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HIF isoforms have divergent effects on invasion, metastasis, metabolism and formation of lipid droplets.

ABSTRACT: Cancer cells adapt to hypoxia by the stabilization of hypoxia inducible factor (HIF)-? isoforms that increase the transcription of several genes. Among the genes regulated by HIF are enzymes that play a role in invasion, metastasis and metabolism. We engineered triple (estrogen receptor/progesterone receptor/HER2/neu) negative, invasive MDA-MB-231 and SUM149 human breast cancer cells to silence the expression of HIF-1?, HIF-2? or both isoforms of HIF-?. We determined the metabolic consequences of HIF silencing and the ability of HIF-? silenced cells to invade and degrade the extracellular matrix (ECM) under carefully controlled normoxic and hypoxic conditions. We found that silencing HIF-1? alone was not sufficient to attenuate invasiveness in both MDA-MB-231 and SUM149 cell lines. Significantly reduced metastatic burden was observed in single (HIF-1? or HIF-2?) and double ?-isoform silenced cells, with the reduction most evident when both HIF-1? and HIF-2? were silenced in MDA-MB-231 cells. HIF-2? played a major role in altering cell metabolism. Lipids and lipid droplets were significantly reduced in HIF-2? and double silenced MDA-MB-231 and SUM149 cells, implicating HIF in their regulation. In addition, lactate production and glucose consumption were reduced. These results suggest that in vivo, cells in or near hypoxic regions are likely to be more invasive. The data indicate that targeting HIF-1? alone is not sufficient to attenuate invasiveness, and that both HIF-1? and HIF-2? play a role in the metastatic cascade in these two cell lines.

SUBMITTER: Shah T

PROVIDER: S-EPMC4695047 | biostudies-literature | 2015 Sep

REPOSITORIES: biostudies-literature

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HIF isoforms have divergent effects on invasion, metastasis, metabolism and formation of lipid droplets.

Shah Tariq T Krishnamachary Balaji B Wildes Flonne F Mironchik Yelena Y Kakkad Samata M SM Jacob Desmond D Artemov Dmitri D Bhujwalla Zaver M ZM

Oncotarget 20150901 29

Cancer cells adapt to hypoxia by the stabilization of hypoxia inducible factor (HIF)-α isoforms that increase the transcription of several genes. Among the genes regulated by HIF are enzymes that play a role in invasion, metastasis and metabolism. We engineered triple (estrogen receptor/progesterone receptor/HER2/neu) negative, invasive MDA-MB-231 and SUM149 human breast cancer cells to silence the expression of HIF-1α, HIF-2α or both isoforms of HIF-α. We determined the metabolic consequences o ...[more]

PMID: 26305551

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Project description:SR-BI binds various lipoproteins, including HDL, LDL as well as VLDL, and mediates selective cholesteryl ester (CE) uptake. HDL derived CE accumulates in cellular lipid droplets (LDs), which also store triacylglycerol (TAG). We hypothesized that SR-BI could significantly facilitate LD formation, in part, by directly transporting LDL derived neutral lipids (NL) such as CE and TAG into LDs without lipolysis and de novo lipid synthesis. SR-BI overexpression greatly increased LDL uptake and LD formation in stably transfected HeLa cells (SR-BI-HeLa). LDs isolated from SR-BI-HeLa contained 4- and 7-times more CE and TAG, respectively, than mock-transfected HeLa (Mock-HeLa). In contrast, LDL receptor overexpression in HeLa (LDLr-HeLa) greatly increased LDL uptake, degradation with moderate 1.5- and 2-fold increases of CE and TAG, respectively. Utilizing CE and TAG analogs, BODIPY-TAG (BP-TAG) and BODIPY-CE (BP-CE), for tracking LDL NL, we found that after initial binding of LDL to SR-BI-HeLa, apoB remained at the cell surface, while BP-CE and BP-TAG were sorted and simultaneously transported together to LDs. Both lipids demonstrated limited internalization to lysosomes or endoplasmic reticulum in SR-BI-HeLa. In LDLr-HeLa, NLs demonstrated clear lysosomal sequestration without their sorting to LDs. An inhibition of TAG and CE de novo synthesis by 90-95% only reduced TAG and CE LD content by 45-50%, and had little effect on BP-CE and BP-TAG transport to LDs in SR-BI HeLa. Furthermore, intravenous infusion of 1-2 mg of LDL increased liver LDs in normal (WT) but not in SR-BI KO mice. Mice transgenic for human SR-BI demonstrated higher liver LD accumulation than WT mice. Finally, Electro Spray Infusion Mass Spectrometry (ESI-MS) using deuterated d-CE found that LDs accumulated up to 40% of unmodified d-CE LDL. We conclude that SR-BI mediates LDL-induced LD formation in vitro and in vivo. In addition to cytosolic NL hydrolysis and de novo lipid synthesis, this process includes selective sorting and transport of LDL NL to LDs with limited lysosomal NL sequestration and the transport of LDL CE, and TAG directly to LDs independently of de novo synthesis.

Dataset Information

HIF isoforms have divergent effects on invasion, metastasis, metabolism and formation of lipid droplets.

Publications

HIF isoforms have divergent effects on invasion, metastasis, metabolism and formation of lipid droplets.

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