ETHE1 Accelerates Triple-Negative Breast Cancer Metastasis by Activating GCN2/eIF2α/ATF4 Signaling.

Triple-negative breast cancer (TNBC) is the most fatal subtype of breast cancer; however, effective treatment strategies for TNBC are lacking. Therefore, it is important to explore the mechanism of TNBC metastasis and identify its therapeutic targets. Dysregulation of ETHE1 leads to ethylmalonic encephalopathy in humans; however, the role of ETHE1 in TNBC remains elusive. Stable cell lines with ETHE1 overexpression or knockdown were constructed to explore the biological functions of ETHE1 during TNBC progression in vitro and in vivo. Mass spectrometry was used to analyze the molecular mechanism through which ETHE1 functions in TNBC progression. ETHE1 had no impact on TNBC cell proliferation and xenograft tumor growth but promoted TNBC cell migration and invasion in vitro and lung metastasis in vivo. The effect of ETHE1 on TNBC cell migratory potential was independent of its enzymatic activity. Mechanistic investigations revealed that ETHE1 interacted with eIF2α and enhanced its phosphorylation by promoting the interaction between eIF2α and GCN2. Phosphorylated eIF2α in turn upregulated the expression of ATF4, a transcriptional activator of genes involved in cell migration and tumor metastasis. Notably, inhibition of eIF2α phosphorylation through ISRIB or ATF4 knockdown partially abolished the tumor-promoting effect of ETHE1 overexpression. ETHE1 has a functional and mechanistic role in TNBC metastasis and offers a new therapeutic strategy for targeting ETHE1-propelled TNBC using ISRIB.

Dynamic SUMOylation of MORC2 orchestrates chromatin remodelling and DNA repair in response to DNA damage and drives chemoresistance in breast cancer.

Rationale: SUMOylation regulates a plethora of biological processes, and its inhibitors are currently under investigation in clinical trials as anticancer agents. Thus, identifying new targets with site-specific SUMOylation and defining their biological functions will not only provide new mechanistic insights into the SUMOylation signaling but also open an avenue for developing new strategy for cancer therapy. MORC family CW-type zinc finger 2 (MORC2) is a newly identified chromatin-remodeling enzyme with an emerging role in the DNA damage response (DDR), but its regulatory mechanism remains enigmatic. Methods: In vivo and in vitro SUMOylation assays were used to determine the SUMOylation levels of MORC2. Overexpression and knockdown of SUMO-associated enzymes were used to detect their effects on MORC2 SUMOylation. The effect of dynamic MORC2 SUMOylation on the sensitivity of breast cancer cells to chemotherapeutic drugs was examined through in vitro and in vivo functional assays. Immunoprecipitation, GST pull-down, MNase, and chromatin segregation assays were used to explore the underlying mechanisms. Results: Here, we report that MORC2 is modified by small ubiquitin-like modifier 1 (SUMO1) and SUMO2/3 at lysine 767 (K767) in a SUMO-interacting motif dependent manner. MORC2 SUMOylation is induced by SUMO E3 ligase tripartite motif containing 28 (TRIM28) and reversed by deSUMOylase sentrin-specific protease 1 (SENP1). Intriguingly, SUMOylation of MORC2 is decreased at the early stage of DNA damage induced by chemotherapeutic drugs that attenuate the interaction of MORC2 with TRIM28. MORC2 deSUMOylation induces transient chromatin relaxation to enable efficient DNA repair. At the relatively late stage of DNA damage, MORC2 SUMOylation is restored, and SUMOylated MORC2 interacts with protein kinase CSK21 (casein kinase II subunit alpha), which in turn phosphorylates DNA-PKcs (DNA-dependent protein kinase catalytic subunit), thus promoting DNA repair. Notably, expression of a SUMOylation-deficient mutant MORC2 or administration of SUMO inhibitor enhances the sensitivity of breast cancer cells to DNA-damaging chemotherapeutic drugs. Conclusions: Collectively, these findings uncover a novel regulatory mechanism of MORC2 by SUMOylation and reveal the intricate dynamics of MORC2 SUMOylation important for proper DDR. We also propose a promising strategy to sensitize MORC2-driven breast tumors to chemotherapeutic drugs by inhibition of the SUMO pathway.

Inositol monophosphatase 1 (IMPA1) promotes triple-negative breast cancer progression through regulating mTOR pathway and EMT process.

