C-terminal binding proteins 1 and 2 in traumatic brain injury-induced inflammation and their inhibition as an approach for anti-inflammatory treatment.

Traumatic brain injury (TBI) induces an acute inflammatory response in the central nervous system that involves both resident and peripheral immune cells. The ensuing chronic neuroinflammation causes cell death and tissue damage and may contribute to neurodegeneration. The molecular mechanisms involved in the maintenance of this chronic inflammation state remain underexplored. C-terminal binding protein (CtBP) 1 and 2 are transcriptional coregulators that repress diverse cellular processes. Unexpectedly, we find that the CtBPs can transactivate a common set of proinflammatory genes both in lipopolysaccharide-activated microglia, astrocytes and macrophages, and in a mouse model of the mild form of TBI. We also find that the expression of these genes is markedly enhanced by a single mild injury in both brain and peripheral blood leukocytes in a severity- and time-dependent manner. Moreover, we were able to demonstrate that specific inhibitors of the CtBPs effectively suppress the expression of the CtBP target genes and thus improve neurological outcome in mice receiving single and repeated mild TBIs. This discovery suggests new avenues for therapeutic modulation of the inflammatory response to brain injury.

Inflammation-dependent overexpression of c-Myc enhances CRL4DCAF4 E3 ligase activity and promotes ubiquitination of ST7 in colitis-associated cancer.

Inflammation is well known as an important driver of the initiation of colitis-associated cancer (CAC). Some cytokines, such as IL-6 and TNF-α can activate expression of the oncogene c-Myc (MYC) and regulate its downstream effects. Cullin-RING E3 Ligases (CRLs) are emerging as master regulators controlling tumorigenesis. Here, we demonstrate that two cullin genes, CUL4A and CUL4B, but not other members, are specifically overexpressed in CAC tumour samples and positively correlate with levels of the proinflammatory cytokines IL-1ß and IL-6. In vitro experiments revealed that the transcription factor c-Myc can specifically activate the expression of CUL4A and CUL4B by binding to a conserved site (CACGTG) located in their promoters. Additionally, we found that both CUL4A and CUL4B can form an E3 complex with DNA damage-binding protein 1 (DDB1) and DDB1-CUL4-associated factor 4 (DCAF4). In vitro and in vivo ubiquitination analyses indicate that CRL4DCAF4 E3 ligase specifically directs degradation of ST7 (suppression of tumorigenicity 7). Overexpression of c-Myc in human colon epithelial cells resulted in the accumulation of CUL4A, CUL4B and DCAF4, but degradation of ST7. In contrast, knockdown of c-Myc, CUL4A or CUL4B in the colon adenocarcinoma cell line HT29 caused accumulation of ST7 and inhibition of cell proliferation, colony formation ability and in vivo tumour growth. Collectively, our results provide in vitro and in vivo evidence that c-Myc regulates CRL4DCAF4 E3 ligase activity to mediate ubiquitination of ST7, whose presence is physiologically essential for CAC tumorigenesis. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

CPP-E1A fusion peptides inhibit CtBP-mediated transcriptional repression.

The carboxyl-terminal binding proteins (CtBP) are transcriptional corepressors that regulate the expression of multiple epithelial-specific and pro-apoptotic genes. Overexpression of CtBP occurs in many human cancers where they promote the epithelial-to-mesenchymal transition, stem cell-like features, and cell survival, while knockdown of CtBP in tumor cells results in p53-independent apoptosis. CtBPs are recruited to their target genes by binding to a conserved PXDLS peptide motif present in multiple DNA-binding transcription factors. Disrupting the interaction between CtBP and its transcription factor partners may be a means of altering CtBP-mediated transcriptional repression and a potential approach for cancer therapies. However, small molecules targeting protein-protein interactions have traditionally been difficult to identify. In this study, we took advantage of the fact that CtBP binds to a conserved peptide motif to explore the feasibility of using peptides containing the PXDLS motif fused to cell-penetrating peptides (CPP) to inhibit CtBP function. We demonstrate that these peptides disrupt the ability of CtBP to interact with its protein partner, E1A, in an AlphaScreen assay. Moreover, these peptides can enter both lung carcinoma and melanoma cells, disrupt the interaction between CtBP and a transcription factor partner, and inhibit CtBP-mediated transcriptional repression. Finally, the constitutive expression of one such peptide, Pep1-E1A-WT, in a melanoma cell line reverses CtBP-mediated oncogenic phenotypes including proliferation, migration, and sphere formation and limits tumor growth in vivo. Together, our results suggest that CPP-fused PXDLS-containing peptides can potentially be developed into a research tool or therapeutic agent targeting CtBP-mediated transcriptional events in various biological pathways.

