TRIM2 promotes metabolic adaptation to glutamine deprivation via enhancement of CPT1A activity.

Cancer cells undergo metabolic adaptation to promote their survival and growth under energy stress conditions, yet the underlying mechanisms remain largely unclear. Here, we report that tripartite motif-containing protein 2 (TRIM2) is upregulated in response to glutamine deprivation by the transcription factor cyclic AMP-dependent transcription factor (ATF4). TRIM2 is shown to specifically interact with carnitine O-palmitoyltransferase 1 (CPT1A), a rate-limiting enzyme of fatty acid oxidation. Via this interaction, TRIM2 enhances the enzymatic activity of CPT1A, thereby regulating intracellular lipid levels and protecting cells from glutamine deprivation-induced apoptosis. Furthermore, TRIM2 is able to promote both in vitro cell proliferation and in vivo xenograft tumor growth via CPT1A. Together, these findings establish TRIM2 as an important regulator of the metabolic adaptation of cancer cells to glutamine deprivation and implicate TRIM2 as a potential therapeutic target for cancer.

Energy stress-induced circDDX21 promotes glycolysis and facilitates hepatocellular carcinogenesis.

Cancer cells undergo metabolic reprogramming in response to hostile microenvironments, such as energy stress; however, the underlying mechanisms remain largely unclear. It is also unknown whether energy stress-responsive circular RNA (circRNA) is involved in the regulation of glucose metabolism. Here we report that circDDX21 is upregulated in response to glucose deprivation by the transcription factor c-Myc. Functionally, circDDX21 is shown to promote glycolysis by increasing PGAM1 expression. Mechanistically, circDDX21 interacts with the RNA binding protein PABPC1, disrupting its association with the ubiquitin E3 ligase MKRN3. This disassociation attenuates MKRN3-mediated PABPC1 ubiquitination and enhances the binding of PABPC1 to PGAM1 mRNA, thereby leading to PGAM1 mRNA stabilization. The ability of the circDDX21-PGAM1 axis to promote hepatocellular carcinogenesis is validated in a xenograft mouse model. Additionally, in clinical hepatocellular carcinoma tissues, there is a positive correlation between circDDX21 and PGAM1 expression. These findings establish circDDX21 as an important regulator of glycolysis and suggest circDDX21 as a potential therapeutic target for hepatocellular carcinoma.

MLF2 Negatively Regulates P53 and Promotes Colorectal Carcinogenesis.

Inactivation of the p53 pathway is linked to a variety of human cancers. As a critical component of the p53 pathway, ubiquitin-specific protease 7 (USP7) acts as a deubiquitinase for both p53 and its ubiquitin E3 ligase mouse double minute 2 homolog. Here, myeloid leukemia factor 2 (MLF2) is reported as a new negative regulator of p53. MLF2 interacts with both p53 and USP7. Via these interactions, MLF2 inhibits the binding of USP7 to p53 and antagonizes USP7-mediated deubiquitination of p53, thereby leading to p53 destabilization. Functionally, MLF2 plays an oncogenic role in colorectal cancer, at least partially, via the negative regulation of p53. Clinically, MLF2 is elevated in colorectal cancer and its high expression is associated with poor prognosis in patients with colorectal cancer. In wild-type-p53-containing colorectal cancer, MLF2 and p53 expressions are inversely correlated. These findings establish MLF2 as an important suppressor of p53 function. The study also reveals a critical role for the MLF2-p53 axis in promoting colorectal carcinogenesis.

c-Myc-induced long noncoding RNA MIRE cooperates with hnRNPK to stabilize ELF2 mRNA and promotes clear cell renal cell carcinogenesis.

Elevated expression of c-Myc is associated with a variety of human cancers including clear cell renal cell carcinoma (ccRCC). Increasing evidence suggests that long noncoding RNAs (lncRNAs) are an important class of molecules that regulate both tumor initiation and progression. Here, we report the lncRNA c-Myc-induced regulator of ELF2 (MIRE) as a transcriptional target of c-Myc. MIRE functions as an oncogenic molecule in ccRCC by increasing ELF2 expression. Mechanistically, MIRE promotes phase separation of the RNA binding protein hnRNPK and facilitates the binding of hnRNPK to ELF2 mRNA, thereby resulting in the stabilization of ELF2 mRNA. Interestingly, MIRE is also under transcriptional control by ELF2, establishing an ELF2-MIRE positive feedback loop. Together, these findings provide new insights into the mechanisms by which c-Myc promotes tumorigenesis. They also implicate MIRE as an important regulator of ccRCC carcinogenesis.

The RNA binding protein RALY suppresses p53 activity and promotes lung tumorigenesis.

