells (Doetsch et al., 1997; Doetsch et al., 2002).”

“Arrays were normalized with dCHIP (www.dchip.org), and expression values were calculated (Li and Wong, 2001).”

“…throughout the germinal zones of the developing embryonic brain and persist into adulthood, providing for ongoing neurogenesis in select regions of the mammalian brain, offering hope for neural repair strategies (Palmer et al., 1999; Gage, 2000; Lie et al., 2004; Morshead and van der Kooy, 2004).”

“…normal progenitors, they are proliferative neural tissue, and data from several sources have demonstrated recently that some tumors, including gliomas, contain a multipotent tumor stem cell from which they derive (Ignatova et al., 2002; Hemmati et al., 2003; Singh et al., 2003; Galli et al., 2004).”

“This revealed that in the group of genes most correlated to PBK/TOPK, there is a highly significant overrepresentation of genes involved in the cell cycle (EASE statistic; p 10e-15), especially M-phase genes (EASE statistic; p 10e-12) (supplemental Fig.”

“Cytological investigations have identified three major morphologically defined populations of cells in the SEZ: ultrastructurally migrating neurons, GFAP-positive glial cells, and highly proliferative ultrastructurally immature cells (Doetsch et al., 1997; Doetsch et al., 2002).”

“The MCM2 protein, which is expressed in the G1 phase, was used to determine whether any of the slowly cycling cells were reentering the cell cycle (Maslov et al., 2004).”

“…normal progenitors, they are proliferative neural tissue, and data from several sources have demonstrated recently that some tumors, including gliomas, contain a multipotent tumor stem cell from which they derive (Ignatova et al., 2002; Hemmati et al., 2003; Singh et al., 2003; Galli et al., 2004).”

“Recent studies have suggested that multipotent neural stem cells may be found in this region as late as the first postnatal week (Goldman et al., 1997; Milosevic and Goldman, 2002; Lee et al., 2005).”

“These data in neural tissues was parallel to data showing cell-cycle regulation of PBK/ TOPK expression in synchronized HeLa cells (Whitfield et al., 2002; Matsumoto et al., 2004).”

“, 2002) and questioning (Morshead et al., 1994) this role.”

“This is supported by data showing that purified CGP in vitro expresses PBK/TOPK, which is mitogen dependent, parallel with the SSH requirement for CGP proliferation (Wechsler-Reya and Scott, 1999).”

“Thus, the PBK/TOPK target P38 appears to be phosphorylated specifically during mitosis in proliferating CGPs, consistent with other reports of mitotically phosphorylated P38 in neuronal progenitors in other germinal zones (Campos et al., 2002).”

“Cytological investigations have identified three major morphologically defined populations of cells in the SEZ: ultrastructurally migrating neurons, GFAP-positive glial cells, and highly proliferative ultrastructurally immature cells (Doetsch et al., 1997; Doetsch et al., 2002).”

“Recent evidence strongly suggests that the primary stem cell of the adult central nervous system is GFAP positive (Doetsch et al., 1999b; Imura et al., 2003; Garcia et al., 2004).”

“Transgenic mice were created and treated as described previously (Bush et al., 1999; Imura et al., 2003; Garcia et al., 2004).”

“Extensive studies have shown that during this period, this region produces granule cells exclusively (Altman, 1972; Lee et al., 2005).”

“Furthermore, how the process of cell division may provide critical windows in which daughter cell fate or the commitment to renew are integrated with environmental signals remains a fundamental question (Lu et al., 2000).”

“NSCs exist throughout the germinal zones of the developing embryonic brain and persist into adulthood, providing for ongoing neurogenesis in select regions of the mammalian brain, offering hope for neural repair strategies (Palmer et al., 1999; Gage, 2000; Lie et al., 2004; Morshead and van der Kooy, 2004).”

“This set included 85 gliomas from 79 patients hybridized onto Affymetrix (Santa Clara, CA) HG133A and B arrays (Freije et al., 2004).”

“To provide additional functional annotation that would aid in identifying genes involved in NSC proliferation or self-renewal, we identified genes expressed in multiple CNS germinal zones and non-neural stem cell populations (Terskikh et al., 2001; Easterday et al., 2003).”

