Publications by authors named "Yongchao Zhao"

66 Publications

P90 RSK confers cancer cell survival by mediating CHK1 degradation in response to glucose stress.

Cancer Sci 2021 Oct 20. Epub 2021 Oct 20.

Department of Hepatobiliary and Pancreatic Surgery, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

In solid tumors, cancer cells have devised multiple approaches to survival and proliferate in response to glucose starvation that is often observed in solid tumor microenvironments. However, the precise mechanisms are far less known. Herein, we report that glucose deprivation activates p90 RSK, a highly conserved Ser/Thr kinase, and activated p90 RSK promotes cancer cell survival. Mechanistically, activated p90 RSK by glucose deprivation phosphorylates CHK1, a key transducer in checkpoint signaling pathways, at Ser280 and triggers CHK1 ubiquitination mediated by SCF ubiquitin ligase and proteasomal degradation, subsequently suppressing cancer cell apoptosis induced by glucose deprivation. Importantly, we identified an inverse correlation between p90 RSK activity and CHK1 levels within the solid tumor mass, with lower levels of CHK1 and higher activity of p90 RSK in the center of the tumor where low glucose concentrations are often observed. Thus, our study indicates that p90 RSK promotes CHK1 phosphorylation at Ser280 and its subsequent degradation, which allows cancer cells to escape from checkpoint signals under the stress of glucose deprivation, leading to cell survival and thus contributing to tumorigenesis.
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http://dx.doi.org/10.1111/cas.15168DOI Listing
October 2021

Neddylation of Coro1a determines the fate of multivesicular bodies and biogenesis of extracellular vesicles.

J Extracell Vesicles 2021 Oct;10(12):e12153

Institute of Immunology, Department of Orthopaedics of the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Multivesicular bodies (MVBs) fuse with not only the plasma membranes to release extracellular vesicles (EVs) but also lysosomes for degradation. Rab7 participates in the lysosomal targeting of MVBs. However, the proteins on MVB that directly bind Rab7, causing MVB recruitment of Rab7 remain unidentified. Here, we show that Coro1a undergoes neddylation modification at K233 by TRIM4. Neddylated Coro1a is associated with the MVB membrane and facilitates MVB recruitment and activation of Rab7 by directly binding Rab7. Subsequently, MVBs are targeted to lysosomes for degradation in a Rab7-dependent manner, leading to reduced EV secretion. Furthermore, a decrease in neddylated Coro1a enhances the production of tumour EVs, thereby promoting tumour progression, indicating that neddylated Coro1a is an ideal target for the regulation of EV biogenesis. Altogether, our data identify a novel substrate of neddylation and reveal an unknown mechanism for MVB recruitment of Rab7, thus providing new insight into the regulation of EV biogenesis.
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http://dx.doi.org/10.1002/jev2.12153DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8500273PMC
October 2021

Trehalose can effectively protect sheep epididymis epithelial cells from oxidative stress.

Arch Anim Breed 2021 18;64(2):335-343. Epub 2021 Aug 18.

College of Animal Science, Inner Mongolia Agricultural University, Hohhot, Inner Mongolia, 010018, China.

Trehalose, a naturally nontoxic disaccharide that does not exist in mammals, stabilizes cell membrane integrity under oxidative stress conditions, the mechanism of which is still unclear. Here, we analyzed the effects of trehalose on sheep epididymis epithelial cell (EEC) proliferation and its possible mechanisms. To study the effect of trehalose on EECs, EECs were isolated from testes of 12-month-old sheep; cell counting kit-8 (CCK-8) was used to measure the growth of the cells. Cell proliferation was evaluated by assaying cell cycle and apoptosis, and RT-PCR was utilized to identify the epididymal molecular markers glutathione peroxidase 5 () and androgen receptor (). Next, reactive oxygen species (ROS) content was evaluated by a dichloro-dihydro-fluorescein diacetate (DCFH-DA) probe. Superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) activities were evaluated by enzyme chemistry methods, and expression was evaluated by qRT-PCR and enzyme-linked immunosorbent assay (ELISA). The results showed that 100  trehalose significantly improved the proliferation potential of EECs, in which the cells could be serially passaged 14 times with continued normal and marker gene expression in vitro. The trehalose can increase significantly a proportion of EECs in S phase ( ) and decrease significantly the apoptotic rate of EECs ( ) compared to the control. Moreover, the trehalose decreased ROS significantly ( ) and increased CAT ( ) and GSH-Px ( ) activities significantly in EECs. mRNA and protein expression were also significantly upregulated in trehalose-treated EECs ( and respectively). Our study suggested that exogenous trehalose exhibited antioxidant activity through increasing the activities of CAT, GSH-Px, and the expression level of and could be employed to maintain vitality of sheep EECs during long-term in vitro culture.
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http://dx.doi.org/10.5194/aab-64-335-2021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8386192PMC
August 2021

USP2 is an SKP2 deubiquitylase that stabilizes both SKP2 and its substrates.

J Biol Chem 2021 Aug 21;297(4):101109. Epub 2021 Aug 21.

Cancer Institute of the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China; Institute of Translational Medicine, Zhejiang University School of Medicine, Hangzhou, China; Cancer Center, Zhejiang University, Hangzhou, China. Electronic address:

The stability of a protein is regulated by a balance between its ubiquitylation and deubiquitylation. S-phase kinase-associated protein 2 (SKP2) is an oncogenic F-box protein that recognizes tumor suppressor substrates for targeted ubiquitylation by the E3 ligase SKP1-Cullin1-F-box and degradation by proteasome. SKP2 is itself ubiquitylated by the E3 ligases APC/C and SCF, and deubiquitylated by deubiquitylases (DUBs) USP10 and USP13. Given the biological significance of SKP2, it is likely that the other E3s or DUBs may also regulate its stability. Here, we report the identification and characterization of USP2 as a new DUB. We first screened a panel of DUBs and found that both USP2 and USP21 bound to endogenous SKP2, but only USP2 deubiquitylated and stabilized SKP2 protein. USP2 inactivation via siRNA knockdown or small-molecule inhibitor treatment remarkably shortened SKP2 protein half-life by enhancing its ubiquitylation and subsequent degradation. Unexpectedly, USP2-stabilized SKP2 did not destabilize its substrates p21 and p27. Mechanistically, USP2 bound to SKP2 via the leucine-rich repeat substrate-binding domain on SKP2 to disrupt the SKP2-substrate binding, leading to stabilization of both SKP2 and these substrates. Biologically, growth suppression induced by USP2 knockdown or USP2 inhibitor is partially mediated via modulation of SKP2 and its substrates. Our study revealed a new mechanism of the cross-talk among the E3-DUB substrates and its potential implication in targeting the USP2-SKP2 axis for cancer therapy.
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http://dx.doi.org/10.1016/j.jbc.2021.101109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8446802PMC
August 2021

DEPTOR inhibits lung tumorigenesis by inactivating the EGFR-mTOR signals.