The tumor suppressor p53 plays a pivotal role in tumor prevention. The activity of p53 is mainly restrained by the ubiquitin E3 ligase Mdm2. However, it is not well understood how the Mdm2-p53 pathway is intricately regulated. Here we report that the RNA binding protein RALY functions as an oncogenic factor in lung cancer. RALY simultaneously binds to Mdm2 and the deubiquitinating enzyme USP7. Via these interactions, RALY not only stabilizes Mdm2 by stimulating the deubiquitinating activity of USP7 toward Mdm2 but also increases the trans-E3 ligase activity of Mdm2 toward p53. Consequently, RALY enhances Mdm2-mediated ubiquitination and degradation of p53. Functionally, RALY promotes lung tumorigenesis, at least partially, via negative regulation of p53. These findings suggest that RALY destabilizes p53 by modulating the function of Mdm2 at multiple levels. Our study also indicates a critical role for RALY in promoting lung tumorigenesis via p53 inhibition.

Decoding transcriptional regulation via a human gene expression predictor.

Transcription factors (TFs) regulate cellular activities by controlling gene expression, but a predictive model describing how TFs quantitatively modulate human transcriptomes is lacking. We construct a universal human gene expression predictor named EXPLICIT-Human and utilize it to decode transcriptional regulation. Using the expression of 1613 TFs, the predictor reconstitutes highly accurate transcriptomes for samples derived from a wide range of tissues and conditions. The broad applicability of the predictor indicates that it recapitulates the quantitative relationships between TFs and target genes ubiquitous across tissues. Significant interacting TF-target gene pairs are extracted from the predictor and enable downstream inference of TF regulators for diverse pathways involved in development, immunity, metabolism, and stress response. A detailed analysis of the hematopoiesis process reveals an atlas of key TFs regulating the development of different hematopoietic cell lineages, and a portion of these TFs are conserved between humans and mice. The results demonstrate that our method is capable of delineating the TFs responsible for fate determination. Compared to other existing tools, EXPLICIT-Human shows a better performance in recovering the correct TF regulators.

The c-Myc targeting hnRNPAB promotes lung adenocarcinoma cell proliferation via stabilization of CDK4 mRNA.

The c-Myc oncoprotein plays a pivotal role in tumorigenesis. The deregulated expression of c-Myc has been linked to a variety of human cancers including lung adenocarcinoma. The oncogenic function of c-Myc has been largely attributed to its intrinsic nature as a transcription factor. Here we reported the RNA binding protein hnRNPAB as a direct transcriptional target of c-Myc by performing quantitative real-time polymerase chain reaction (qRT-PCR), western blot, chromatin immunoprecipitation (ChIP), and luciferase reporter analyses. Flow cytometry, colony formation, and RNA immunoprecipitation (RIP) assays were used to investigate the role of hnRNPAB in lung adenocarcinoma cell proliferation, as well as the underlying mechanism. HnRNPAB was functionally shown to promote lung adenocarcinoma cell proliferation by accelerating G1/S cell cycle progression. Mechanistically, hnRNPAB interacted with and stabilized CDK4 mRNA, thereby increasing CDK4 expression. Moreover, hnRNPAB was able to promote G1/S cell cycle progression and cell proliferation via the regulation of CDK4. HnRNPAB was also revealed as a mediator of the promoting effect of c-Myc on cell proliferation. Together, these findings demonstrate that hnRNPAB is an important regulator of lung adenocarcinoma cell proliferation. They also add new insights into the mechanisms of how c-Myc promotes tumorigenesis.

Reciprocal modulation of long noncoding RNA EMS and p53 regulates tumorigenesis.

p53 plays a central role in tumor suppression. Emerging evidence suggests long noncoding RNA (lncRNA) as an important class of regulatory molecules that control the p53 signaling. Here, we report that the oncogenic lncRNA E2F1 messenger RNA (mRNA) stabilizing factor (EMS) and p53 mutually repress each other's expression. EMS is negatively regulated by p53. As a direct transcriptional repression target of p53, EMS is surprisingly shown to inhibit p53 expression. EMS associates with cytoplasmic polyadenylation element-binding protein 2 (CPEB2) and thus, disrupts the CPEB2-p53 mRNA interaction. This disassociation attenuates CPEB2-mediated p53 mRNA polyadenylation and suppresses p53 translation. Functionally, EMS is able to exert its oncogenic activities, at least partially, via the CPEB2-p53 axis. Together, these findings reveal a double-negative feedback loop between p53 and EMS, through which p53 is finely controlled. Our study also demonstrates a critical role for EMS in promoting tumorigenesis via the negative regulation of p53.

c-Myc-activated USP2-AS1 suppresses senescence and promotes tumor progression via stabilization of E2F1 mRNA.

