NEET Proteins: A New Link Between Iron Metabolism, Reactive Oxygen Species, and Cancer

Abstract

Significance: Cancer cells accumulate high levels of iron and reactive oxygen species (ROS) to promote their high metabolic activity and proliferation rate. However, high levels of iron and ROS can also lead to enhanced oxidative stress and the activation of cell death pathways such as apoptosis and ferroptosis. This has led to the proposal that different drugs that target iron and/or ROS metabolism could be used as anticancer drugs. However, due to the complex role iron and ROS play in cells, the majority of these drugs yielded mixed results, highlighting a critical need to identify new players in the regulation of iron and ROS homeostasis in cancer cells. Recent Advances: NEET proteins belong to a newly discovered class of iron-sulfur proteins (2Fe-2S) required for the regulation of iron and ROS homeostasis in cells. Recent studies revealed that the NEET proteins NAF-1 (CISD2) and mitoNEET (CISD1) play a critical role in promoting the proliferation of cancer cells, supporting tumor growth and metastasis. Moreover, the function of NEET proteins in cancer cells was found to be dependent of the degree of lability of their 2Fe-2S clusters.

Critical issues: NEET proteins could represent a key regulatory link between the maintenance of high iron and ROS in cancer cells, the activation of cell death and survival pathways, and cellular proliferation.

Future directions: Because the function of NEET proteins depends on the lability of their clusters, drugs that target the 2Fe2S clusters of NEET proteins could be used as promising anticancer drugs.

Keywords: NAF-1; NEET proteins; ROS; cancer; iron–sulfur; mitoNEET.

FIG. 1.

Iron-related cellular pathways of cancer cells relative to normal cells. The iron addiction phenotype of cancer cells is accompanied by alterations in a number of iron-related pathways. These are compared in the figure between normal (left) and cancer cells (right), and include the enhanced expression of the iron-uptake protein TFR1 on the PM of cancer cells and the suppressed accumulation of the iron-secretion protein FPN. These changes, among others, lead to an increase in the cLIP in cancer cells. Iron is then used as a cofactor for the synthesis of different proteins, as well as transported into mitochondria and used for the synthesis of heme and Fe-S moieties and proteins. The enhanced accumulation of iron in cancer cells is also accompanied by higher levels of ROS. Thick and thin arrows indicate high or low flow rate, respectively. Upward facing red and lower facing blue thick arrows indicate enhanced or suppressed expression/flow/accumulation, respectively. CIA system, cytosolic iron–sulfur cluster assembly; cLIP, cytosolic labile iron pool; DMT1, divalent metal transporter 1; ER, endoplasmic reticulum; Fe-S, iron-sulfur; FPN, ferroprotein; ISC system, iron–sulfur cluster biogenesis system; mFt, mitochondrial ferritin; PM, plasma membrane; ROS, reactive oxygen species; TAM, tumor-associated macrophages; TFR1, transferrin receptor-1.

FIG. 2.

Involvement of NEET proteins in mediating the mobilization of iron and Fe-S clusters between the mitochondria and cytosol. The NEET proteins, mNT (colored in red) and NAF-1 (colored in blue), are localized to the outer mitochondrial membrane and the ER and could link between these compartments and the cytosol. The labile nature of the Fe-S cluster of NEET proteins highlights them as ideal candidates for bridging between the mitochondrial and the cytosolic ISC biogenesis system (CIA). MitoNEET and NAF-1 were shown to mobilize 2Fe-2S clusters from the mitochondria to apo-acceptor proteins such as anamorsin and IRP1 in the cytosol. In addition, in cases in which the cluster transfer reactions are not complete, NEET proteins may also mediate the export/import of iron and sulfur ions between the mitochondria to the cytosol. Upward facing red thick arrow indicates enhanced accumulation. 2Fe-2S, iron-sulfur proteins; IRP1, iron regulatory protein 1; ISC, iron–sulfur cluster; MAMs; mitochondria-associated membranes; mNT, mitoNEET.

FIG. 3.

Evidence for the involvement of NEET proteins in cancer. A survey of the cBio portal for Cancer Genomics database for alterations in expression or mutations in DNA sequences of NEET proteins showing that a large number of different tumors are accompanied by alterations in altered expression and/or in mutations in NEET proteins. The analysis was performed with search tools from (www.cbioportal.org).

FIG. 4.

Altered expression of the NEET proteins mNT and NAF-1 affects the levels of iron and ROS in cancer cells. (A) NEET proteins are shown to help reduce the mitochondrial levels of iron and ROS by mobilizing iron–sulfur (Fe-S) clusters from the mitochondria to the cytosol, thereby promoting tumor growth. (B) Suppression of mNT or NAF-1 expression in cancer cells (e.g., via shRNA) is shown to result in the suppressed mobilization of Fe-S clusters from the mitochondria to the cytosol, the enhanced accumulation of iron and ROS in mitochondria, the activation of autophagy and apoptosis, and the suppression of tumor growth. (C) Overexpression of the NEET proteins mNT or NAF-1 in cancer cells is shown to support tumor growth by allowing the efficient mobilization of iron–sulfur clusters from the mitochondria to the cytosol, by blocking apoptosis and autophagy and by inducing a heightened state of oxidative stress tolerance. NEET, the NEET proteins mNT and NAF-1. (17, 33, 59, 61). Fe, iron; S, sulfur; mLIP, mitochondrial labile iron pool; mROS, mitochondrial ROS.

FIG. 5.

Mutations in mNT (e.g., His87 to Cys), or NAF-1 (e.g., His114 to Cys), or drugs that stabilize the clusters of NEET proteins can block the mobilization of iron–sulfur clusters from the mitochondria to the cytosol. This blockage would result in the overaccumulation of iron and ROS in mitochondria (high mLIP and high mROS), the activation of autophagy and apoptosis, and the suppression of tumor growth (17).

FIG. 6.

A model for the function of the NEET proteins mNT and NAF-1 in cancer cells. (A). The elevated expression level of mNT and/or NAF-1 proteins in cancer cells is proposed to alleviate the iron and ROS stress from the mitochondria via iron and/or Fe-S transfer reactions. This allows cancer cells to accumulate high levels of ROS without cellular damage and to promote cellular proliferation (thick arrows). The high level of NEET proteins in cancer cells is also needed to suppress the activation of the high iron-induced apoptosis and ROS-induced apoptosis and autophagy via interactions with BCL-2 and other proteins (red suppressive line). The state of the mNT and/or NAF-1 clusters and its effect on the interaction of NEET proteins with proteins such as BCL-2 are, therefore, proposed to mediate a link between the iron/Fe-S clusters/ROS metabolic state of the cell and regulatory functions that determine life-or-death decisions. (B). Because NEET proteins could undergo oxidation and cluster loss under high oxidative stress conditions or low pH, the requirement for high levels of NEET proteins in cancer cells could also drive a vicious cycle in which NEET proteins accumulate, undergo oxidation that results in cluster transfer/loss (right), and degradation leading to increased labile iron/Fe-S levels and more oxidative stress that requires even more NEET proteins. This process (underlined in B) is shown to maintain high levels of ROS in cells and to drive cancer proliferation. Thick and thin arrows indicate high or low flow rate, respectively. Upward facing red and lower facing blue thick arrows indicate enhanced or suppressed expression/flow/accumulation, respectively. TF, transferrin.