Cancer Lett 2021 Oct 25;519:263-276. Epub 2021 Jul 25.

Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China; Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China; Institute of Translational Medicine, Zhejiang University School of Medicine, Hangzhou, China; Cancer Center, Zhejiang University, Hangzhou, China. Electronic address:

DEPTOR plays vital roles in the regulation of cell proliferation and survival by directly modulating the activity of mTORC1/2. However, the physiological role of DEPTOR in lung tumorigenesis, as well as its clinical significance, remains elusive. In this study, we revealed that decreased DEPTOR expression correlated with increased tumor size, poor differentiation, and worse survival in patients with lung cancer. DEPTOR depletion promoted cell proliferation, survival, migration, and invasion in human lung cancer cells. Mechanistically, DEPTOR bound to the kinase domain of EGFR via its PDZ domain to inactivate EGFR signal. Thus, DEPTOR depletion not only directly activated mTORC1/2, but also relieved the inhibition of EGFR to subsequently activate mTOR signals, leading to the induction of cell proliferation and survival. Additionally, activated EGFR-mTOR signals upregulated the expression of ZEB1 and SLUG to induce epithelial-mesenchymal transition, resulting in enhanced migration and invasion. Importantly, Deptor deletion accelerated Kras;p53-induced lung tumorigenesis and shortened mouse life span via the activation of EGFR-mTOR signals. Collectively, our study demonstrated that DEPTOR acts as a tumor suppressor in lung tumorigenesis, and its reduction may advance the progression of human lung cancer.
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http://dx.doi.org/10.1016/j.canlet.2021.07.031DOI Listing
October 2021

A modified endocardial radiofrequency ablation approach for hypertrophic obstructive cardiomyopathy guided by transthoracic echocardiography: a case series.

Ann Transl Med 2021 Jun;9(12):1006

Department of Cardiology, Zhongshan Hospital of Fudan University, Shanghai, China.

Background: A series of studies showed that endocardial radiofrequency ablation (ERFA) could reduce the left ventricular outflow tract (LVOT) gradient in patients with septal hypertrophy. This study aimed to determine the safety and efficacy of a modified ERFA approach guided by transthoracic echocardiography (TTE) as an alternative to ablation performed under a three-dimensional (3D) electroanatomical system or intracardiac echocardiography (ICE).

Methods: Twenty-five patients with hypertrophic obstructive cardiomyopathy (HOCM) underwent ERFA of septal hypertrophy, guided by echocardiography. The LVOT gradient, left ventricular ejection fraction (LVEF), LV thickness, New York Heart Association (NYHA) class, and biochemical laboratory values were recorded before ablation and during follow-up.

Results: The patients' peak and stress-induced LVOT gradients were significantly reduced after 12 months of follow-up (resting gradient: from 123.2±17.7 to 15.7±7.8 mmHg, P<0.05; provocative gradient: from 140.2±20.8 to 18.4±8.0 mmHg, P<0.05). Compared with baseline, the septal diameter was reduced slightly after 12 months, but the difference was not significant (24.8±3.5 24.2±3.4 mm, P>0.05). The reduction in LVOT gradient was associated with an improvement in NYHA functional classification (from 3.0±0.0 to 1.6±0.7, P<0.05), the 6-minute walking distance (413±129 m at baseline; 458±108 m immediately after ERFA; 471±139 m after 12 months, P<0.05), and pro B-type natriuretic peptide levels (from 924.00±139 to 137.45 ±75.73 pg/mL, P<0.05). After the procedure, the patients showed no worsening of LVEF compared with baseline (64%±5.3%), and no cases of bundle branch block nor complete heart block occurred.

Conclusions: ERFA guided by TTE provides a new treatment option for HOCM which can achieve symptomatic improvement as well as a significant and sustained reduction of the LVOT gradient. Moreover, by avoiding the use of the 3D electroanatomical system or ICE, this treatment has an acceptable cost.
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http://dx.doi.org/10.21037/atm-21-2783DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8267310PMC
June 2021

Mononuclear phagocyte system blockade using extracellular vesicles modified with CD47 on membrane surface for myocardial infarction reperfusion injury treatment.

Biomaterials 2021 08 28;275:121000. Epub 2021 Jun 28.

Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University, Shanghai, China; Institute of Biomedical Science, Fudan University, Shanghai, China. Electronic address:

Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) with anti-apoptotic and anti-inflammatory properties have been intensively studied. However, rapid clearance by the mononuclear phagocyte system remains a huge barrier for the delivery of extracellular vesicle contents into target organs and restricts its wider application, particularly in the heart. CD47 is a transmembrane protein that enables cancer cells to evade clearance by macrophages through CD47 signal regulatory proteinα binding, which initiates a "don't eat me" signal. This study aimed to explore the biodistribution and delivery efficiency of EVs carrying the membrane protein CD47 and specific anti-apoptotic miRNAs. EVs were isolated from MSCs overexpressing CD47 (CD47-EVs) and identified. Fluorescence-labeled EVs were injected through the tail vein and tracked using fluorescence imaging. In silico analysis was performed to determine miRNA profiles in MSCs and in a heart-derived H9c2 cardiomyoblast cell line under hypoxia vs. normoxia conditions. Electro CD47-EV was constructed by encapsulating purified CD47-EV with miR-21a via electroporation. The effect of miR21-EVs on the pro-apoptotic gene encoding phosphatase and tensin homolog (PTEN) was evaluated by dual-luciferase assay, qPCR, and western blotting. Exogenous miR21 distribution, PTEN protein level, blood vessel density, anti-apoptotic effect by TdT-mediated dUTP nick-end labeling staining, and macrophage and leukocyte infiltration in the myocardium were assessed by immunofluorescence staining. Cardiac functional recovery during the early stage and recovery period was evaluated using echocardiography. The results showed that CD47-EVs were still detectable in the plasma 120 min after the tail vein injection, compared to the detection time of less than 30 min observed with the unmodified EVs. More strikingly, CD47-EVs preferentially accumulated in the heart in the ischemia-reperfusion (I/R) + CD47-EV group [heart total fluorescence radiance ( × 10 Photons/sec/cm/sr) 51.62 ± 11.30 v.s. 10.08 ± 3.15 in the I/R + unmodified EVs group] 8 h post-injection. Exogenous miR-21 is efficiently internalized into cardiomyocytes, inhibits apoptosis, alleviates inflammation, and improves cardiac function. In conclusion, electro CD47-EVs efficiently improve biodistribution in the heart, shedding new light on the application of a two-step EV delivery method (CD47 genetic modification followed by therapeutic content electrotransfection) as a potential therapeutic tool for myocardial I/R injury that may benefit patients in the future.
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http://dx.doi.org/10.1016/j.biomaterials.2021.121000DOI Listing
August 2021

