Functional & Integrative Genomics最新文献

{"title":"Circ-IP6K2 suppresses tumor progression by modulating the miR-1292-5p/CAMK2N1 signal in clear cell renal cell carcinoma","authors":"Jian-ying Tang,&nbsp;Lu Yang,&nbsp;Qing-Jian Wu,&nbsp;Ying Yang,&nbsp;Yuan-Yuan Su,&nbsp;Yi-Rong Chen,&nbsp;Jiao Mu","doi":"10.1007/s10142-024-01398-9","DOIUrl":"10.1007/s10142-024-01398-9","url":null,"abstract":"<div><p>Renal cell carcinoma (RCC) is a malignant tumor originating from the epithelial cells of the renal tubules. The clear cell RCC subtype is closely linked to a poor prognosis due to its rapid progression. Circular RNA (circRNA) is a novel class of regulatory RNA molecules that play a role in the development of ccRCC, although their functions have not been fully elucidated. In this study, we identified a significant downregulation of circ-IP6K2 in ccRCC tissues based on data from the GSE100186 dataset. The decreased expression of circ-IP6K2 correlated with the progression of TNM stage and histological grade, and was also associated with decreased overall survival rates in ccRCC patients. Moreover, our findings revealed that circ-IP6K2 expression suppressed proliferation, migration, and invasion capabilities in vitro, and inhibited xenograft growth in vivo. Mechanistically, circ-IP6K2 acted as a sponge for miR-1292-5p in ccRCC cells, which in turn targeted the 3’UTR of CAMK2N1, leading to a decrease in its expression. CAMK2N1 was identified as a tumor suppressor that negatively regulated the β-catenin/c-Myc oncogenic signaling pathway. Additionally, we confirmed a positive correlation between the expression of circ-IP6K2 and CAMK2N1 in ccRCC. Circ-IP6K2 functions to impede the progression of ccRCC by modulating the miR-1292-5p/CAMK2N1 axis. These findings shed new light on the molecular mechanisms driving ccRCC progression and suggest potential therapeutic targets for the treatment of ccRCC.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141557769","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Purifying selection drove the adaptation of mitochondrial genes along with correlation of gene rearrangements and evolutionary rates in two subfamilies of Whitefly (Insecta: Hemiptera)","authors":"Abhishek Ghosh,&nbsp;Kaomud Tyagi,&nbsp;Anil Kumar Dubey,&nbsp;Andrew D. Sweet,&nbsp;Devkant Singha,&nbsp;Prathana Goswami,&nbsp;Vikas Kumar","doi":"10.1007/s10142-024-01400-4","DOIUrl":"10.1007/s10142-024-01400-4","url":null,"abstract":"<div><p>Insect mitochondrial genomes (mitogenomes) are usually represented by a conserved gene order. Whiteflies exhibit gene rearrangement in their mitogenomes; however, understanding how nucleotide substitution rates shape gene rearrangement in whiteflies is unclear due to the limited number of mitogenomes. Additionally, the mechanisms by which selection pressure drives adaptations in mitochondrial genes in the two subfamilies of whiteflies are not yet known. Here, we analyzed 18 whitefly mitogenomes, including one newly generated mitogenome, to compare nucleotide substitution rates, selection pressure, and gene arrangements. The newly generated mitogenome is reported along with reannotation of <i>Pealius mori</i> and comparisons to other whitefly mitogenomes. Comparative studies on nucleotide composition of 18 whiteflies revealed the positive GC skewness, confirming the reversal of strand asymmetry. We found 11 rearranged gene orders within two subfamilies of whiteflies with 8–18 breakpoints of gene rearrangements. Members of the subfamily Aleyrodinae exhibit more complex pathways in the evolution of gene order as compared to the subfamily Aleurodicinae. Our findings also revealed that the increase or reduction of nucleotide substitution rates does not have an impact on any of the gene rearrangement scenarios depicting neutral correlation. Selection pressure analysis revealed that the mitogenomes from members of both the subfamilies Aleurodicinae and Aleyrodinae are characterized by intense purifying selection pressure.