“…their rich potential for therapeutic applications, research on NSCs has been hampered by a lack of markers to identify NSCs prospectively and an incomplete understanding of the pathways involved in the regulation of their proliferation and differentiation (Anderson, 2001; Lindvall et al., 2004).”

“In the EGL, scaffolding for cell division and migration are provided by GLAST-positive Bergmann glia (Furuta et al., 1997), which are PBK/TOPK negative (Fig.”

“These data in neural tissues was parallel to data showing cell-cycle regulation of PBK/ TOPK expression in synchronized HeLa cells (Whitfield et al., 2002; Matsumoto et al., 2004).”

“To provide additional functional annotation that would aid in identifying genes involved in NSC proliferation or self-renewal, we identified genes expressed in multiple CNS germinal zones and non-neural stem cell populations (Terskikh et al., 2001; Easterday et al., 2003).”

“Considered together, all of the evidence clearly demonstrates PBK/TOPK is expressed by multiple neuronal progenitor populations during development, and possibly multipotent progenitors as well (Parras et al., 2004).”

“To identify genes involved in NSC proliferation and differentiation, we performed extensive analysis of the gene expression in neural stem and progenitor cell cultures (Geschwind et al., 2001; Karsten et al., 2003).”

“To provide additional functional annotation that would aid in identifying genes involved in NSC proliferation or self-renewal, we identified genes expressed in multiple CNS germinal zones and non-neural stem cell populations (Terskikh et al., 2001; Easterday et al., 2003).”

“Results PBK/TOPK transcript is expressed exclusively in neurogenic regions in embryonic and adult CNS Previous array studies demonstrated that PBK/TOPK was expressed by a wide variety of stem and progenitor populations in vitro, consistent with a role in stem/progenitor cell proliferation (Geschwind et al., 2001; Ters…”

“Double-label immunohistochemistry of PBK/TOPK and Tuj1 or Dcx (data not shown), both markers of immature neurons, reveal clear expression of PBK/TOPK in the PCNA-positive mitotic (Fig.”

“Recently, a subset of NG2positive cells in SEZ have also been identified as sphere forming, possibly C cells, but in contrast to EGFR, it is expressed in many regions beyond the neurogenic SEZ (Doetsch et al., 2002; Aguirre et al., 2004).”

“…throughout the germinal zones of the developing embryonic brain and persist into adulthood, providing for ongoing neurogenesis in select regions of the mammalian brain, offering hope for neural repair strategies (Palmer et al., 1999; Gage, 2000; Lie et al., 2004; Morshead and van der Kooy, 2004).”

“Previous work has shown that when these mice are treated with ganciclovir, there is a complete ablation of stem cell potential, as assessed by multipotent neurosphere forming ability in vitro (Imura et al., 2003; Morshead et al., 2003) and a complete ablation of neurogenesis in vivo (Garcia et al.”

“To extend these findings to other primary progenitor populations in addition to the CGPs, we also exposed attached neural progenitors cultured from E12 mouse telencephalic germinal zones to varying doses of SB203580 in the presence of the mitogen basic fibroblast growth factor (bFGF).”

“This is supported by data showing that purified CGP in vitro expresses PBK/TOPK, which is mitogen dependent, parallel with the SSH requirement for CGP proliferation (Wechsler-Reya and Scott, 1999).”

“In non-neural cells, PBK/TOPK appears to require cyclinB/CDK1 phosphorylation for activation (Gaudet et al., 2000).”

“The PBK/TOPK target P38 MAPK is also phosphorylated in a cell-cycle-dependant manner in neural progenitor cells Next, we wished to examine whether the PBK/TOPK target P38 MAPK is also phosphorylated in neural progenitors, because in non-neural cells, PBK/TOPK phosphorylates P38 MAPK, activating it (Abe et al., 2000).”

“Recent studies have suggested that multipotent neural stem cells may be found in this region as late as the first postnatal week (Goldman et al., 1997; Milosevic and Goldman, 2002; Lee et al., 2005).”