m7G Methyltransferase METTL1 Promotes Post-ischemic Angiogenesis via Promoting VEGFA mRNA Translation.

Front Cell Dev Biol 2021 31;9:642080. Epub 2021 May 31.

Department of Cardiology, Affiliated Hospital of Zunyi Medical University, Zunyi, China.

Post-transcriptional modifications play pivotal roles in various pathological processes and ischemic disorders. However, the role of N7-methylguanosine (m7G), particularly m7G in mRNA, on post-ischemic angiogenesis remains largely unknown. Here, we identified that methyltransferase like 1 (METTL1) was a critical candidate responsible for a global decrease of m7G within mRNA from the ischemic tissues. The gene transfer of METTL1 improved blood flow recovery and increased angiogenesis with enhanced mRNA m7G upon post-ischemic injury. Increased METTL1 expression using plasmid transfection promoted HUVECs proliferation, migration, and tube formation with a global increase of m7G in mRNA. Mechanistically, METTL1 promoted VEGFA mRNA translation in an m7G methylation-dependent manner. Our findings emphasize a critical link between mRNA m7G and ischemia and provide a novel insight of targeting METTL1 in the therapeutic angiogenesis for ischemic disorders, including peripheral arterial disease.
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http://dx.doi.org/10.3389/fcell.2021.642080DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8200671PMC
May 2021

Loss of m6A demethylase ALKBH5 promotes post-ischemic angiogenesis via post-transcriptional stabilization of WNT5A.

Clin Transl Med 2021 05;11(5):e402

Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai Institute of Cardiovascular Diseases, Shanghai, China.

Background: Post-ischemic angiogenesis is critical for blood flow recovery and ischemic tissue repair. N6-methyladenosine (m6A) plays essential roles in numerous biological processes. However, the impact and connected mechanism of m6A on post-ischemic angiogenesis are not fully understood.

Methods: AlkB homolog 5 (ALKBH5) was screened out among several methyltransferases and demethylases involved in dynamic m6A regulation. Cardiac microvascular endothelial cells (CMECs) angiogenesis and WNT family member 5A (WNT5A) stability were analyzed upon ALKBH5 overexpression with adenovirus or knockdown with small interfering RNAs in vitro. The blood flow recovery, capillary, and small artery densities were evaluated in adeno-associated virus (AAV)-ALKBH5 overexpression or ALKBH5 knockout (KO) mice in a hind-limb ischemia model. The same experiments were conducted to explore the translational value of transient silencing of ALKBH5 with adenovirus.

Results: ALKBH5 was significantly upregulated in hypoxic CMECs and led to a global decrease of m6A level. ALKBH5 overexpression further reduced m6A level in normoxic and hypoxic CMECs, impaired proliferation, migration, and tube formation only in hypoxic CMECs. Conversely, ALKBH5 knockdown preserved m6A levels and promoted angiogenic phenotypes in hypoxic but not in normoxic CMECs. Mechanistically, ALKBH5 regulated WNT5A expression through post-transcriptional mRNA modulation in an m6A-dependent manner, which decreased its stability and subsequently impeded angiogenesis in hypoxic CMECs. Furthermore, ALKBH5 overexpression hindered blood flow recovery and reduced CD31 and alpha-smooth muscle actin expression in hind-limb ischemia mice. As expected, ALKBH5-KO mice exhibited improved blood flow recovery, increased capillary, and small artery densities after hind-limb ischemia, and similar beneficial effects were observed in mice with transient adenoviral ALKBH5 gene silencing.

Conclusion: We demonstrate that ALKBH5 is a negative regulator of post-ischemic angiogenesis via post-transcriptional modulation and destabilization of WNT5A mRNA in an m6A-dependent manner. Targeting ALKBH5 may be a potential therapeutic option for ischemic diseases, including peripheral artery disease.
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http://dx.doi.org/10.1002/ctm2.402DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8087997PMC
May 2021

DEPTOR stabilizes ErbB2 to promote the proliferation and survival of ErbB2-positive breast cancer cells.

Theranostics 2021 19;11(13):6355-6369. Epub 2021 Apr 19.

