Defective arginine synthesis, due to the silencing of argininosuccinate synthase 1 (ASS1), is a common metabolic vulnerability in cancer, known as arginine auxotrophy. Understanding how arginine depletion kills arginine-auxotrophic cancer cells will facilitate the development of anti-cancer therapeutic strategies. Here we show that depletion of extracellular arginine in arginine-auxotrophic cancer cells causes mitochondrial distress and transcriptional reprogramming. Mechanistically, arginine starvation induces asparagine synthetase (ASNS), depleting these cancer cells of aspartate, and disrupting their malate-aspartate shuttle. Supplementation of aspartate, depletion of mitochondria, and knockdown of ASNS all protect the arginine-starved cells, establishing the causal effects of aspartate depletion and mitochondrial dysfunction on the arginine starvation-induced cell death. Furthermore, dietary arginine restriction reduced tumor growth in a xenograft model of ASS1-deficient breast cancer. Our data challenge the view that ASNS promotes homeostasis, arguing instead that ASNS-induced aspartate depletion promotes cytotoxicity, which can be exploited for anti-cancer therapies.

The role of fatty acid metabolism, including both anabolic and catabolic reactions in cancer has gained increasing attention in recent years. Many studies have shown that aberrant expression of the genes involved in fatty acid synthesis or fatty acid oxidation correlate with malignant phenotypes including metastasis, therapeutic resistance and relapse. Such phenotypes are also strongly associated with the presence of a small percentage of unique cells among the total tumor cell population. This distinct group of cells may have the ability to self-renew and propagate or may be able to develop resistance to cancer therapies independent of genetic alterations. Therefore, these cells are referred to as cancer stem cells/tumor-initiating cells/drug-tolerant persisters, which are often refractory to cancer treatment and difficult to target. Moreover, interconversion between cancer cells and cancer stem cells/tumor-initiating cells/drug-tolerant persisters may occur and makes treatment even more challenging. This review highlights recent findings on the relationship between fatty acid metabolism, cancer stemness and therapeutic resistance and prompts discussion about the potential mechanisms by which fatty acid metabolism regulates the fate of cancer cells and therapeutic resistance.

The DNA damage response (DDR) involves both the control of DNA damage repair and signaling to cell cycle checkpoints. Therefore, unraveling the underlying mechanisms of the DDR is important for understanding tumor suppression and cellular resistance to clastogenic cancer therapeutics. Because the DDR is likely to be influenced by chromatin regulation at the sites of DNA damage, we investigated the role of heterochromatin protein 1 (HP1) during the DDR process. We monitored double-strand breaks (DSBs) using the γH2AX foci marker and found that depleting cells of HP1 caused genotoxic stress, a delay in the repair of DSBs and elevated levels of apoptosis after irradiation. Furthermore, we found that these defects in repair were associated with impaired BRCA1 function. Depleting HP1 reduced recruitment of BRCA1 to DSBs and caused defects in two BRCA1-mediated DDR events: (i) the homologous recombination repair pathway and (ii) the arrest of cell cycle at the G2/M checkpoint. In contrast, depleting HP1 from cells did not affect the non-homologous end-joining (NHEJ) pathway: instead it elevated the recruitment of the 53BP1 NHEJ factor to DSBs. Notably, all three subtypes of HP1 seemed to be almost equally important for these DDR functions. We suggest that the dynamic interaction of HP1 with chromatin and other DDR factors could determine DNA repair choice and cell fate after DNA damage. We also suggest that compromising HP1 expression could promote tumorigenesis by impairing the function of the BRCA1 tumor suppressor.

from conditional knockout mouse models reveal that KAP1-deficiency significantly impairs vital physiological processes, such as immune maturation, stress vulnerability, hepatic metabolism, gamete development and erythropoiesis. In this review, we summarize and evaluate current literatures involving the biochemical and physiological functions of KAP1. In addition, increasing studies on the clinical relevance of KAP1 in cancer will also be discussed. Key words: KRAB domain-associated protein 1; Transcriptional co-repressor; Post-translational modification; Chromatin remodeling; KRAB-containing zinc finger protein Core tip: This review article primarily summarizes the current findings of KAP1/TRIM28/TIF1β, with focuses on its biochemical and physiological functions. Both the canonical transcriptional co-repressor function and the transcriptional-independent roles of KAP1 are discussed in detail. We highlight the post-translational modifications and the compartmentalized localization of KAP1 and suggest that the function of KAP1 could be spatial and temporal regulated in multiple physiological circumstances. Finally, we summarize the clinical relevance of KAP1 in cancer and discuss the possibility to translate the mechanistic studies of KAP1 to human pathophysiology in the future. Abstract KAP1/TRIM28/TIF1β was identified nearly twenty years ago as a universal transcriptional co-repressor because it interacts with a large KRAB-containing zinc finger protein (KRAB-ZFP) transcription factor family. Many studies demonstrate that KAP1 affects gene expression by regulating the transcription of KRAB-ZFP-specific loci, trans-repressing as a transcriptional co-repressor or epigenetically modulating chromatin structure. Emerging evidence suggests that KAP1 also functions independent of gene regulation by serving as a SUMO/ubiquitin E3 ligase or signaling scaffold protein to mediate signal transduction. KAP1 is subjected to multiple post-translational modifications (PTMs), including serine/tyrosine phosphorylation, SUMOylation, and acetylation, which coordinately regulate KAP1 function and its protein abundance. KAP1 is involved in multiple aspects of cellular activities, including DNA damage response, virus replication, cytokine production and stem cell pluripotency. Moreover, knockout of KAP1 results in embryonic lethality, indicating that KAP1 is crucial for embryonic development and possibly impacts a wide-range of (patho)physiological manifestations. Indeed, studies World J Biol Chem 2014 August 26; 5(3): 308-320 ISSN 1949-8454 (online)