“These data support a role for PBK/TOPK in neural progenitor proliferation and self-renewal and, in combination with our studies of other genes (Nakano et al., 2005), supports our general genomic screening approach to identify progenitor markers. ploratory use of published microarray data sets is…”

“Furthermore, many proliferating cells in the SEZ and RMS, structures that give rise to neurons throughout the life of the animal (Luskin, 1993), are positive for both PBK/TOPK and the proneu- ral Mash1 gene.”

“To identify genes involved in NSC proliferation and differentiation, we performed extensive analysis of the gene expression in neural stem and progenitor cell cultures (Geschwind et al., 2001; Karsten et al., 2003).”

“, 2004), and Mash1 knock-out mice have morphological defects in the olfactory bulb (Guillemot, 1995; Murray et al., 2003; Parras et al., 2004).”

“However, it was absent in WiDr and HT-29 colon cancer cells, indicating that it was not ubiquitously expressed in cycling cells (Abe et al., 2000; Simons-Evelyn et al., 2001; Zhao et al., 2001).”

“Recent studies have suggested that multipotent neural stem cells may be found in this region as late as the first postnatal week (Goldman et al., 1997; Milosevic and Goldman, 2002; Lee et al., 2005).”

“The EGL contains a mitotic layer with PCNA-positive proliferating progenitors (EGLa), a premigratory layer with postmitotic immature neurons (EGLb), and the radially oriented processes of Bergmann glia (Migheli et al., 1999).”

“Recent studies have suggested that multipotent neural stem cells may be found in this region as late as the first postnatal week (Goldman et al., 1997; Milosevic and Goldman, 2002; Lee et al., 2005).”

“Mash1 has recently been shown to specify neuronal and oligodendritic fate in the postnatal brain in vitro and in vivo (Parras et al., 2004), and Mash1 knock-out mice have morphological defects in the olfactory bulb (Guillemot, 1995; Murray et al., 2003; Parras et al., 2004).”

“Furthermore, activation of PBK/TOPK required phosphorylation by both the M-phase kinase complex cyclinB/cyclindependent kinase 1 (CDK1) and another unknown kinase, possibly RafC or RafA (Gaudet et al., 2000; Yuryev and Wennogle, 2003).”

“Thus, the PBK/TOPK target P38 appears to be phosphorylated specifically during mitosis in proliferating CGPs, consistent with other reports of mitotically phosphorylated P38 in neuronal progenitors in other germinal zones (Campos et al., 2002).”

“However, it was absent in WiDr and HT-29 colon cancer cells, indicating that it was not ubiquitously expressed in cycling cells (Abe et al., 2000; Simons-Evelyn et al., 2001; Zhao et al., 2001).”

“Thus, gene products that are involved in an unknown cell type or biological process can be annotated on the basis of their proximity to marker genes of known function (‘guilt by association’) 26,33,34 .”

“Cell lines—Glioma spheres were established from 14 GBM patient samples (sample ID: GSC157, GSC84, GSC185A, GSC83, GSC267, GSC11, GSC20, GSC28, GSC23, GSC11, GSC1010, GSC1020, GSC1051, GSC1079) (Nakano et al., 2008; Dougherty et al., 2005; Gu et al., 2013; Mao et al., 2013; Bhat et al., 2013).”

“In normal adult organs, PBK is most abundant in the testis, mainly in proliferating spermatocytes, as well as in several fetal tissues and in proliferating brain neural stem cells (Dougherty et al., 2005; Fujibuchi et al., 2005).”

“Furthermore, based on its restricted pattern of expression to tumor cells, testis, embryonic tissues and adult progenitor cells (Simons-Evelyn et al., 2001; Dougherty et al., 2005; Park et al., 2006), PBK may be considered a novel cancer/testis antigen that may also have a role in maintenance of tumor stem cells.”

“It was cloned as a new MAPKK-like protein kinase, highly conserved among vertebrates, specifically expressed in the testis, placenta, activated lymphoid cells, and neural lineages (Abe et al, 2000; Dougherty et al, 2005).”

“PBK, a serine/ threonine kinase, is functionally related to the AKT pathway and can regulate cell cycle [41, 42].”

“Among normal cell types, PBK/TOPK is expressed in highly proliferating cells such as spermatocytes, in several fetal tissues as well as in neural stem and progenitor cells [18, 23].”