MicroRNA-300 Regulates the Ubiquitination of PTEN through the CRL4BDCAF13 E3 Ligase in Osteosarcoma Cells.

Cullins, critical members of the cullin-RING ubiquitin ligases (CRLs), are often aberrantly expressed in different cancers. However, the underlying mechanisms regarding aberrant expression of these cullins and the specific substrates of CRLs in different cancers are mostly unknown. Here, we demonstrate that overexpressed CUL4B in human osteosarcoma cells forms an E3 complex with DNA damage binding protein 1 (DDB1) and DDB1- and CUL4-associated factor 13 (DCAF13). In vitro and in vivo analyses indicated that the CRL4BDCAF13 E3 ligase specifically recognized the tumor suppressor PTEN (phosphatase and tensin homolog deleted on chromosome 10) for degradation, and disruption of this E3 ligase resulted in PTEN accumulation. Further analyses indicated that miR-300 directly targeted the 3' UTR of CUL4B, and DNA hypermethylation of a CpG island in the miR-300 promoter region contributed to the downregulation of miR-300. Interestingly, ectopic expression of miR-300 or treatment with 5-AZA-2'-deoxycytidine, a DNA methylation inhibitor, decreased the stability of CRL4BDCAF13 E3 ligase and reduced PTEN ubiquitination. By applying in vitro screening to identify small molecules that specifically inhibit CUL4B-DDB1 interaction, we found that TSC01131 could greatly inhibit osteosarcoma cell growth and could disrupt the stability of the CRL4BDCAF13 E3 ligase. Collectively, our findings shed new light on the molecular mechanism of CUL4B function and might also provide a new avenue for osteosarcoma therapy.

Activation of TNF-α/NF-κB axis enhances CRL4BDCAF11 E3 ligase activity and regulates cell cycle progression in human osteosarcoma cells.

Cullin 4B, a member of the Cullins, which serve as scaffolds to facilitate the assembly of E3 ligase complexes, is aberrantly expressed in many cancers, including osteosarcoma. Recently, we observed that CUL4B forms the CRL4BDCAF11 E3 ligase, which specifically ubiquitinates and degrades the cyclin-dependent kinase (CDK) inhibitor p21Cip1 in human osteosarcoma cells. However, the underlying mechanisms regarding the aberrant expression of CUL4B and the upstream members of this signaling pathway are mostly unknown. In this study, we demonstrate that nuclear factor kappaB (NF-κB) is a direct modulator of CUL4B expression. The CUL4B promoter is responsive to several NF-κB subunits, including RelA, RelB, and c-Rel, but not to p50 or p52. Additional studies reveal that the tumor necrosis factor alpha (TNF-α)/NF-κB axis pathway is activated in human osteosarcoma cells. This activation causes both CUL4B and NF-κB subunits to become abundant in the nucleus of human osteosarcoma cells. The down-regulation of individual genes, including TNFR1, RelA, RelB, c-Rel, and CUL4B, or pairs of them, including TNFR1 + RelA, TNFR1 + RelB, TNFR1 + c-Rel, and RelA+CUL4B, has similar effects on cell growth inhibition, colony formation, cell invasion, and in vivo tumor formation, whereas the overexpression of CUL4B in these knockdown cells significantly reverses their phenotypes. The inhibition of the TNF-α/NF-κB pathway greatly attenuates CRL4BDCAF11 E3 ligase activity and causes the accumulation of p21Cip1 , thereby leading to cell cycle arrest at the S phase. Taken together, our results support a model in which the activation of the TNF-α/NF-κB axis contributes to an increase in CRL4BDCAF11 activity and a decrease in p21Cip1 protein levels, thereby controlling cell cycle progression in human osteosarcoma cells.

DNA Methylation Mediated Downregulation of miR-449c Controls Osteosarcoma Cell Cycle Progression by Directly Targeting Oncogene c-Myc.

