we noted that the overexpression of myt1l correlated with cxcr1 mrna levels in 4 out of 6 brain cancer cell lines ( figure 3a, b ) and the induction of cxcr1 in the normal dna - pk hek293 and m059k cells in response to the ectopically expressed myt1l ( figure 3c, d ), supporting the opinion that cxcr1 is a transcriptional target of myt1l. surprisingly, in m059j cells with loss of dna - pk function ( supplementary figure s4 ), the enforced expression of myt1l failed to induce cxcr1 transcription ( figure 3c ), although the protein levels of cxcr1 were not consistent with its mrna levels ( figure 3c, d ). taken together, our results demonstrate that myt1l functionality depends on dna - pk activity. furthermore, the ectopically expressed myt1l increased the luciferase activity of the wild - type cxcr1 promoter / luciferase reporter in hek - 293 and m059k cells with the normal expression of dna - pk, which was abolished when the mutant construct was used ( figure 3g ). however, the ectopic expression of myt1l suppressed the luciferase activity in the dna - pk - deficient m059j cells ( figure 3g ). moreover, the enforced expression of myt1l resulted in the sustained activation of the akt pathway in all three cell lines examined ( figure 1k and figure 2i ). the erk1 / 2 pathway was activated in m059k and m059j cells, while it was inhibited in hek293 cells in response to myt1l ( figure 1k and figure 2i ). the activation of the erk1 / 2 pathway was cxcr1 - dependent, whereas the akt pathway was not ( figure 4j ). based on these findings, we may propose that under non - stimulated conditions, myt1l may act as a transcriptional repressor suppressing cxcr1 transcription; dna - pk may function as a coactivator, and once it is phosphorylated by the cxcr1 - dependent activation of the erk1 / 2 kinase, it, in turn, phosphorylates myt1l; the phosphorylated myt1l recruits histone acetyltransferases ( hats ) and transcription factors ( tfs ) to the cxcr1 promoter and promotes cxcr1 transcription ( figure 5 ). dna - pk ( also known as prkdc and p350 ) is a nuclear serine / threonine protein kinase complex that is compose d of a catalytic subunit of dna - pkcs and a heterodimer of ku proteins ( ku70 / ku80 ). although one of the most well - defined functions of dna - pk is to govern the repair of dna double - strand breaks through both non - homologous end - joining ( nhej ) and homologous recombination ( hr ) [ 42 ], it was originally identified as a component of the sp1 transcription complex in which it may modulate the transcriptional activity of the complex by phosphorylating sp1 [ 43 ]. for the first time, our findings demonstrated that dna - pk may functionally modulate the transcriptional activity of myt1l by phosphorylation, eventually leading to the transcriptional activation of cxcr1, although this requires further validation using chromatin immunoprecipitation quantitative pcr ( chip - qpcr ) and the electrophoretic mobility shift assay ( emsa ) when a chip - grade antibod y to the phosphorylated myt1l is commercially available. our previous work demonstrated that dna - pk may also function as a repressor of myt1l, resulting in the transcriptional inhibition of tumor suppressor p21, a transcriptional target of myt1l [ 29 ]. several publications support the dual role of dna - pk in the transcriptional control of gene expression. dna - pk inhibited the transcription of the xanthine oxidoreductase gene via e - box / tata - like elements [ 44 ]. simultaneously, ku proteins may also act as a transcriptional recycling coactivator of the androgen receptor [ 45 ]. chemok ines are a superfamily of small ( about 8 to 14 kda ) cytokine - like proteins that selectively regulate the recruitment and trafficking of leukocytes to inflammatory sites through chemoat traction [ 46 ]. members of this family have been divided into four subfamilies, cxc, cc, c, and cx3c, based on the arrangement of the first two of the four conserved cysteine residues in the amino terminus of the proteins, which can bind cxcr, ccr, xcr1, and cx3cr1, respectively. chemok ines play a critical role in inflammatory reactions. il - 8 is a well - characterized elr + cxc chemok ine that attracts neutrophils through binding cxcr1 and cxcr2 receptors on the cell surface [ 47 ]. it is suspected that the signature of chemok ines that persist at inflammatory sites may be important in the development of chronic diseases and can contribute to the development of malignancies [ 48 ]. there is sufficient evidence to show that chemok ines also play a significant role in cancer, including melanoma [ 49 ], breast [ 50 ], colon [ 51 ], esophageal [ 52 ], prostate [ 53 ], and non - small cell lung [ 54 ] cancers, in addition to its role in the development of inflammatory responses. here, we showed that il - 8 was significantly up - regulated in hek293 and m059j cells, while it was only slightly increased in m059k cells in response to the ectopically expressed myt1l ( figure 4a ). unlike il - 8, groα more selectively binds to the cxcr2 receptor, while the affinity to cxcr1 is significantly lower [ 55 ]. since the cxcr2 receptor was undetectable in hek293 and m059k cells ( figure 3e ), groα in these cells could only function through cxcr1. the enforced expression of myt1l caused an elevation of groα in hek293 and m059j cells ( figure 4b – e ). although groα mrna was down-regulate d, its protein level was unaffected in m059k cells in response to the ectopically expressed myt1l ( figure 4f, g ). il - 8 and / or groα bind to cxcr1, eventually promoting glioblastoma proliferation via the erk1 / 2 pathway that can be blocked by cxcr1 sirna ( figure 4j, k ), suggesting that the activation of the proliferative erk1 / 2 pathway in glioblastoma is, at least in part, cxcr1 - dependent. although this is the first report regarding the critical role of dna - pk / myt1l in the proliferative il - 8 / groα - cxcr1 signaling loop in glioblastoma, the significant contributing role of il - 8 - cxcr1 / 2 axes in the proliferation and angiogenesis of glioblastoma cells has been demonstrated before [ 28 ]. taken together, we propose a potential dna - pk / myt1l - cxcr1 signaling loop in the progression of glioblastoma ( figure 5 ). we speculate that the dna - pk / myt1l - cxcr1 - erk1 / 2 might be up - regulated at who grade iv and / or recurrent glioblastomas due to chemo - or radio - resistance. dna - pk is a key player in dna damage response on double - strand breaks. activation of dna damage response facilitates glioma stem cells to develop radio - resistance [ 56 ]. furthermore, dna - pk is a master kinase of the proliferative / progenitor subtype of glioblastoma, guiding targeted cancer therapy [ 57 ]. immunoh istochemical staining has indicated the expression of il - 8 in 66. 7 – 67. 3 % of who grade iv glioblastoma tissues, with the expression of its receptor cxcr1 found primarily in both tumor - associated vessels and grade iv glioblastomas [ 28 ], supporting the previous report showing a crucial role of il - 8 / cxcr1 / 2 signaling in glioblastoma stem cells [ 58 ]. moreover, the il - 8 / cxcr1 / stat3 pathway is crucial for the maintenance of glioblastoma stem cells [ 59 ]. glioblastoma and neuroblastoma are two common types of brain tumors that occur in different aged populations. our previous studies indicated that myt1l was overexpressed in glioblastoma cell lines and 46. 9 % of malignant glioma tissue samples ( n = 32 ) [ 29 ], functioning as an oncogene in glioblastoma cells with normal dna - pk activity. to see whether myt1l is also up - regulated in neuroblastoma, we compared its expression in both glioblastoma and neuroblastoma cell lines. it was found that myt1l was overexpressed in 3 out of 7 neuroblastoma cell lines examined ( figure 3a ), suggesting that myt1l up - regulation may be a common event in both glioblastoma and neuroblastoma.