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141553969","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Shaping Drosophila eggs: unveiling the roles of Arpc1 and cpb in morphogenesis","authors":"Poulami Chatterjee,&nbsp;Sandipan Mukherjee,&nbsp;Pralay Majumder","doi":"10.1007/s10142-024-01396-x","DOIUrl":"10.1007/s10142-024-01396-x","url":null,"abstract":"<div><p>The <i>Drosophila</i> egg chamber (EC) starts as a spherical tissue at the beginning. With maturation, the outer follicle cells of EC collectively migrate in a direction perpendicular to the anterior-posterior axis, to shape EC from spherical to ellipsoidal. Filamentous actin (F-actin) plays a significant role in shaping individual migratory cells to the overall EC shape, like in every cell migration. The primary focus of this article is to unveil the function of different Actin Binding Proteins (ABPs) in regulating mature <i>Drosophila</i> egg shape. We have screened 66 ABPs, and the genetic screening data revealed that individual knockdown of Arp2/3 complex genes and the “capping protein β” (<i>cpb</i>) gene have severely altered the egg phenotype. <i>Arpc1</i> and <i>cpb</i> RNAi mediated knockdown resulted in the formation of spherical eggs which are devoid of dorsal appendages. Studies also showed the role of <i>Arpc1</i> and <i>cpb</i> on the number of laid eggs and follicle cell morphology. Furthermore, the depletion of <i>Arpc1</i> and <i>cpb</i> resulted in a change in F-actin quantity. Together, the data indicate that <i>Arpc1</i> and <i>cpb</i> regulate <i>Drosophila</i> egg shape, F-actin management, egg-laying characteristics and dorsal appendages formation.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141496801","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comprehensive pan-cancer analysis reveals that C5orf34 regulates the proliferation and mortality of lung cancer","authors":"Meng Yang,&nbsp;Yuhan Deng,&nbsp;Yu Ma,&nbsp;Chunli Song,&nbsp;Zhenhua Wu,&nbsp;Xiayimaierdan Yibulayin,&nbsp;Xiaohong Sun,&nbsp;Yunquan Guo,&nbsp;Dan He","doi":"10.1007/s10142-024-01397-w","DOIUrl":"10.1007/s10142-024-01397-w","url":null,"abstract":"<p>The gene C5orf34 exhibits evolutionary conservation among mammals, and emerging evidence suggests its potential involvement in tumor development; however, comprehensive investigations of this gene are lacking. This study aims to elucidate the functional attributes and underlying mechanisms of C5orf34 in cancer. To evaluate its clinical predictive value, we conducted an analysis of the pan-cancerous expression, clinical data, mutation, and methylation data of C5orf34. Additionally, we investigated the correlation between C5orf34 and tumor mutant load (TMB), immune cell infiltration, and microsatellite instability (MSI) through relevant analyses. Furthermore, immunohistochemical (IHC) staining was employed to validate clinical samples, while knockdown and overexpression experiments and transcriptome RNA sequencing were utilized to examine the impact of C5orf34 on LUAD cells. According to our study, C5orf34 exhibits high expression levels in the majority of malignant tumors. The upregulation of C5orf34 is governed by DNA copy number alterations and methylation patterns, and it is closely associated with patients' survival prognosis and immune characteristics, thereby holding significant clinical implications. Furthermore, IHC staining analysis, cellular experiments, and transcriptome RNA sequencing have provided evidence supporting the role of C5orf34 in modulating the cell cycle to promote LUAD proliferation, migration, and invasion. This highlights its potential as a promising therapeutic target. The findings of this investigation suggest that C5orf34 may serve as a valuable biomarker for various tumor types and represent a potential target for immunotherapy, particularly in relation to the proliferation, migration, and apoptosis of LUAD cells.