MicroRNAs (miRNAs) are critical regulators of gene expression, and they have broad roles in the pathogenesis of different diseases including cancer. Limited studies and expression profiles of miRNAs are available in human osteosarcoma cells. By applying a miRNA microarray analysis, we observed a number of miRNAs with abnormal expression in cancerous tissues from osteosarcoma patients. Of particular interest in this study was miR-449c, which was significantly downregulated in osteosarcoma cells and patients, and its expression was negatively correlated with tumor size and tumor MSTS stages. Ectopic expression of miR-449c significantly inhibited osteosarcoma cell proliferation and colony formation ability, and caused cell cycle arrest at the G1 phase. Further analysis identified that miR-449c was able to directly target the oncogene c-Myc and negatively regulated its expression. Overexpression of c-Myc partially reversed miR-449c-mimic-inhibited cell proliferation and colony formation. Moreover, DNA hypermethylation was observed in two CpG islands adjacent to the genomic locus of miR-449c in osteosarcoma cells. Conversely, treatment with the DNA methylation inhibitor AZA caused induction of miR-449c. In conclusion, our results support a model that DNA methylation mediates downregulation of miR-449c, diminishing miR-449c mediated inhibition of c-Myc and thus leading to the activation of downstream targets, eventually contributing to osteosarcoma tumorigenesis.

The Emerging Roles of Forkhead Box (FOX) Proteins in Osteosarcoma.

Osteosarcoma is the most common bone cancer primarily occurring in children and young adults. Over the past few years, the deregulation of a superfamily transcription factors, known as forkhead box (FOX) proteins, has been demonstrated to contribute to the pathogenesis of osteosarcoma. Molecular mechanism studies have demonstrated that FOX family proteins participate in a variety of signaling pathways and that their expression can be regulated by multiple factors. The dysfunction of FOX genes can alter osteosarcoma cell differentiation, metastasis and progression. In this review, we summarized the evidence that FOX genes play direct or indirect roles in the development and progression of osteosarcoma, and evaluated the emerging role of FOX proteins as targets for therapeutic intervention.

Autophagic degradation of FOXO3a represses the expression of PUMA to block cell apoptosis in cisplatin-resistant osteosarcoma cells.

Autophagy and apoptosis are the two major modes of cell death, and autophagy usually inhibits apoptosis. The current understanding has shown that there is a complex crosstalk between the components of these two pathways. Here, we describe a transcriptional mechanism that links autophagy to apoptosis. We show that the cisplatin-resistant MG63-R12 and U2OS-R5 osteosarcoma sublines, in comparison to their parental MG63 and U2OS cells, respectively, exhibit increased autophagy but decreased apoptosis levels after treatment with cisplatin. We then used a microarray assay to examine the gene expression changes in these two cisplatin-resistant sublines and found that the expression of the transcription factor FOXO3a was dramatically decreased. Pharmacological treatment with either 3-methyladenine to inhibit autophagy or with rapamycin to activate autophagy in these two cisplatin-resistant sublines resulted in the accumulation or degradation of FOXO3a, respectively. Ectopic expression of FOXO3a in MG63-R12 and U2OS-R5 cells significantly enhanced cell sensitivity to cisplatin through a mechanism in which FOXO3a directly binds to the PUMA promoter and activates its expression, as well as its downstream event, the intrinsic apoptosis pathway. Importantly, this overexpression resulted in tumor growth inhibition in vivo. In conclusion, our results provide new insights into the molecular link between autophagy and apoptosis that involves a FOXO3a-mediated transcriptional mechanism. Importantly, our results may facilitate the development of therapeutic strategies for osteosarcoma patients who have become resistant to cisplatin therapy.

The diferric-tyrosyl radical cluster of ribonucleotide reductase and cytosolic iron-sulfur clusters have distinct and similar biogenesis requirements.

