Hypoxia‐induced HIF1α targets in melanocytes reveal a molecular profile associated with poor melanoma prognosis

Abstract

Summary Hypoxia and HIF1α signaling direct tissue-specific gene responses regulating tumor progression, invasion and metastasis. By integrating HIF1α knockdown and hypoxia-induced gene expression changes, this study identifies a melanocyte-specific, HIF1α-dependent/hypoxia-responsive gene expression signature. Integration of these gene expression changes with HIF1α ChIP-Seq analysis identifies 81 HIF1α direct target genes in melanocytes. The expression levels for ten of the HIF1α direct targets – GAPDH, PKM, PPAT, DARS, DTWD1, SEH1L, ZNF292, RLF, AGTRAP, and GPC6 – are significantly correlated with reduced time of Disease Free Status (DFS) in melanoma by logistic regression (P-value =0.0013) and ROC curve analysis (AUC= 0.826, P-value<0.0001). This HIF1α-regulated profile defines a melanocyte-specific response under hypoxia, and demonstrates the role of HIF1α as an invasive cell state gatekeeper in regulating cellular metabolism, chromatin and transcriptional regulation, vascularization and invasion.

Non-Canonical Targets of HIF1a Drive Cell-Type-Specific Dysfunction

“…Whether sustained activation of unique cell-type-specific HIF1a target genes drive cellular dysfunction in hypoxia is unknown. To categorize canonical and cell-type-specific targets of HIF1a, we overlapped HIF1a targets in OPCs with HIF1a targets from the limited number of publicly available datasets derived from other mouse cell types including melanocytes (Loftus et al, 2017), T-cells (Ciofani et al, 2012), and embryonic heart (Guimaraes-Camboa et al, 2015). This analysis identified 51 genes that were HIF1a targets across all 4 cell types (“HIF1a core targets”), 152 genes that were HIF1a targets only in OPCs based on these datasets (“OPC-specific HIF1a targets”) and 2250 genes that were specific to either heart, T-cells, or melanocytes (“Other tissue-specific HIF1a targets”) (Figure 3A).…”

“…HIF1a ChIP-seq raw data were re-analyzed for: two HIF1a replicates ChIP-seqs and 1 input of E12.5 heart (GSM1500750, GSM1500751, GSM1500749) (Guimaraes-Camboa et al, 2015), two HIF1a replicates ChIP-seqs and 1 input of melanocyte (GSM2305570, GSM2305571, GSM2305572) (Loftus et al, 2017), and two HIF1a ChIP-seqs and 2 inputs of Th17 cells (GSM1004819, GSM1004991, GSM1004820, GSM1004993) (Ciofani et al, 2012). For each of the HIF1a ChIP-seqs, the peaks were called as described above (MACS2, narrow peak, FDR<0.001).…”

GPC6 Promotes Cell Proliferation, Migration, and Invasion in Nasopharyngeal Carcinoma

“…Moreover, GPC6 was found to be frequently mutated and aberrantly methylated in colorectal cancer (in 12 paired colorectal tumors and adjacent healthy mucosal tissues using the Illumina Infinium Human Methylation 450 BeadChip) [55]. Similarly, a previous study demonstrated that increased GPC6 expression was significantly correlated with a reduced progression-free survival time in patients with melanoma, and furthermore, that this effect occurred in an HIF1α-dependent/hypoxiaresponsive manner (as shown via a ROC curve) [56]. Likewise, high GPC6 expression has been shown to be closely associated with gastric cancer progression [57].…”

Absence of Nuclear p16 Is a Diagnostic and Independent Prognostic Biomarker in Squamous Cell Carcinoma of the Cervix

“…Moreover, it helps deliver cyclin D/E to CDK4 [40] and interacts with a kinase which phosphorylates p27 to prevent its translocation to the nucleus [41]. Finally, we identified AGTRAP, a HIF1α direct target related to reduced PFS in melanoma [25] and to tumor growth and angiogenesis in Lewis lung carcinoma [42].…”

“…Some of the cytoplasmic p16 interactors identified by LC-MS/MS were selected for further examination by double immunofluorescence because of their involvement in other carcinogenic processes [23][24][25][26][27][28][29]. As seen in Figure 4C, we found that CDK4, the A-kinase anchor protein 8 (AKAP8), the barrier to autointegration factor (BANF1), and the type-1 angiotensin II receptor-associated protein (AGTRAP) colocalized with p16 in the cytoplasm of SiHa cells, confirming that these proteins interact with cytoplasmic p16 in SiHa cells.…”