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MITF controls the TCA cycle to modulate the melanoma hypoxia response.

ABSTRACT: In response to the dynamic intra-tumor microenvironment, melanoma cells adopt distinct phenotypic states associated with differential expression of the microphthalmia-associated transcription factor (MITF). The response to hypoxia is driven by hypoxia-inducible transcription factors (HIFs) that reprogram metabolism and promote angiogenesis. HIF1α indirectly represses MITF that can activate HIF1α expression. Although HIF and MITF share a highly related DNA-binding specificity, it is unclear whether they co-regulate subset of target genes. Moreover, the genomewide impact of hypoxia on melanoma and whether melanoma cell lines representing different phenotypic states exhibit distinct hypoxic responses is unknown. Here we show that three different melanoma cell lines exhibit widely different hypoxia responses with only a core 23 genes regulated in common after 12 hr in hypoxia. Surprisingly, under hypoxia MITF is transiently up-regulated by HIF1α and co-regulates a subset of HIF targets including VEGFA. Significantly, we also show that MITF represses itself and also regulates SDHB to control the TCA cycle and suppress pseudo-hypoxia. Our results reveal a previously unsuspected role for MITF in metabolism and the network of factors underpinning the hypoxic response in melanoma.

SUBMITTER: Louphrasitthiphol P

PROVIDER: S-EPMC6777998 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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MITF controls the TCA cycle to modulate the melanoma hypoxia response.

Louphrasitthiphol Pakavarin P Ledaki Ioanna I Chauhan Jagat J Falletta Paola P Siddaway Robert R Buffa Francesca M FM Mole David R DR Soga Tomoyoshi T Goding Colin R CR

Pigment cell & melanoma research 20190708 6

In response to the dynamic intra-tumor microenvironment, melanoma cells adopt distinct phenotypic states associated with differential expression of the microphthalmia-associated transcription factor (MITF). The response to hypoxia is driven by hypoxia-inducible transcription factors (HIFs) that reprogram metabolism and promote angiogenesis. HIF1α indirectly represses MITF that can activate HIF1α expression. Although HIF and MITF share a highly related DNA-binding specificity, it is unclear wheth ...[more]

PMID: 31207090

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Project description:IntroductionMacrophage phenotypes have been linked to progression and prognosis of cutaneous melanoma. However, the association between Warburg effect in A375 melanoma and macrophages polarization, as well as the underlying mechanisms, remains less well documented.ObjectiveThe present study aimed to investigate the effect of lactate derived from A375 melanoma on macrophage polarization, melanoma phenotype responses and the underlying mechanisms.MethodsFlow cytometry was performed to evaluate the expression of M1 and M2 markers, cell cycle and apoptosis. Levels of transforming growth factor β (TGF-β) and tumor necrosis factor α (TNF-α) were determined with enzyme-linked immunosorbent assay (ELISA) kit. Proliferation and invasion were assessed by CCK8 and transwell assays, respectively. The extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) were analyzed using an XF96 extracellular flux analyzer. Protein expressions were determined by Western blotting.ResultsOur results revealed that melanoma A375 conditioned medium (A375-CM) induced peripheral blood mononuclear cells (PBMCs) to polarize toward anti-inflammatory M2 macrophages. M2 markers CD206 and ARG1 expression increased, as did TGF-β secretion. Conversely, M1 marker CD68 expression decreased. Furthermore, hypoxia promoted macrophage M2 polarization induced by A375-CM. Elevated lactate level in PIG1-conditioned medium (PIG1-CM) induced M2 polarization, whereas the lactate transport inhibitor AZD3965 suppressed this effect in PBMCs cultured with A375-CM. Additionally, lactate derived from melanoma regulated M1/M2 polarization by the tricarboxylic acid (TCA) cycle instead of glycolysis. Significantly, polarized macrophages altered melanoma phenotypes including proliferation, clone formation, cell cycle, apoptosis, migration and invasion via TCA cycle and TGF-β.ConclusionOur data collectively demonstrate that lactate derived from melanoma facilitates polarization of M2 macrophages, which subsequently leads to modifications in melanoma phenotypes via TCA cycle and TGF-β signaling.

Dataset Information

MITF controls the TCA cycle to modulate the melanoma hypoxia response.

Publications

MITF controls the TCA cycle to modulate the melanoma hypoxia response.

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