How each metalloprotein assembles the correct metal at the proper binding site presents challenges to the cell. The di-iron enzyme ribonucleotide reductase (RNR) uses a diferric-tyrosyl radical (FeIII2-Y•) cofactor to initiate nucleotide reduction. Assembly of this cofactor requires O2, FeII, and a reducing equivalent. Recent studies show that RNR cofactor biosynthesis shares the same source of iron, in the form of [2Fe-2S]-GSH2 from the monothiol glutaredoxin Grx3/4, and the same electron source, in the form of the Dre2-Tah18 electron transfer chain, with the cytosolic iron-sulfur protein assembly (CIA) machinery required for maturation of [4Fe-4S] clusters in cytosolic and nuclear proteins. Here, we further investigated the interplay between the formation of the FeIII2-Y• cofactor in RNR and the cellular iron-sulfur (Fe-S) protein biogenesis pathways by examining both the iron loading into the RNR ß subunit and the RNR catalytic activity in yeast mutants depleted of individual components of the mitochondrial iron-sulfur cluster assembly (ISC) and the CIA machineries. We found that both iron loading and cofactor assembly in RNR are dependent on the ISC machinery. We also found that Dre2 is required for RNR cofactor formation but appears to be dispensable for iron loading. None of the CIA components downstream of Dre2 was required for RNR cofactor formation. Thus, the pathways for RNR and Fe-S cluster biogenesis bifurcate after the Dre2-Tah18 step. We conclude that RNR cofactor biogenesis requires the ISC machinery to mature the Grx3/4 and Dre2 Fe-S proteins, which then function in iron and electron delivery to RNR, respectively.

DNA Methylation Mediated Down-Regulation of miR-370 Regulates Cell Growth through Activation of the Wnt/ß-Catenin Signaling Pathway in Human Osteosarcoma Cells.

MicroRNA-370 (miR-370) has been observed to act as a tumor suppressor through the targeting of different proteins in a variety of tumors. Our previous study indicated that miR-370 was able to target forkhead box protein M1 (FOXM1) to inhibit cell growth and metastasis in human osteosarcoma cells. In this study, we reported that FOXM1 interacted with ß-catenin in vitro and in vivo. Similar to FOXM1, critical components of the Wnt signaling pathway, including ß-catenin, c-Myc, and Cyclin D1, were also highly expressed in different human osteosarcoma cells lines. Pharmacological inhibition of FOXM1 or ß-catenin but not of c-Myc was associated with the increased expression of miR-370. Ectopic expression of miR-370 inhibited the downstream signaling of ß-catenin. Moreover, osteosarcoma cells treated with 5-AZA-2'-deoxycytidine (AZA), a DNA methylation inhibitor, exhibited increased levels of miR-370 and decreased levels of ß-catenin downstream targets, which resulted in inhibition of cell proliferation and colony formation ability. In conclusion, our results supported a model in which the DNA methylation-mediated down-regulation of miR-370 reduced its inhibitory effect on FOXM1, thereby promoting FOXM1-ß-catenin interaction and activating the Wnt/ß-Catenin signaling pathway in human osteosarcoma cells.

CRL4BDCAF11 E3 ligase targets p21 for degradation to control cell cycle progression in human osteosarcoma cells.

Cell cycle progression in mammals is strictly controlled by a number of cyclin-dependent kinases (CDKs) and CDK inhibitors (CKIs), the expression of which is often dysregulated in cancer cells. Our previous work revealed that Cullin 4B (CUL4B), a critical component of the Cullin4B-RING E3 ligase complex (CRL4B), is overexpressed in human osteosarcoma cells through an unknown mechanism. Here, we demonstrated that CUL4B forms an E3 ligase with RBX1 (RING-box 1), DDB1 (DNA damage binding protein 1), and DCAF11 (DDB1 and CUL4 associated factor 11) in human osteosarcoma cells. In vitro and in vivo ubiquitination analyses indicated that CRL4BDCAF11 E3 ligase was able to specifically ubiquitinate a CDK inhibitor-p21Cip1 at K16, K154, K161 and K163 but not at K75 and K141. Knocking down any component of the CRL4BDCAF11 complex, including CUL4B, DDB1 or DCAF11, using short hairpin RNAs (shRNAs) attenuated the ubiquitination level of p21Cip1, inhibited osteosarcoma cell proliferation, led to cell cycle arrest at S phase, and decreased colony formation rate. Taken together, our data suggest that the CRL4BDCAF11 complex represents a unique E3 ligase that promotes the ubiquitination of p21Cip1 and regulates cell cycle progression in human osteosarcoma cells.

MiR-34a and miR-203 Inhibit Survivin Expression to Control Cell Proliferation and Survival in Human Osteosarcoma Cells.