Department of Hepatobiliary and Pancreatic Surgery, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Dysregulation of the PI3K/AKT/mTOR pathway occurs frequently in cancers, providing an attractive therapeutic target for anticancer treatments. DEPTOR plays essential roles in regulation of cell proliferation and survival by directly modulating mTOR activity. However, whether DEPTOR regulates the growth of ErbB2-positive breast cancer cells remains unknown. DEPTOR expression was determined by TCGA data analysis and immunohistochemistry of human breast tissue microarrays. The membrane localization of DEPTOR was demonstrated by immunofluorescence and subcellular fractionation. The interaction of DEPTOR with ErbB2 was determined by immunoprecipitation. Furthermore, the biological significance of this interaction was assessed by ATPlite cell growth, clonogenic survival, and flow cytometry-based apoptosis assays. DEPTOR promoted the proliferation and survival of ErbB2-positive breast cancer cells by directly interacting with and stabilizing ErbB2. Specifically, DEPTOR translocates to cell membrane and interacts with ErbB2 to disrupt ErbB2 polyubiquitination and degradation promoted by β-TrCP, an E3 ubiquitin ligase. DEPTOR knockdown destabilizes ErbB2 by shortening its protein half-life to inactivate ErbB2-PI3K-AKT-mTOR signaling, leading to the suppression of cell proliferation and survival by inducing apoptosis. Ectopic expression of a constitutively active ErbB2 mutant completely rescued the reduction in cell proliferation and survival by DEPTOR knockdown. Importantly, DEPTOR expression is increased in human breast cancer tissues and its overexpression correlates with poor patient survival. Moreover, DEPTOR is located on the cell membrane in ErbB2-positive breast cancer tissues, but not in tumor-adjacent normal tissues, indicating that DEPTOR may contribute to the oncogenic characteristics of ErbB2. Our study reveals a novel mechanism by which DEPTOR promotes breast cancer cell proliferation and survival by stabilizing ErbB2.
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http://dx.doi.org/10.7150/thno.51286DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8120212PMC
July 2021

Nuclear ErbB2 represses DEPTOR transcription to inhibit autophagy in breast cancer cells.

Cell Death Dis 2021 04 14;12(4):397. Epub 2021 Apr 14.

Department of Hepatobiliary and Pancreatic Surgery, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

ErbB2, a classical receptor tyrosine kinase, is frequently overexpressed in breast cancer cells. Although the role of ErbB2 in the transmission of extracellular signals to intracellular matrix has been widely studied, the functions of nuclear ErbB2 remain largely elusive. Here, we report a novel function of nuclear ErbB2 in repressing the transcription of DEPTOR, a direct inhibitor of mTOR. Nuclear ErbB2 directly binds to the consensus binding sequence in the DEPTOR promoter to repress its transcription. The kinase activity of ErbB2 is required for its nuclear translocation and transcriptional repression of DEPTOR. Moreover, the repressed DEPTOR by nuclear ErbB2 inhibits the induction of autophagy by activating mTORC1. Thus, our study reveals a novel mechanism for autophagy regulation by functional ErbB2, which translocates to the nucleus and acts as a transcriptional regulator to suppress DEPTOR transcription, leading to activation of the PI3K/AKT/mTOR pathway to inhibit autophagy.
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http://dx.doi.org/10.1038/s41419-021-03686-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8047043PMC
April 2021

Shexiang Baoxin Pill Attenuates Ischemic Injury by Promoting Angiogenesis by Activation of Aldehyde Dehydrogenase 2.

J Cardiovasc Pharmacol 2021 03;77(3):408-417

Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University, Shanghai, China.

Abstract: Promoting angiogenesis is a critical treatment strategy for ischemic cardiovascular diseases. Shexiang Baoxin Pill (SBP), a traditional Chinese medicine, has been reported to be capable of relieving angina and improve heart function by promoting angiogenesis. The aim of this study was to determine the role of mitochondrial aldehyde dehydrogenase 2 (ALDH2) in SBP-induced angiogenesis. Left femoral artery ligation was performed in wild-type mice (WT) and ALDH2 knockout mice, which were administrated with SBP (20 mg/kg/d) or equal volume saline per day by gastric gavage for 2 weeks. Perfusion recovery, angiogenesis in chronic hind limb ischemia, was significantly improved in the WT + SBP group than in the WT group. However, these beneficial effects were absent in ALDH2 knockout mice. In vitro, hypoxia impaired the ability of proliferation, migration and tube formation, sprouting angiogenesis, and promoted apoptosis in cardiovascular microvascular endothelial cells, whereas the hypoxia damage was restored by SBP. The protective effect of SBP was remarkably weakened by ALDH2 knockdown. Furthermore, SBP suppressed hypoxia-induced ALDH2/protein kinase B (AKT)/mammalian target of rapamycin pathways. In conclusion, this study demonstrated that SBP protected lower limb from ischemia injury through the ALDH2-dependent pathway. The protective mechanism of SBP in cardiovascular microvascular endothelial cells was partly mediated through ALDH2/AKT/mammalian target of rapamycin pathways.
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http://dx.doi.org/10.1097/FJC.0000000000000967DOI Listing
March 2021

Qiliqiangxin alleviates Ang II-induced CMECs apoptosis by downregulating autophagy via the ErbB2-AKT-FoxO3a axis.

Life Sci 2021 May 27;273:119239. Epub 2021 Feb 27.

Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai Institute of Cardiovascular Diseases, Shanghai 200032, China; Institute of Biomedical Sciences, Fudan University, Shanghai 200032, China. Electronic address:

Our previous work revealed the protective effect of Qiliqiangxin (QLQX) on cardiac microvascular endothelial cells (CMECs), but the underlying mechanisms remain unclear. We aimed to investigate whether QLQX exerts its protective effect against high-concentration angiotensin II (Ang II)-induced CMEC apoptosis through the autophagy machinery. CMECs were cultured in high-concentration Ang II (1 μM) medium in the presence or absence of QLQX for 48 h. We found that QLQX obviously inhibited Ang II-triggered autophagosome synthesis and apoptosis in cultured CMECs. QLQX-mediated protection against Ang II-induced CMEC apoptosis was reversed by the autophagy activator rapamycin. Specifically, deletion of ATG7 in cultured CMECs indicated a detrimental role of autophagy in Ang II-induced CMEC apoptosis. QLQX reversed Ang II-mediated ErbB2 phosphorylation impairment. Furthermore, inhibition of ErbB2 phosphorylation with lapatinib in CMECs revealed that QLQX-induced downregulation of Ang II-activated autophagy and apoptosis was ErbB2 phosphorylation-dependent via the AKT-FoxO3a axis. Activation of ErbB2 phosphorylation by Neuregulin-1β achieved a similar CMEC-protective effect as QLQX in high-concentration Ang II medium, and this effect was also abolished by autophagy activation. These results show that the CMEC-protective effect of QLQX under high-concentration Ang II conditions could be partly attributable to QLQX-mediated ErbB2 phosphorylation-dependent downregulation of autophagy via the AKT-FoxO3a axis.
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http://dx.doi.org/10.1016/j.lfs.2021.119239DOI Listing
May 2021

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition).

Autophagy 2021 Jan 8;17(1):1-382. Epub 2021 Feb 8.