“In the sea urchin, there are six identifiedM2Ks: 5 STE7 kinases and 1 kinase from the Other group, TOPK.”

“PBK/TOPK has also been demonstrated in specific neural progenitor cells in vivo [14], and is found to be upregulated in hormone refractory metastatic prostate carcinoma as seen by querying the Oncomine database [15,16].”

“Furthermore, PBK/TOPK expression was shown to prompt the proliferation of Ewing sarcoma cells and neural cells.(8,11) According to our results and other relevant reports, we speculate that PBK/TOPK overexpression may not only inhibit p53 function but also promote TP53 mutation.”

“Not surprisingly, the subset of OPC transcripts identified by GO as involved in cell cycle/cell proliferation, such as Melk (27) and Pbk (28), are highly enriched in the tumor samples, consistent with the Olig2: Egfp-L10a population being in a highly proliferative state inside of the tumor.”

“Most of the inhibitors exhibited general cytotoxicity and were not studied further; others have been described as exerting negative effects on neuronal function, e.g., p38 MAP kinase inhibitors [10, 11 ].”

“In the neurogenic SVZ, Cerulean from an Aldh1L1 BAC (in Prism line JD1849) is found in a pattern consistent with known localization of “Type B” neural stem cells (Doetsch, 2003; Dougherty et al., 2005; Nakano et al., 2007): Cerulean1 cells surround pockets of Cerulean–, DAPI1 nuclei (presumptive neuroblasts) (Fig.”

“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 earl…”

“These progenitor cells have differential expression profiles and activities of several kinases (MSK1/2, PBK/TOPK, ERK5MAPK, PI3K/Akt) as comparedwith adult neurons (Choi et al., 2012; Dougherty et al., 2005; Pan et al., 2012; Peltier et al., 2007).”

“Some authors reported that TOPK expression did not affect cell proliferation [17,35], but others reported that high TOPK expression might promote cell proliferation [9,11–13].”

“…tissues and in a wide variety of tumours in vitro, whereas the inhibition of TOPK is shown to lead to apoptosis in breast and melanoma cell lines (Simons-Evelyn et al, 2001; Zhao et al, 2001; Matsumoto et al, 2004; Nandi et al, 2004; Dougherty et al, 2005; Park et al, 2006; Zykova et al, 2006).”

“As most PBK/TOPK-positive cells were all PCNApositive in neurons system [18], it was necessary to know if there was a correlation between PBK/TOPK expression and cell proliferation in this study.”

“[77] found that TOPK also plays a crucial role in progenitors and is generally associated with the p38/MAPK pathway, indicating that TOPK is an important regulator of growth and self-renewal in progenitor.”

“PDZ-binding kinase (PBK), a serine/threonine kinase, functions as the upstream kinase of p38/ERK signaling [7].”

“As one example, we found a novel interaction between the PDZ of Erbin, which mediates the localization of its target proteins such as ErbB2[44], and PBK (PDZ domain binding kinase), which is recruited to MAPK14 through interacting with hDLG PDZ and activates MAPK14 signaling[45, 46].”

“Inhibition of TOPK disrupts neural progenitor self-renewal and proliferation [11].”

“This approach also provides the basis for gene discovery, using a ‘guilt by association’ approach.(30,35) Indeed, our analysis has identified”

“…as T-LAK cell-originated protein kinase (TOPK), is a serine/threonine kinase that promotes the proliferation and self-renewal of neural stem cells [28]; 3) PBK is overexpressed in diverse adult histo-types and implicated in multiple hallmarks of cancer, including cell cycle regulation, apoptosis,…”

“PBK is highly enriched in neural stem cells (20).”

“2001); and phosphorylation of p38 is involved in both the proliferation of neuronal precursors (Dougherty et al. 2005) and neuronal death (Xia et al.”

“Knockdown of PBK inhibits the proliferation of human epidermal keratinocytes and neural progenitor cells [25, 26].”

“PBK is hyperexpressed in cancer cells, T - LAK cells, and activated lymphoid cells ( 14 , 15 ), and capable of regulating phosphorylated levels of JNK, p38, and ERK involved in the proliferation and metastatic potential of cancer cells ( 13 , 16 ).”