The c-Myc oncoprotein plays a prominent role in cancer initiation, progression, and maintenance. Long noncoding RNAs (lncRNAs) are recently emerging as critical regulators of the c-Myc signaling pathway. Here, we report the lncRNA USP2-AS1 as a direct transcriptional target of c-Myc. Functionally, USP2-AS1 inhibits cellular senescence and acts as an oncogenic molecule by inducing E2F1 expression. Mechanistically, USP2-AS1 associates with the RNA-binding protein G3BP1 and facilitates the interaction of G3BP1 to E2F1 3'-untranslated region, thereby leading to the stabilization of E2F1 messenger RNA. Furthermore, USP2-AS1 is shown as a mediator of the oncogenic function of c-Myc via the regulation of E2F1. Together, these findings suggest that USP2-AS1 is a negative regulator of cellular senescence and also implicates USP2-AS1 as an important player in mediating c-Myc function.

Energy stress-induced linc01564 activates the serine synthesis pathway and facilitates hepatocellular carcinogenesis.

Cancer cells undergo metabolic adaption to sustain their survival and growth under metabolic stress conditions, yet the underlying mechanism remains largely unclear. It is also not known if lncRNAs contribute to this metabolic adaption of cancer cells. Here we show that linc01564 is induced in response to glucose deprivation by the transcription factor ATF4. Linc01564 functions to facilitate hepatocellular carcinoma cell survival under glucose deprivation by activating the serine synthesis pathway. Mechanistically, linc01564 acts as a competing endogenous RNA for miR-107/103a-3p and attenuates the inhibitory effect of miR-107/103a-3p on PHGDH, the rate-limiting enzyme of the serine synthesis pathway, thereafter leading to increased PHGDH expression. Furthermore, linc01564 is able to promote hepatocellular carcinogenesis via PHGDH. Together, these findings suggest that linc01564 is an important player in the regulation of metabolic adaption of cancer cells and also implicate linc01564 as a potential therapeutic target for cancer.

Tumor Immune Microenvironment Characteristics and Their Prognostic Value in Non-Small-Cell Lung Cancer.

INTRODUCTION: Cancer progression is determined not only by the malignant behavior of tumors but also by the immune microenvironment. The tumor immune microenvironment also plays a pivotal role in determining the clinical response of non-small-cell lung cancer (NSCLC) to immunotherapies. To understand the possible mechanisms and explore new targets in lung cancer immunotherapy, we characterized the immune profiles in NSCLC patients. METHODS: Seventy-one NSCLC patients who underwent radical resection were selected. The immune cell composition in paired tumor and adjacent normal lung tissues was tested by flow cytometry. The associations of tumor immune microenvironment characteristics with clinicopathological factors and overall survival were analyzed. Kaplan-Meier curves and Cox proportional hazards models were used to determine differences in survival. RESULTS: Compared with adjacent normal lung tissues, an increased proportion of CD45+ hematopoietic-derived cells, CD4+ T cell subtypes, Tregs and B cells was observed in tumor samples with a reduced frequency of myeloid cell populations. There was no significant increase in total CD8+ T cells, but both PD1+ and CD38+ CD8+ T cells were significantly enriched in tumor samples and statistically significantly associated with tumor size. In addition, positive CD38 expression was highly correlated with PD1 positivity. A high proportion of CD8+ T cells and a low percentage of PD1+ CD8+ T cells were statistically significantly associated with better survival in stage II and III patients, whereas a low frequency of CD38+ CD8+ T cells was statistically significantly associated with better survival in all patients and identified as an independent prognostic factor (p=0.049). CONCLUSION: We profiled the immune cells in the tumor tissues of NSCLC patients using flow cytometry. The results revealed significant enrichment of infiltrating immune cells. A strong correlation was identified between CD38 and PD-1 expression on CD8+ T cells in tumors. CD8+ T cells and their subtypes play a critical role in the prediction of prognosis.

Identified lung adenocarcinoma metabolic phenotypes and their association with tumor immune microenvironment.