University of Crete, School of Medicine, Laboratory of Clinical Microbiology and Microbial Pathogenesis, Voutes, Heraklion, Crete, Greece; Foundation for Research and Technology, Institute of Molecular Biology and Biotechnology (IMBB), Heraklion, Crete, Greece.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
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http://dx.doi.org/10.1080/15548627.2020.1797280DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7996087PMC
January 2021

Alda-1 treatment promotes the therapeutic effect of mitochondrial transplantation for myocardial ischemia-reperfusion injury.

Bioact Mater 2021 Jul 8;6(7):2058-2069. Epub 2021 Jan 8.

Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University, Shanghai, 200032, China.

Mitochondrial damage is a critical driver in myocardial ischemia-reperfusion (I/R) injury and can be alleviated via the mitochondrial transplantation. The efficiency of mitochondrial transplantation is determined by mitochondrial vitality. Because aldehyde dehydrogenase 2 (ALDH2) has a key role in regulating mitochondrial homeostasis, we aimed to investigate its potential therapeutic effects on mitochondrial transplantation via the use of ALDH2 activator, Alda-1. Our present study demonstrated that time-dependent internalization of exogenous mitochondria by cardiomyocytes along with ATP production were significantly increased in response to mitochondrial transplantation. Furthermore, Alda-1 treatment remarkably promoted the oxygen consumption rate and baseline mechanical function of cardiomyocytes caused by mitochondrial transplantation. Mitochondrial transplantation inhibited cardiomyocyte apoptosis induced by the hypoxia-reoxygenation exposure, independent of Alda-1 treatment. However, promotion of the mechanical function of cardiomyocytes exposed to hypoxia-reoxygenation treatment was only observed after mitochondrial Alda-1 treatment and transplantation. By using a myocardial I/R mouse model, our results revealed that transplantation of Alda-1-treated mitochondria into mouse myocardial tissues limited the infarction size after I/R injury, which was at least in part due to increased mitochondrial potential-mediated fusion. In conclusion, ALDH2 activation in mitochondrial transplantation shows great potential for the treatment of myocardial I/R injury.
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http://dx.doi.org/10.1016/j.bioactmat.2020.12.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7809100PMC
July 2021

Analysis of changes in circular RNA expression and construction of ceRNA networks in human dilated cardiomyopathy.

J Cell Mol Med 2021 03 22;25(5):2572-2583. Epub 2021 Jan 22.

Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai, China.

Dilated cardiomyopathy (DCM) is a severe life-threatening disease worldwide, and the underlying mechanisms remain unclear. Circular RNAs (circRNAs) have been reported to play important roles in various cardiovascular diseases and can function as competitive endogenous RNAs (ceRNAs). However, their role in human DCM has not been fully elucidated. In the present study, heart samples from DCM patients and healthy controls were used to identify circRNAs by RNA sequencing. Real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was conducted to validate differentially expressed circRNAs and mRNAs. A total of 9585 circRNAs and 22050 mRNAs were detected in the two groups. Overall, 213 circRNAs and 617 mRNAs were significantly up-regulated in the DCM group compared with the control group. Similarly, 85 circRNAs and 1125 mRNAs were significantly down-regulated. According to the ceRNA theory, circRNAs can indirectly interact with mRNAs by directly binding to microRNAs (miRNAs), and circRNAs and mRNAs should be concurrently either up-regulated or down-regulated. Based on this theory, we constructed two circRNA-miRNA-mRNA networks by using the RNA sequencing data and prediction by proprietary software. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to probe the potential functions of differentially expressed circRNAs. In conclusion, this study revealed that the expression of cardiac circRNAs was altered in human DCM and explored the potential functions of circRNAs by constructing ceRNA networks. These findings provide a foundation for future studies of circRNAs in DCM.
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http://dx.doi.org/10.1111/jcmm.16251DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7933965PMC
March 2021

Effect of mutations in capsid shell protein on the assembly of BmCPV virus-like particles.

J Gen Virol 2021 03 17;102(3). Epub 2020 Dec 17.

Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science, South China Agricultural University, Guangzhou 510642, PR China.

cytoplasmic polyhedrosis virus (BmCPV) is a typical single-layer capsid dsRNA virus belonging to the genus in the family . The results of cryo-electron microscopy showed that the BmCPV capsid consists of 60 asymmetric units, and each asymmetric unit contains one turret protein (TP), two large protrusion proteins (LPP) and two capsid shell proteins (CSP). CSP has the ability to self-assemble into virus-like particles (VLPs), and the small protrusion domain (SPD) in CSP may play an essential role in the assembly of viral capsids. In this study, three critical amino acid sites, D828, S829 and V945, in the SPD were efficiently mutated (point mutation) based on the principle of PCR circular mutagenesis. Moreover, a multi-gene expression system, Ac-MultiBac baculovirus, was used to produce eight different recombinant VLPs . Transmission electron microscopy showed that the single site and double site mutations had little effect on the efficiency and morphology of the assembly of VLPs. Still, the simultaneous mutation of the three sites had a significant impact. The experimental results demonstrate that the SPD of CSP plays an essential role in assembly of the viral capsid, which lays the foundation for further analysis of the molecular and structural mechanism of BmCPV capsid assembly.
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http://dx.doi.org/10.1099/jgv.0.001542DOI Listing
March 2021

DEPTOR is a direct p53 target that suppresses cell growth and chemosensitivity.

Cell Death Dis 2020 11 12;11(11):976. Epub 2020 Nov 12.

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

DEP-domain containing mTOR-interacting protein (DEPTOR), a natural mTOR inhibitor, has essential roles in several processes, including cell growth, metabolism, apoptosis, and immunity. DEPTOR expression has been shown to be diversely controlled at transcriptional levels in cell- and context-specific manners. However, whether there is a general mechanism for the regulation of DEPTOR expression remains largely unknown. Here, we report that DEPTOR is a downstream target of the tumor suppressor, p53, whose activity is positively correlated with DEPTOR expression both in vitro in cell cultures and in vivo in mouse tissues. Mechanistically, p53 directly binds to the DEPTOR promoter and transactivates its expression. Depletion of the p53-binding site on the DEPTOR promoter by CRISPR-Cas9 technology decreases DEPTOR expression and promotes cell proliferation and survival by activating AKT signaling. Importantly, inhibition of AKT by small molecular inhibitors or genetic knockdown abrogates the induction of cell growth and survival induced by deletion of the p53-binding region on the DEPTOR promoter. Furthermore, p53, upon activation by the genotoxic agent doxorubicin, induces DEPTOR expression, leading to cancer cell resistance to doxorubicin. Together, DEPTOR is a direct p53 downstream target and contributes to p53-mediated inhibition of cell proliferation, survival, and chemosensitivity.
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http://dx.doi.org/10.1038/s41419-020-03185-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7661726PMC
November 2020

EAG1 enhances hepatocellular carcinoma proliferation by modulating SKP2 and metastasis through pseudopod formation.