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, which is characterized by high heterogeneity and metabolic dysregulation. Inositol monophosphatase 1(IMPA1) is critical for the metabolism of inositol, which has profound effects on gene expression and other biological processes. Here, we report for the first time that IMPA1 was upregulated in TNBC cell lines and tissues, and enhanced cell colony formation and proliferation in vitro and tumorigenicity in vivo. Additionally, IMPA1 promoted cell motility in vitro and metastatic lung colonization in vivo. Mechanistic investigations by transcriptome sequencing revealed that 4782 genes were differentially expressed between cells with IMPA1 knockdown and control cells. Among the differentially expressed genes after IMPA1 knockdown, five significantly altered genes were verified via qRT-PCR assays. Morerover, we found that the expression profile of those five targets as a gene set was significantly associated with IMPA1 status in TNBC cells. As this gene set was associated with mTOR pathway and epithelial-mesenchymal transition (EMT) process, we further confirmed that IMPA1 induced mTOR activity and EMT process, which at least in part contributed to IMPA1-induced TNBC progression. Collectively, our findings reveal a previously unrecognized role of IMPA1 in TNBC progression and identify IMPA1 as a potential target for TNBC therapy.

TOLLIP-mediated autophagic degradation pathway links the VCP-TMEM63A-DERL1 signaling axis to triple-negative breast cancer progression.

Triple-negative breast cancer (TNBC) is the most challenging breast cancer subtype to treat due to the lack of effective targeted therapies. Transmembrane (TMEM) proteins represent attractive drug targets for cancer therapy, but biological functions of most members of the TMEM family remain unknown. Here, we report for the first time that TMEM63A (transmembrane protein 63A), a poorly characterized TMEM protein with unknown functions in human cancer, functions as a novel oncogene to promote TNBC cell proliferation, migration, and invasion in vitro and xenograft tumor growth and lung metastasis in vivo. Mechanistic investigations revealed that TMEM63A localizes in endoplasmic reticulum (ER) and lysosome membranes, and interacts with VCP (valosin-containing protein) and its cofactor DERL1 (derlin 1). Furthermore, TMEM63A undergoes autophagy receptor TOLLIP-mediated autophagic degradation and is stabilized by VCP through blocking its lysosomal degradation. Strikingly, TMEM63A in turn stabilizes oncoprotein DERL1 through preventing TOLLIP-mediated autophagic degradation. Notably, pharmacological inhibition of VCP by CB-5083 or knockdown of DERL1 partially abolishes the oncogenic effects of TMEM63A on TNBC progression both in vitro and in vivo. Collectively, these findings uncover a previously unknown functional and mechanistic role for TMEM63A in TNBC progression and provide a new clue for targeting TMEM63A-driven TNBC tumors by using a VCP inhibitor.Abbreviations: ATG16L1, autophagy related 16 like 1; ATG5, autophagy related 5; ATP5F1B/ATP5B, ATP synthase F1 subunit beta; Baf-A1, bafilomycin A1; CALCOCO2/NDP52, calcium binding and coiled-coil domain 2; CANX, calnexin; DERL1, derlin 1; EGFR, epidermal growth factor receptor; ER, endoplasmic reticulum; ERAD, endoplasmic reticulum-associated degradation; HSPA8, heat shock protein family A (Hsp70) member 8; IP, immunoprecipitation; LAMP2A, lysosomal associated membrane protein 2; NBR1, NBR1 autophagy cargo receptor; OPTN, optineurin; RT-qPCR, reverse transcription-quantitative PCR; SQSTM1/p62, sequestosome 1; TAX1BP1, Tax1 binding protein 1; TMEM63A, transmembrane protein 63A; TNBC, triple-negative breast cancer; TOLLIP, toll interacting protein; VCP, valosin containing protein.

Dimerization of MORC2 through its C-terminal coiled-coil domain enhances chromatin dynamics and promotes DNA repair.

Decondesation of the highly compacted chromatin architecture is essential for efficient DNA repair, but how this is achieved remains largely unknown. Here, we report that microrchidia family CW-type zinc finger protein 2 (MORC2), a newly identified ATPase-dependent chromatin remodeling enzyme, is required for nucleosome destabilization after DNA damage through loosening the histone-DNA interaction. Depletion of MORC2 attenuates phosphorylated histone H2AX (γH2AX) focal formation, compromises the recruitment of DNA repair proteins, BRCA1, 53BP1, and Rad51, to sites of DNA damage, and consequently reduces cell survival following treatment with DNA-damaging chemotherapeutic drug camptothecin (CPT). Furthermore, we demonstrate that MORC2 can form a homodimer through its C-terminal coiled-coil (CC) domain, a process that is enhanced in response to CPT-induced DNA damage. Deletion of the C-terminal CC domain in MORC2 disrupts its homodimer formation and impairs its ability to destabilize histone-DNA interaction after DNA damage. Consistently, expression of dimerization-defective MORC2 mutant results in impaired the recruitment of DNA repair proteins to damaged chromatin and decreased cell survival after CPT treatment. Together, these findings uncover a new mechanism for MORC2 in modulating chromatin dynamics and DDR signaling through its c-terminal dimerization.