BACKGROUND: Lung adenocarcinoma (LUAD), a subtype of non-small cell lung cancer (NSCLC), causes high mortality around the world. Previous studies have suggested that the metabolic pattern of tumor is associated with tumor response to immunotherapy and patient's survival outcome. Yet, this relationship in LUAD is still unknown. METHODS: Therefore, in this study, we identified the immune landscape in different tumor subtypes classified by metabolism-related genes expression with a large-scale dataset (tumor samples, n = 2181; normal samples, n = 419). We comprehensively correlated metabolism-related phenotypes with diverse clinicopathologic characteristics, genomic features, and immunotherapeutic efficacy in LUAD patients. RESULTS: And we confirmed tumors with activated lipid metabolism tend to have higher immunocytes infiltration and better response to checkpoint immunotherapy. This work highlights the connection between the metabolic pattern of tumor and tumor immune infiltration in LUAD. A scoring system based on metabolism-related gene expression is not only able to predict prognosis of patient with LUAD but also applied to pan-cancer. LUAD response to checkpoint immunotherapy can also be predicted by this scoring system. CONCLUSIONS: This work revealed the significant connection between metabolic pattern of tumor and tumor immune infiltration, regulating LUAD patients' response to immunotherapy.

Erratum: LukS-PV Inhibits Hepatocellular Carcinoma Progression by Downregulating HDAC2 Expression.

[This corrects the article DOI: 10.1016/j.omto.2020.05.006.].

LukS-PV Inhibits Hepatocellular Carcinoma Progression by Downregulating HDAC2 Expression.

Hepatocellular carcinoma (HCC) is a common malignant tumor. LukS-PV is the S component of Panton-Valetine leukocidin (PVL), which is secreted by Staphylococcus aureus. This study investigated the effects of LukS-PV on the proliferation, apoptosis, and cell-cycle progression of HCC cells and the mechanisms of its activity. The HCC cells were treated with different LukS-PV concentrations in vitro. Cell Counting Kit-8 and 5-Ethynyl-2'-deoxyuridine (EdU) assays were used to study cell proliferation. Flow cytometry was used to measure apoptosis and cell-cycle progression. Quantitative reverse transcriptase PCR and western blot assays were used to determine mRNA and protein expression levels. Xenograft experiments were performed to determine the in vivo antitumor effect of LukS-PV. Immunostaining was performed to analyze Ki-67 and HDAC2 (histone deacetylase 2) expression. Our results showed that LukS-PV inhibited cell proliferation and induced apoptosis in a concentration-dependent manner in HCC cell lines. LukS-PV also can induce cell-cycle arrest. Moreover, we discovered that LukS-PV attenuated HDAC2 expression and upregulated PTEN; phosphorylated AKT was also reduced. Further studies demonstrated that LukS-PV treatment significantly reduced tumor growth in nude mice and suppressed Ki-67 and HDAC2 levels. Our data revealed a vital role of LukS-PV in suppressing HCC progression by downregulating HDAC2 and upregulating PTEN.

WDR63 inhibits Arp2/3-dependent actin polymerization and mediates the function of p53 in suppressing metastasis.

Accumulating evidence suggests that p53 plays a suppressive role in cancer metastasis, yet the underlying mechanism remains largely unclear. Regulation of actin dynamics is essential for the control of cell migration, which is an important step in metastasis. The Arp2/3 complex is a major nucleation factor to initiate branched actin polymerization that drives cell migration. However, it is unknown whether p53 could suppress metastasis through modulating Arp2/3 function. Here, we report that WDR63 is transcriptionally upregulated by p53. We show with migration assays and mouse xenograft models that WDR63 negatively regulates cell migration, invasion, and metastasis downstream of p53. Mechanistically, WDR63 interacts with the Arp2/3 complex and inhibits Arp2/3-mediated actin polymerization. Furthermore, WDR63 overexpression is sufficient to dampen the increase in cell migration, invasion, and metastasis induced by p53 depletion. Together, these findings suggest that WDR63 is an important player in the regulation of Arp2/3 function and also implicate WDR63 as a critical mediator of p53 in suppressing metastasis.

Bclaf1 is a direct target of HIF-1 and critically regulates the stability of HIF-1α under hypoxia.

Hypoxic stress is intimately connected with tumor progression, with hypoxia-inducible factor-1α (HIF-1α) being a critical regulator in this process. HIF-1α is stabilized in response to hypoxia, which is required for the induction of gene transcriptions important for hypoxic adaptation. Bclaf1 is a multifunctional protein involved in tumorigenesis, however, its role in this process is not well characterized. Here we report Bclaf1 is a direct transcriptional target of HIF-1 and upregulated in multiple cell lines during hypoxia. Importantly, we found Bclaf1 is involved in the stabilization of HIF-1α during long-term hypoxic treatments. Compared with the control cells, the protein level and stability of HIF-1α in Bclaf1 knockdown or knockout cells is greatly compromised after long-term hypoxic treatments, concomitant with the impaired inductions of HIF-1 target gene transcription. Bclaf1 knockout HeLa cells exhibit a reduced tumor growth in mice xenografts, in which the expressions of HIF-1α and its target genes are also decreased. Bclaf1 binds to HIF-1α in the nucleus, and this interaction is required for Bclaf1 to stabilize HIF-1α in hypoxic condition. These results uncover a positive feedback loop, HIF-1-Bclaf1, that sustains HIF-1 activity during long-term hypoxic conditions by binding to and protecting HIF-1α from degradation, and suggest that Bclaf1 may promote tumor progression by enhancing HIF-1α stability.