Oncogene 2021 01 24;40(1):163-176. Epub 2020 Oct 24.

Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Ether-à-go-go-1 (EAG1), one of the potassium channels, is involved in various physiological processes and plays an important role in the tumorigenesis of many kinds of cancer. EAG1 is highly expressed in hepatocarcinoma cells and is closely related to clinical prognosis, but the molecular mechanism remains elusive. In this study, we verified that EAG1 promotes the proliferation of hepatocellular carcinoma (HCC) both in vitro and in vivo. It promotes cell cycle progression by inhibiting the ubiquitination of SKP2. In addition, EAG1 promotes the migration and invasion of HCC by promoting cell pseudopod formation. Furthermore, in a high-pressure plasmid-injected mouse liver orthotopic carcinoma model, astemizole, an EAG family blocker, can significantly inhibit the formation of liver cancer. Meanwhile, liver-specific EAG1 knockout mice show resistance to hepatocarcinogenesis. This research demonstrated that EAG1 plays an important role in the progression of HCC, and could be a potential therapeutic target for HCC.
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http://dx.doi.org/10.1038/s41388-020-01522-6DOI Listing
January 2021

The characteristics and roles of β-TrCP1/2 in carcinogenesis.

FEBS J 2021 06 23;288(11):3351-3374. Epub 2020 Oct 23.

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

β-transducin repeat-containing protein (β-TrCP), one of the well-characterized F-box proteins, acts as a substrate receptor and constitutes an active SCF E3 ligase with a scaffold protein CUL1, a RING protein RBX1, and an adaptor protein SKP1. β-TrCP plays a critical role in the regulation of various physiological and pathological processes, including signal transduction, cell cycle progression, cell migration, DNA damage response, and tumorigenesis, by governing burgeoning amounts of key regulators for ubiquitination and proteasomal degradation. Given that a variety of β-TrCP substrates are well-known oncoproteins and tumor suppressors, and dysregulation of β-TrCP is frequently identified in human cancers, β-TrCP plays a vital role in carcinogenesis. In this review, we first briefly introduce the characteristics of β-TrCP1, β-TrCP2, and SCF ubiquitin ligase, and then discuss SCF ubiquitin ligase regulated biological processes by targeting its substrates for degradation. Moreover, we summarize the regulation of β-TrCP1 and β-TrCP2 at multiple layers and further discuss the various roles of β-TrCP1 and β-TrCP2 in human cancer, functioning as either an oncoprotein or a tumor suppressor in a manner dependent of cellular context. Finally, we provide novel insights for future perspectives on the potential of targeting β-TrCP1 and β-TrCP2 for cancer therapy.
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http://dx.doi.org/10.1111/febs.15585DOI Listing
June 2021

Emerging role of FBXO22 in carcinogenesis.

Cell Death Discov 2020 27;6:66. Epub 2020 Jul 27.

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

The F-box protein 22 (FBXO22), one of F-box proteins, has been identified to be critically involved in carcinogenesis. FBXO22 promotes proliferation in breast cancer and lung cancer, but suppresses migration and metastasis. FBXO22 exerts oncogenetic functions via promoting the ubiquitination and degradation of its substrates, including KDM4A, KDM4B, methylated p53, p21, KLF4, LKB1, Snail, CD147, Bach1, PTEN, and HDM2. FBXO22 is also regulated by several regulatory factors such as p53, miR-155, SNHG14, and circ_0006282. In this review, we summarize the regulatory factors and downstream targets of FBXO22 in cancers, discuss its functions in tumorigenesis, and further highlight the alteration of FBXO22 expression in a variety of human malignancies. Finally, we provide novel insights for future perspectives on targeting FBXO22 as a promising strategy for cancer therapy.
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http://dx.doi.org/10.1038/s41420-020-00303-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7385156PMC
July 2020

Neddylation modification of ribosomal protein RPS27L or RPS27 by MDM2 or NEDP1 regulates cancer cell survival.

FASEB J 2020 10 11;34(10):13419-13429. Epub 2020 Aug 11.

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, P. R. China.

Neddylation plays a distinct role in stabilization of a subset of ribosomal proteins. Whether the family of ribosomal proteins S27 (RPS27 and RPS27-like) is subjected to neddylation regulation with associated biological consequence is totally unknown. Here, we report that both family members are subjected to neddylation by MDM2 E3 ubiquitin ligase, and deneddylation by NEDP1. Blockage of neddylation with MLN4924, a small molecule inhibitor of neddylation-activating enzyme, destabilizes RPS27L and RPS27 by shortening their protein half-lives. Biologically, knockdown of RPS27L and RPS27 sensitizes, whereas ectopic expression of RPS27L and RPS27 desensitizes cancer cells to MLN4924-induced apoptosis. Taken together, our study demonstrates that neddylation stabilizes RPS27L and RPS27 to confer the survival of cancer cells.
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http://dx.doi.org/10.1096/fj.202000530RRRDOI Listing
October 2020

Cullin-RING Ligase 5: Functional characterization and its role in human cancers.

Semin Cancer Biol 2020 12 22;67(Pt 2):61-79. Epub 2020 Apr 22.