Mitochondrial dynamics during nutrient starvation of cancer cells likely exert profound effects on their capability for metastatic progression. Here we report that KAP1 (TRIM28), a transcriptional co-adaptor protein implicated in metastatic progression in breast cancer, is a pivotal regulator of mitochondrial fusion in glucose-starved cancer cells. Diverse metabolic stresses induced Ser473-phosphorylation of KAP1 (pS473-KAP1) in a ROS- and p38-dependent manner. Results from live cell imaging and molecular studies revealed that during the first 6–8 hr of glucose starvation mitochondria initially underwent extensive fusion, but then subsequently fragmented in a pS473-KAP1-dependent manner. Mechanistic investigations using phosphorylation-defective mutants revealed that KAP1 Ser473-phosphorylation limited mitochondrial hyperfusion in glucose-starved breast cancer cells, as driven by downregulation of the mitofusin protein MFN2, leading to reduced oxidative phosphorylation and ROS production. In clinical specimens of breast cancer, reduced expression of MFN2 corresponded to poor prognosis in patients. In a mouse xenograft model of human breast cancer, there was an association in the core region of tumors between MFN2 downregulation and the presence of highly fragmented mitochondria. Collectively, our results suggest that KAP1 Ser473 phosphorylation acts through MFN2 reduction to restrict mitochondrial hyperfusion, thereby contributing to cancer cell survival under conditions of sustained metabolic stress.

Background:The function of KAP1 is regulated by multiple posttranslational modifications during DNA damage response. Results: A previously unidentified arginine-rich motif (ARM) of RNF4 regulates the phosphorylation-induced, SUMO-dependent recruitment and degradation of KAP1. Conclusion:The ARM of RNF4 enhances SIM-SUMO-dependent recruitment. Significance: The bimodular recognition by RNF4 could be critical for fine-tuning substrate selection during DNA damage response and other stress conditions.

The high-mobility group A2 gene (HMGA2) is one of the most frequently amplified genes in human cancers. However, functions of HMGA2 in tumorigenesis are not fully understood due to limited knowledge of its targets in tumor cells. Our study reveals a novel link between HMGA2 and the regulation of human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase, which offers critical insight into how HMGA2 contributes to tumorigenesis. The expression of HMGA2 modulates the expression of hTERT, resulting in cells with enhanced telomerase activities and increased telomere length. Treatment with suberoylanilide hydroxamide (SAHA), a histone deacetylase (HDAC) inhibitor, causes dose-dependent hTERT reporter activation, mimicking HMGA2 overexpression. By interacting with Sp1, HMGA2 interferes with the recruitment of HDAC2 to the hTERT proximal promoter, enhancing localized histone H3-K9 acetylation and thereby stimulating hTERT expression and telomerase activity. Moreover, HMGA2 knockdown by short hairpin HMGA2 in HepG2 cells leads to progressive telomere shortening and a concurrent decrease of steady-state hTERT mRNA levels, attenuating their ability to form colonies in soft agar. Importantly, HMGA2 partially replaces the function of hTERT during the tumorigenic transformation of normal human fibroblasts. These findings are potentially clinically relevant, because HMGA2 expression is reported to be upregulated in a number of human cancers as telomere maintenance is essential for tumorigenesis.

Defects in basal autophagy limit the nutrient supply from recycling of intracellular constituents. Despite our understanding of the prosurvival role of macroautophagy/autophagy, how nutrient deprivation, caused by compromised autophagy, affects oncogenic KRAS-driven tumor progression is poorly understood. Here, we demonstrate that conditional impairment of the autophagy gene Atg5 (atg5-KO) extends the survival of KRAS-driven tumor-bearing mice by 38%. atg5-KO tumors spread more slowly during late tumorigenesis, despite a faster onset. atg5-KO tumor cells displayed reduced mitochondrial function and increased mitochondrial fragmentation. Metabolite profiles indicated a deficiency in the nonessential amino acid asparagine despite a compensatory overexpression of ASNS (asparagine synthetase), key enzyme for de novo asparagine synthesis. Inhibition of either autophagy or ASNS reduced KRAS-driven tumor cell proliferation, migration, and invasion, which was rescued by asparagine supplementation or knockdown of MFF (mitochondrial fission factor). Finally, these observations were reflected in human cancer-derived data, linking ASNS overexpression with poor clinical outcome in multiple cancers. Together, our data document a widespread yet specific asparagine homeostasis control by autophagy and ASNS, highlighting the previously unrecognized role of autophagy in suppressing the metabolic barriers of low asparagine and excessive mitochondrial fragmentation to permit malignant KRAS-driven tumor progression.