Long noncoding RNA EMS connects c-Myc to cell cycle control and tumorigenesis.

Deregulated expression of c-Myc is an important molecular hallmark of cancer. The oncogenic function of c-Myc has been largely attributed to its intrinsic nature as a master transcription factor. Here, we report the long noncoding RNA (lncRNA) E2F1 messenger RNA (mRNA) stabilizing factor (EMS) as a direct c-Myc transcriptional target. EMS functions as an oncogenic molecule by promoting G1/S cell cycle progression. Mechanistically, EMS cooperates with the RNA binding protein RALY to stabilize E2F1 mRNA, and thereby increases E2F1 expression. Furthermore, EMS is able to connect c-Myc to cell cycle control and tumorigenesis via modulating E2F1 mRNA stability. Together, these findings reveal a previously unappreciated mechanism through which c-Myc induces E2F1 expression and also implicate EMS as an important player in the regulation of c-Myc function.

RNF12 catalyzes BRF1 ubiquitination and regulates RNA polymerase III-dependent transcription.

RNA polymerase III (Pol III) is responsible for the production of small noncoding RNA species, including tRNAs and 5S rRNA. Pol III-dependent transcription is generally enhanced in transformed cells and tumors, but the underlying mechanisms remain not well-understood. It has been demonstrated that the BRF1 subunit of TFIIIB is essential for the accurate initiation of Pol III-dependent transcription. However, it is not known whether BRF1 undergoes ubiquitin modification and whether BRF1 ubiquitination regulates Pol III-dependent transcription. Here, we show that RNF12, a RING domain-containing ubiquitin E3 ligase, physically interacts with BRF1. Via direct interaction, RNF12 catalyzes Lys27- and Lys33-linked polyubiquitination of BRF1. Furthermore, RNF12 is able to negatively regulate Pol III-dependent transcription and cell proliferation via BRF1. These findings uncover a novel mechanism for the regulation of BRF1 and reveal RNF12 as an important regulator of Pol III-dependent transcription.

TRMP, a p53-inducible long noncoding RNA, regulates G1/S cell cycle progression by modulating IRES-dependent p27 translation.

The tumor suppressor p53 plays a pivotal role in the protection against cancer. Increasing evidence suggests that long noncoding RNA (lncRNA) plays an important role in the regulation of the p53 pathway, however, the detailed mechanisms remain to be further elucidated. In this study, we report a new p53-inducible lncRNA that we termed TRMP (TP53-regulated modulator of p27). As a direct transcriptional target of p53, TRMP plays an unexpected pro-survival function. Knockdown of TRMP inhibits cell proliferation by inducing a G1 cell cycle arrest. Mechanistically, TRMP suppresses internal ribosomal entry site (IRES)-dependent translation of p27 by competing p27 mRNA for polypyrimidine tract-binding protein 1 (PTBP1) binding. Furthermore, TRMP is able to regulate cell proliferation, G1/S cell cycle progression, and tumor xenograft growth via the inhibition of p27. Taken together, these findings suggest lncRNA as a new layer to fine-tune the p53 response and reveal TRMP as an important downstream effector of p53 activity.

Regulation of the Mdm2-p53 pathway by the ubiquitin E3 ligase MARCH7.

The tumor suppressor p53 plays a prominent role in the protection against cancer. The activity of p53 is mainly controlled by the ubiquitin E3 ligase Mdm2, which targets p53 for proteasomal degradation. However, the regulation of Mdm2 remains not well understood. Here, we show that MARCH7, a RING domain-containing ubiquitin E3 ligase, physically interacts with Mdm2 and is essential for maintaining the stability of Mdm2. MARCH7 catalyzes Lys63-linked polyubiquitination of Mdm2, which impedes Mdm2 autoubiquitination and degradation, thereby leading to the stabilization of Mdm2. MARCH7 also promotes Mdm2-dependent polyubiquitination and degradation of p53. Furthermore, MARCH7 is able to regulate cell proliferation, DNA damage-induced apoptosis, and tumorigenesis via a p53-dependent mechanism. These findings uncover a novel mechanism for the regulation of Mdm2 and reveal MARCH7 as an important regulator of the Mdm2-p53 pathway.