Institute of Translational Medicine, Zhejiang University School of Medicine, Hangzhou, China; Cancer Institute of the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China; Division of Radiation and Cancer Biology, Department of Radiation Oncology, University of Michigan, Ann Arbor, MI, USA. Electronic address:

Cullin-RING ligase 5 (CRL5) is a multi-protein complex and consists of a scaffold protien cullin 5, a RING protein RBX2 (also known as ROC2 or SAG), adaptor proteins Elongin B/C, and a substrate receptor protein SOCS. Through targeting a variety of substrates for proteasomal degradation or modulating various protein-protein interactions, CRL5 is involved in regulation of many biological processes, such as cytokine signal transduction, inflammation, viral infection, and oncogenesis. As many substrates of CRL5 are well-known oncoproteins or tumor suppressors, abnormal regulation of CRL5 is commonly found in human cancers. In this review, we first briefly introduce each of CRL5 components, and then discuss the biological processes regulated by four members of SOCS-box-containing substrate receptor family through substrate degradation. We next describe how CRL5 is hijacked by a variety of viral proteins to degrade host anti-viral proteins, which facilitates virus infection. We further discuss the regulation of CUL5 and its various roles in human cancers, acting as either a tumor suppressor or an oncoprotein in a context-dependent manner. Finally, we propose novel insights for future perspectives on the validation of cullin5 and other CRL5 components as potential targets, and possible targeting strategies to discover CRL5 inhibitors for anti-cancer and anti-virus therapies.
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http://dx.doi.org/10.1016/j.semcancer.2020.04.003DOI Listing
December 2020

SCF-mediated degradation of TOP2β promotes cancer cell survival in response to chemotherapeutic drugs targeting topoisomerase II.

Oncogenesis 2020 Feb 3;9(2). Epub 2020 Feb 3.

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Topoisomerase II (TOP2)-targeting anticancer chemotherapeutic drugs, termed TOP2 poisons, are widely used and effective in the clinic by stabilizing TOP2-DNA covalent complexes to induce DNA double-strand breaks (DSBs) and ultimately, cause cell death. The stabilized TOP2-DNA complex is known to be degraded by proteasome, whereas the underlying mechanism for instant TOP2β degradation in response to TOP2 poisons and the subsequent biological consequence remain elusive. Here, we reported that TOP2 poison-induced TOP2β degradation is mediated by SCF ubiquitin ligase. Specifically, DNA damage signal, triggered by teniposide (VM-26) treatment, activates ATM, cooperating with CK1 to phosphorylate TOP2β on Ser1134 and Ser1130, respectively, in a canonical degron motif to facilitate β-TrCP binding and subsequent degradation. Inactivation of ATM, CK1 or SCF by small molecular inhibitors or genetic knockdown/knockout abrogates TOP2β degradation. Biologically, blockage of TOP2β degradation in combination with VM-26 treatment impairs DNA damage response and repair, leading to an accelerated cell death via apoptosis. Thus, it appears that TOP2β degradation is a cellular defensive mechanism to facilitate the exposure of DSBs to trigger DNA damage response and repair. Collectively, our findings reveal a new strategy to improve the efficacy of TOP2 poisons in combination with small-molecule inhibitors against TOP2β degradation.
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http://dx.doi.org/10.1038/s41389-020-0196-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6997367PMC
February 2020

FBXW7 Confers Radiation Survival by Targeting p53 for Degradation.

Cell Rep 2020 01;30(2):497-509.e4

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China; Institute of Translational Medicine, Zhejiang University School of Medicine, Hangzhou, China. Electronic address:

The tumor suppressor p53 plays a critical role in integrating a wide variety of stress responses. Therefore, p53 levels are precisely regulated by multiple ubiquitin ligases. In this study, we report that FBXW7, a substrate recognition component of the SKP1-CUL1-F-box (SCF) E3 ligase, interacts with and targets p53 for polyubiquitination and proteasomal degradation after exposure to ionizing radiation or etoposide. Mechanistically, DNA damage activates ATM to phosphorylate p53 on Ser33 and Ser37, which facilitates the FBXW7 binding and subsequent p53 degradation by SCF. Inactivation of ATM or SCF by small molecular inhibitors or genetic knockdown/knockout approaches extends the p53 protein half-life upon DNA damage in an MDM2-independent manner. Biologically, FBXW7 inactivation sensitizes cancer cells to radiation or etoposide by stabilizing p53 to induce cell-cycle arrest and apoptosis. Taken together, our study elucidates a mechanism by which FBXW7 confers cancer cell survival during radiotherapy or chemotherapy via p53 targeting.
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http://dx.doi.org/10.1016/j.celrep.2019.12.032DOI Listing
January 2020

Targeting Cullin-RING Ubiquitin Ligases and the Applications in PROTACs.

Adv Exp Med Biol 2020 ;1217:317-347

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Cullin-RING ligases (CRLs), the largest family of E3 ubiquitin ligases, have become an attractive target for drug discovery, primarily due to their ability to regulate the degradation of numerous functionally and structurally diverse proteins, thereby controlling a myriad of biological processes. As the abnormal expressions of CRLs and their substrate proteins are associated with human diseases, elucidating their roles in these physiological and pathological processes will facilitate CRL-targeting drug development for the treatment of these diseases. Notably, these studies are also providing new concepts for the design of potential small-molecule therapeutics targeting CRLs and for the use of CRLs to degrade "undruggable" proteins. In this chapter, we systematically review the development of small molecules that target CRLs and especially emphasize the applications of CRLs in a chemical chimera for protein degradation, termed proteolysis-targeting chimeras (PROTACs).
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http://dx.doi.org/10.1007/978-981-15-1025-0_19DOI Listing
February 2020

DEPTOR is an in vivo tumor suppressor that inhibits prostate tumorigenesis via the inactivation of mTORC1/2 signals.

Oncogene 2020 02 4;39(7):1557-1571. Epub 2019 Nov 4.

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

The DEPTOR-mTORC1/2 axis has been shown to play an important, but a context dependent role in the regulation of proliferation and the survival of various cancer cells in cell culture settings. The in vivo role of DEPTOR in tumorigenesis remains elusive. Here we showed that the levels of both DEPTOR protein and mRNA were substantially decreased in human prostate cancer tissues, which positively correlated with disease progression. DEPTOR depletion accelerated proliferation and survival, migration, and invasion in human prostate cancer cells. Mechanistically, DEPTOR depletion not only activated both mTORC1 and mTORC2 signals to promote cell proliferation and survival, but also induced an AKT-dependent epithelial-mesenchymal transition (EMT) and β-catenin nuclear translocation to promote cell migration and invasion. Abrogation of mTOR or AKT activation rescued the biological consequences of DEPTOR depletion. Importantly, in a Deptor-KO mouse model, Deptor knockout accelerated prostate tumorigenesis triggered by Pten loss via the activation of mTOR signaling. Collectively, our study demonstrates that DEPTOR is a tumor suppressor in the prostate, and its depletion promotes tumorigenesis via the activation of mTORC1 and mTORC2 signals. Thus, DEPTOR reactivation via a variety of means would have therapeutic potential for the treatment of prostate cancer.
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http://dx.doi.org/10.1038/s41388-019-1085-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7018663PMC
February 2020

Experimental study of the kinetics of the adsorption of nitrogen by coal containing different amounts of water.

