Tumor suppressor functions of DAXX through histone H3.3/H3K9me3 pathway in pancreatic NETs

Pancreatic neuroendocrine tumors (PanNETs) have considerable malignant potential. Frequent somatic mutations and loss of DAXX protein expression have been found in PanNETs. DAXX is known as a transcriptional repressor; however, molecular functions underlying DAXX loss remain unclear in PanNETs. We evaluated DAXX expression by immunohistochemistry in 44 PanNETs. DAXX-knockdown (KD) and -knockout (KO) PanNET cells were analyzed for in vitro and vivo The target genes were screened by microarray and chromatin immunoprecipitation (ChIP) assays for DAXX, histone H3.3 and H3K9me3 complex. In clinicopathological features, low DAXX expression was significantly correlated with nonfunctional tumors, higher Ki-67 index and WHO grade. Microarray and ChIP assays of DAXX-KD/KO identified 12 genes as the direct targets of DAXX transcriptional repressor. Among them, expression of five genes including STC2 was suppressed by DAXX/H3.3/H3K9me3 pathway. DAXX-KD/KO cells enhanced sphere forming activity, but its effect was suppressed by knockdown of STC2 In xenograft models, tumorigenicity and tumor vessel density were significantly increased in DAXX-KO cells with high expression of STC2. Clinically, higher recurrence rate was recognized in PanNETs with low expression of DAXX and high expression of STC2 than others (P = 0.018). Our data suggest that DAXX plays as a tumor suppressor and DAXX/H3.3 complex suppresses target genes by promoting H3K9me3 in PanNETs. Combination of DAXX loss and its target gene STC2 overexpression might be effective biomarkers and therapeutic candidates.