Elevated expression of survivin is observed in a number of cancer types, including human osteosarcoma. Few studies have demonstrated that survivin expression levels can be considered an independent predictor of survival for human osteosarcoma patients. However, the underlying molecular mechanisms of survivin in the process of human osteosarcoma carcinogenesis remain unclear. In the current study, we evaluated the biological effects of survivin knockdown on osteosarcoma cell proliferation, colony formation rate, and sensitivity to the chemotherapeutic agent cisplatin. We found that two different osteosarcoma cell lines, U2OS and Saos-2, have relatively higher expression levels of survivin, and specific knockdown of survivin resulted in a number of effects, such as inhibition of cell proliferation, decreased colony formation rate, cell cycle arrest at G2/M phase, induction of apoptosis, and increased sensitivity to cisplatin. In addition, we identified two microRNAs, miR-34a and miR-203, that are aberrantly expressed in human osteosarcoma cells and specifically target survivin by inhibiting its expression, therefore repressing osteosarcoma cell maintenance and proliferation.

Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Small Subunit Localization in Response to Iron Deficiency.

Ribonucleotide reductase (RNR) is an essential iron-dependent enzyme that catalyzes deoxyribonucleotide synthesis in eukaryotes. Living organisms have developed multiple strategies to tightly modulate RNR function to avoid inadequate or unbalanced deoxyribonucleotide pools that cause DNA damage and genome instability. Yeast cells activate RNR in response to genotoxic stress and iron deficiency by facilitating redistribution of its small heterodimeric subunit Rnr2-Rnr4 from the nucleus to the cytoplasm, where it forms an active holoenzyme with large Rnr1 subunit. Dif1 protein inhibits RNR by promoting nuclear import of Rnr2-Rnr4. Upon DNA damage, Dif1 phosphorylation by the Dun1 checkpoint kinase and its subsequent degradation enhances RNR function. In this report, we demonstrate that Dun1 kinase triggers Rnr2-Rnr4 redistribution to the cytoplasm in response to iron deficiency. We show that Rnr2-Rnr4 relocalization by low iron requires Dun1 kinase activity and phosphorylation site Thr-380 in the Dun1 activation loop, but not the Dun1 forkhead-associated domain. By using different Dif1 mutant proteins, we uncover that Dun1 phosphorylates Dif1 Ser-104 and Thr-105 residues upon iron scarcity. We observe that the Dif1 phosphorylation pattern differs depending on the stimuli, which suggests different Dun1 activating pathways. Importantly, the Dif1-S104A/T105A mutant exhibits defects in nucleus-to-cytoplasm redistribution of Rnr2-Rnr4 by iron limitation. Taken together, these results reveal that, in response to iron starvation, Dun1 kinase phosphorylates Dif1 to stimulate Rnr2-Rnr4 relocalization to the cytoplasm and promote RNR function.

Survivin and Tumorigenesis: Molecular Mechanisms and Therapeutic Strategies.

Survivin is the smallest member of the inhibitor of apoptosis protein family, which has key roles in regulating cell division and inhibiting apoptosis by blocking caspase activation. Survivin is highly expressed in most human cancers, such as lung, pancreatic and breast cancers, relative to normal tissues. Aberrant survivin expression is associated with tumor cell proliferation, progression, angiogenesis, therapeutic resistance, and poor prognosis. Studies on the underlying molecular mechanisms indicate that survivin is involved in the regulation of cytokinesis and cell cycle progression, as well as participates in a variety of signaling pathways such as the p53, Wnt, hypoxia, transforming growth factor, and Notch signaling pathways. In this review, recent progress in understanding the molecular basis of survivin is discussed. Therapeutic strategies targeting survivin in preclinical studies are also briefly summarized.

Long noncoding RNAs and tumorigenesis: genetic associations, molecular mechanisms, and therapeutic strategies.

The human genome contains a large number of nonprotein-coding sequences. Recently, new discoveries in the functions of nonprotein-coding sequences have demonstrated that the "Dark Genome" significantly contributes to human diseases, especially with regard to cancer. Of particular interest in this review are long noncoding RNAs (lncRNAs), which comprise a class of nonprotein-coding transcripts that are longer than 200 nucleotides. Accumulating evidence indicates that a large number of lncRNAs exhibit genetic associations with tumorigenesis, tumor progression, and metastasis. Our current understanding of the molecular bases of these lncRNAs that are associated with cancer indicate that they play critical roles in gene transcription, translation, and chromatin modification. Therapeutic strategies based on the targeting of lncRNAs to disrupt their expression or their functions are being developed. In this review, we briefly summarize and discuss the genetic associations and the aberrant expression of lncRNAs in cancer, with a particular focus on studies that have revealed the molecular mechanisms of lncRNAs in tumorigenesis. In addition, we also discuss different therapeutic strategies that involve the targeting of lncRNAs.