An Acad Bras Cienc 2019 Sep 2;91(3):e20180434. Epub 2019 Sep 2.

School of Energy Science and Engineering, Henan Polytechnic University, Jiaozuo 454000, China.

Useful insight can be gained into the underlying adsorption mechanism by studying the adsorption of nitrogen by primary structure coal with different water content. In this work, a coal-adsorption gas experimental system is used to investigate the adsorption of nitrogen by coal samples containing different amounts of water ( 2.894%, 1.871%, 0.897%, and 0%) and different pressures (0.7, 1.2, 1.7, 2.2, 2.7, 3.2, and 3.7MPa). The adsorption rates under the different conditions were calculated using a volume method, and the adsorption kinetics investigated using a kinetic model. The results show that an adsorption model based on an opposing process characterizes the adsorption behavior better than the pseudo-first order kinetic model and pseudo-second order kinetic model. The adsorption rate constant, k, reflects the rate at which the gas can get into the pores of different sizes in the coal a greater k value, implies a greater increment in the rate. The k value is found to decrease when the initial pressure is increased for the same moisture content. Also, the greater the water content, the smaller the value of k for a given initial pressure. As the moisture content continues to increase, the k value tends to a certain value. The results presented can provide an experimental basis for the study of the mechanism responsible for the adsorption of nitrogen by primary structure coal with different moisture content.
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http://dx.doi.org/10.1590/0001-3765201920180434DOI Listing
September 2019

The cross talk of two family members of β-TrCP in the regulation of cell autophagy and growth.

Cell Death Differ 2020 03 13;27(3):1119-1133. Epub 2019 Aug 13.

Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

β-transducin repeat-containing protein (β-TrCP), one of the best-characterized substrate recognition components of the SKP1-CUL1-F-box (SCF) E3 ligase, has two distinct paralogs, β-TrCP1 and β-TrCP2, expressed in mammals. Through governing the ubiquitination and degradation of numerous key regulators, β-TrCP1/2 regulates various cellular physiological and pathological processes. However, whether and how these two proteins cross talk and whether they regulate cell autophagy and proliferation in different manners is completely unknown. Herein, we report that β-TrCP1 and β-TrCP2 are the physiological substrates of SCF E3 ligase and target each other for degradation that is dependent on their β-TrCP degron sequences. Furthermore, glucose deprivation activates AMPK kinase to phosphorylate β-TrCP1 and promotes the subsequent ubiquitination and degradation of β-TrCP1 by β-TrCP2, but does not promote β-TrCP2 degradation by β-TrCP1. Finally, we found that β-TrCP2, not β-TrCP1, preferentially degrades DEPTOR and REDD1, the inhibitors of mTORC1, to activate mTORC1, leading to autophagy inhibition and cell growth. Thus, our study demonstrates that β-TrCP1 and β-TrCP2 mutually target each other for degradation and that β-TrCP2 acts as a dominant paralog in the regulation of cell autophagy and growth, which might be a promising anticancer target.
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http://dx.doi.org/10.1038/s41418-019-0402-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7206145PMC
March 2020

Transcriptional response of immune-related genes after endogenous expression of VP1 and exogenous exposure to VP1-based VLPs and CPV virions in lepidopteran cell lines.

Mol Genet Genomics 2019 Aug 28;294(4):887-899. Epub 2019 Mar 28.

College of Animal Science, South China Agricultural University, Guangzhou, 510642, Guangdong, People's Republic of China.

In insects, RNAi is considered the major antiviral immune defense pathway. DsRNAs produced during viral infection are processed by Dicer enzymes into small RNAs that function as specificity determinants to silence viral genes. By contrast, in mammals, recognition of molecules associated with viral infection, such as dsRNA, by pattern recognition receptors (PRRs) initiates a signaling cascade that culminates in the production and release of signaling proteins with antiviral function such as interferons. However, in insects, the hypothesis that components of virions can be recognized as pathogen-activated molecular patterns (PAMPs) to activate the innate immune response has not been investigated systematically. In this study, the potential of VP1, that constitutes the major capsid protein of cytoplasmic polyhedrosis virus (CPV; Reoviridae), to activate a collection of immune-related genes was examined in silkworm-derived Bm5 cells. Two different methods of VP1 administration were tested, either through endogenous expression in transformed cell lines, or through addition of purified VP1-based viral-like particles to the extracellular medium. In addition, exposure to CPV virions isolated from purified polyhedra was also performed. In general, our results do not show a robust transcriptional response of immune-related genes to VP1 or CPV virions, but two exceptions were noted. First, the expression of the antimicrobial peptide (AMP) gene Attacin was strongly induced after 24 h of exposure to VP1-based VLPs. Second, the expression levels of dcr-2, an essential gene in the RNAi pathway, were greatly increased in VP1-expressing transformed Sf21 cells but not transformed Bm5 cells, indicating the existence of species-specific effects. However, the increased expression of dcr-2 did not result in increased silencing efficiency when tested in an RNAi reporter assay. Our study indicates that the capsid protein VP1 of CPV has the potential to act as a PAMP and to induce a transcriptional response in insect cells that relate both to RNAi and protein effectors such as AMPs. The identity of the PRRs and the signaling cascade that are potentially triggered by VP1 remain to be elucidated in future experiments. While this study was performed on a small scale, it can encourage more comprehensive studies with high-throughput approaches (microarray, deep sequencing) to search more systematically whether viral capsid proteins can act as PAMPs in insects and whether their production results in the induction of immune-related genes with potential antiviral function.
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http://dx.doi.org/10.1007/s00438-019-01551-1DOI Listing
August 2019
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