“Another group of gene transcripts that were upregulated in OLPs treated with PD169316 included the upstream enzymes regulating p38 activity such as MKKKs (Dlk1), and PBK, a MKK that specifically phosphorylates p38 during cell cycle [17, 28].”

“Thus, as nodes, gene products that are associated with biological processes or ill-defined cell types are annotated by their neighbors and marker genes with known functions (Dougherty et al., 2005; Oldham et al., 2008; Wolfe et al., 2005).”

“Although a previous study reported that the pharmacological inhibition of p38 reduces CGNP proliferation of PN 6–8 CGNPs [15], a link between p38a activity and the Shh pathway in CGNPs was not described.”

“TOPK act as a marker as well as involved in the proliferation of neuronal progenitor cells [6].”

“In consequence of its identification in NPC microarray screens, Dougherty et al. (2005) studied Pbk expression and function in these cells.”

“However, PBK has also been shown to be differentially expressed in normal proliferative cells, including neural precursor cells in the subventricular zone of the adult brain, as well as under pathological conditions, such as ischemic tissues, including the brain and plays important roles in their physiological functio…”

“Dougherty JD, Garcia AD, Nakano I, Livingstone M, Norris B, Polakiewicz R, Wexler EM, Sofroniew MW, Kornblum HL, Geschwind DH: PBK/TOPK, a proliferating neural progenitor-specific mitogen-activated protein kinase kinase.”

“phosphorylated by a specific MAPKK, PDZ-binding kinase/ T-LAK cell-originated protein kinase that is activated by cyclin B/CDK1 [38].”

“PDZ-binding kinase (PBK, also known as TOPK) is a serine-threonine kinase belonging to the MAPKkinase (MAPKK) family, that promotes phosphorylation of p38 MAPK [55], leading to the identification of p38 MAPK as a specific substrate of PBK [56] and suggesting the cell-type-dependent involvement of PBK in ERK/ MAPK, p38…”

“PBK is a serine/threonine protein kinase which is associated with the mitogen-activated protein kinase (MAPK) kinase (MAPKK) family (22).”

“Additionally, Dougherty et al (32) reported PBK expression in rapidly proliferating central nervous system cells of mice, such as adult subependymal neuronal progenitors and granular cell precursors of the postnatal cerebellum, and its suppres‐ sion in neurons, mature glia and quiescent cells.”

“Neural progenitor cells also express Pbk during the preneonatal and postneonatal period [14].”

“PDZ-binding kinase (PBK) encodes a serine/threonine protein kinase, which is related to the bispecific mitogen-activated protein kinase (MAPKK) family and participates in many cell processes, such as proliferation, differentiation, transcription regulation and development (6).”

“TOPK, also known as PBK, encodes a serine/threonine protein kinase related to the dual specific mitogen-activated protein kinase kinase (MAPKK) family (13).”

“EGL NPCs were isolated from dissociated tissue using a 35%/65% Percoll fractionation using a 12min centrifugation at 2500 rpm as described previously [23].”

“In the central nervous system, PBK-positive cells are found in rapidly proliferating early postnatal cerebellar granules, the adult subventricular zone, and glial fi brillary acidic protein (GFAP)-positive neural stem cells, while PBK expression is signi fi cantly reduced in mature neurons [11] .”

“Pbk is aberrantly expressed or activated in various cancers and is essential for neuronal progenitor proliferation, self-renewal, and adult neurogenesis [20, 21].”

“In the adult mouse brain, Pbk is expressed in rapidly proliferating NS/PCs of the adult subventricular zone and early postnatal cerebellar external granular layer [6].”

“This mitotic kinase may be involved in the activation of lymphoid cells and support testicular functions, with an implied role in the process of spermatogenesis (SimonsEvelyn et al., 2001; Dougherty et al., 2005; Zhu et al., 2007; Nandi et al., 2007).”

“B cell BANK1 [19], MS4A1 [20] Part of the adaptive immune system; produces antibodies Cell cycle PBK [21], TOP2A [22] Represents proliferating cells across different cell types Cholangiocyte EPCAM [23], KRT19 [24] Epithelial cells lining bile ducts; involved in bile production and secretion…”