The late-annotated small ORF LSO1 is a target gene of the iron regulon of Saccharomyces cerevisiae.

We have identified a new downstream target gene of the Aft1/2-regulated iron regulon in budding yeast Saccharomyces cerevisiae, the late-annotated small open reading frame LSO1. LSO1 transcript is among the most highly induced from a transcriptome analysis of a fet3-1 mutant grown in the presence of the iron chelator bathophenanthrolinedisulfonic acid. LSO1 has a paralog, LSO2, which is constitutively expressed and not affected by iron availability. In contrast, we find that the LSO1 promoter region contains three consensus binding sites for the Aft1/2 transcription factors and that an LSO1-lacZ reporter is highly induced under low-iron conditions in a Aft1-dependent manner. The expression patterns of the Lso1 and Lso2 proteins mirror those of their mRNAs. Both proteins are localized to the nucleus and cytoplasm, but become more cytoplasmic upon iron deprivation consistent with a role in iron transport. LSO1 and LSO2 appear to play overlapping roles in the cellular response to iron starvation since single lso1 and lso2 mutants are sensitive to iron deprivation and this sensitivity is exacerbated when both genes are deleted.

Targeting cancer with sesterterpenoids: the new potential antitumor drugs.

Cancer remains a major cause of death in the world to date. A variety of anticancer drugs have been used in clinical chemotherapy, acting on the particular oncogenic abnormalities that are responsible for malignant transformation and progression. Interestingly, some of these anticancer drugs are developed from natural sources such as plants, marine organisms, and microorganisms. Over the past decades, a family of naturally occuring molecules, namely sesterterpenoids, has been isolated from different organisms and they exhibit significant potential in the inhibition of tumor cells in vitro, while the molecular targets of these compounds and their functional mechanisms are still obscure. In this review, we summarize and discuss the functions of these sesterterpenoids in the inhibition of cancer cells. Moreover, we also highlight and discuss chemical structure-activity relationships of some compounds, demonstrating their pervasiveness and importance in cancer therapy.

The Multifunctions of WD40 Proteins in Genome Integrity and Cell Cycle Progression.

Eukaryotic genome encodes numerous WD40 repeat proteins, which generally function as platforms of protein-protein interactions and are involved in numerous biological process, such as signal transduction, gene transcriptional regulation, protein modifications, cytoskeleton assembly, vesicular trafficking, DNA damage and repair, cell death and cell cycle progression. Among these diverse functions, genome integrity maintenance and cell cycle progression are extremely important as deregulation of them is clinically linked to uncontrolled proliferative diseases such as cancer. Thus, we mainly summarize and discuss the recent understanding of WD40 proteins and their molecular mechanisms linked to genome stability and cell cycle progression in this review, thereby demonstrating their pervasiveness and importance in cellular networks.

Cullin family proteins and tumorigenesis: genetic association and molecular mechanisms.

Cullin family proteins function as scaffolds to form numerous E3 ubiquitin ligases with RING proteins, adaptor proteins and substrate recognition receptors. These E3 ligases further recognize numerous substrates to participate in a variety of cellular processes, such as DNA damage and repair, cell death and cell cycle progression. Clinically, cullin-associated E3 ligases have been identified to involve numerous human diseases, especially with regard to multiple cancer types. Over the past few years, our understanding of cullin proteins and their functions in genome stability and tumorigenesis has expanded enormously. Herein, this review briefly provides current perspectives on cullin protein functions, and mainly summarizes and discusses molecular mechanisms of cullin proteins in tumorigenesis.

Involvement of Iron-Containing Proteins in Genome Integrity in Arabidopsis Thaliana.

The Arabidopsis genome encodes numerous iron-containing proteins such as iron-sulfur (Fe-S) cluster proteins and hemoproteins. These proteins generally utilize iron as a cofactor, and they perform critical roles in photosynthesis, genome stability, electron transfer, and oxidation-reduction reactions. Plants have evolved sophisticated mechanisms to maintain iron homeostasis for the assembly of functional iron-containing proteins, thereby ensuring genome stability, cell development, and plant growth. Over the past few years, our understanding of iron-containing proteins and their functions involved in genome stability has expanded enormously. In this review, I provide the current perspectives on iron homeostasis in Arabidopsis, followed by a summary of iron-containing protein functions involved in genome stability maintenance and a discussion of their possible molecular mechanisms.