Paired Related Homeobox 1 Transactivates Dopamine D2 Receptor To Maintain Propagation And Tumorigenicity Of Glioma-Initiating Cells

Almost 80%of malignant tumors in central nervous system(CNS)are malignant glioma,and nearly half of these tumors are glioblastoma multiforme(GBM),which is the most aggressive tumor in CNS.Traditional treatments such as surgical resection,radiation,and cytotoxic chemotherapy are mostly palliative.GBMs have high recurrence rate with a poor prognosis with patients who are newly diagnosed having only 14 month median survival rate.Recent studies indicate that a small cell population called glioma initiating cells/glioma stem cells(GICs/GSCs),which are responsible for tumor initiation,maintenance and recurrence.They are capable of self-renewal and differentiation similar to normal neural stem cells.Most importantly,they could generate tumors in xenografted immunocompromised mice,these may utilize factors involved in embryonic development to maintain self-renewal of GICs and promote tumor growth.Homeodomain transcription factor PRRX1 plays important roles in skeletogenesis,morphogenesis,and solid tumor invasion,but the underlying molecular mechanisms remain largely unknown.Herein,we investigated the roles of PRRX1 in cortical development of mouse brain and human glioma tumorigenesis.We found Prrx1 is a key regulator to maintain self-renewal of mouse cortical neural progenitor cells,knockdown Prrxl represses mice NPCs self-renewal and promotes neurogenesis.Both overexpression and knockdown of Prrxl impaired neuronal differentiation and migration.Interestingly,PRRX1 is highly expressed in glioma samples,Depletion of PRRX1 significantly inhibits glioma cells proliferation both in vitro and in vivo.To explore the roles of PRRX1 in glioma stem cells,we isolated glioma stem cells from fresh surgical specimens.These cells highly expressed stem cell marker NESTIN and SOX2,and they can differentiated into astrocytes and pericytes under optimal conditions.PRRX1 Knockdown in GICs remarkably impairs their self-renewal and tumor initiation,which can be fully rescued by overexpression PRRX1A or PRRX1B in vitro.Surprisingly,we identified dopamine receptor 2(DRD2)as a potential target of PRRX1 in GICs by using RNA-seq transcriptome analysis.We found both PRRXIAand PRRX1B could active DRD2 promoter-drived luciferase reporter,and they were capable of binding to DRD2 promoter in ChIP-qPCR assay.We tested DRD2 levels in PRRX1-knockdown cells including immortalized hNPCs,GICs,and particular cell lines such as A172 and T98G,and found that DRD2 levels in hNPCs and GICs showed a significantly reduction than other cell lines.It suggest that PRRX1 positively regulates DRD2 expression preferentially in stem cells.Both PRRX1 and DRD2 are enriched in CD133+ population from fresh surgical specimen of malignant glioma.Blockage or knockdown of DRD2 signaling phenocopied PRRX1 knockdown in GICs.DRD2 two variant iso forms D2L and D2S,are equally potent in rescuing GICs propagating defects caused by PRRX1 knockdown.Both D2L and D2S could reverse the cell-cycle arrest and p-ERK levels in PRRX1-knockdown GICs independent of agonist treatment.These findings reveal a new molecular mechanism of PRRX1 in GICs during glioma tumorigenesis,and therapeutic targeting PRRX1-DRD2 axis is a promising strategy for treating malignant gliomas.