</p>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-06-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141475639","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Integrated multi-omics analysis and machine learning to refine molecular subtypes, prognosis, and immunotherapy in lung adenocarcinoma","authors":"Tao Han,&nbsp;Ying Bai,&nbsp;Yafeng Liu,&nbsp;Yunjia Dong,&nbsp;Chao Liang,&nbsp;Lu Gao,&nbsp;Jiawei Zhou,&nbsp;Jianqiang Guo,&nbsp;Jing Wu,&nbsp;Dong Hu","doi":"10.1007/s10142-024-01388-x","DOIUrl":"10.1007/s10142-024-01388-x","url":null,"abstract":"<div><p>Lung adenocarcinoma (LUAD) has a malignant characteristic that is highly aggressive and prone to metastasis. There is still a lack of suitable biomarkers to facilitate the refinement of precision-based therapeutic regimens. We used a combination of 10 known clustering algorithms and the omics data from 4 dimensions to identify high-resolution molecular subtypes of LUAD. Subsequently, consensus machine learning-related prognostic signature (CMRS) was developed based on subtypes related genes and an integrated program framework containing 10 machine learning algorithms. The efficiency of CMRS was analyzed from the perspectives of tumor microenvironment, genomic landscape, immunotherapy, drug sensitivity, and single-cell analysis. In terms of results, through multi-omics clustering, we identified 2 comprehensive omics subtypes (CSs) in which CS1 patients had worse survival outcomes, higher aggressiveness, mRNAsi and mutation frequency. Subsequently, we developed CMRS based on 13 key genes up-regulated in CS1. The prognostic predictive efficiency of CMRS was superior to most established LUAD prognostic signatures. CMRS demonstrated a strong correlation with tumor microenvironmental feature variants and genomic instability generation. Regarding clinical performance, patients in the high CMRS group were more likely to benefit from immunotherapy, whereas low CMRS were more likely to benefit from chemotherapy and targeted drug therapy. In addition, we evaluated that drugs such as neratinib, oligomycin A, and others may be candidates for patients in the high CMRS group. Single-cell analysis revealed that CMRS-related genes were mainly expressed in epithelial cells. The novel molecular subtypes identified in this study based on multi-omics data could provide new insights into the stratified treatment of LUAD, while the development of CMRS could serve as a candidate indicator of the degree of benefit of precision therapy and immunotherapy for LUAD.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141454501","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Engineering circular RNA for molecular and metabolic reprogramming","authors":"Narendra Kumar Sharma,&nbsp;Pragya Dwivedi,&nbsp;Ravi Bhushan,&nbsp;Pawan Kumar Maurya,&nbsp;Abhishek Kumar,&nbsp;Tikam Chand Dakal","doi":"10.1007/s10142-024-01394-z","DOIUrl":"10.1007/s10142-024-01394-z","url":null,"abstract":"<div><p>The role of messenger RNA (mRNA) in biological systems is extremely versatile. However, it’s extremely short half-life poses a fundamental restriction on its application. Moreover, the translation efficiency of mRNA is also limited. On the contrary, circular RNAs, also known as circRNAs, are a common and stable form of RNA found in eukaryotic cells. These molecules are synthesized via back-splicing. Both synthetic circRNAs and certain endogenous circRNAs have the potential to encode proteins, hence suggesting the potential of circRNA as a gene expression machinery. Herein, we aim to summarize all engineering aspects that allow exogenous circular RNA (circRNA) to prolong the time that proteins are expressed from full-length RNA signals. This review presents a systematic engineering approach that have been devised to efficiently assemble circRNAs and evaluate several aspects that have an impact on protein production derived from. We have also reviewed how optimization of the key components of circRNAs, including the topology of vector, 5′ and 3′ untranslated sections, entrance site of the internal ribosome, and engineered aptamers could be efficiently impacting the translation machinery for molecular and metabolic reprogramming. Collectively, molecular and metabolic reprogramming present a novel way of regulating distinctive cellular features, for instance growth traits to neoplastic cells, and offer new possibilities for therapeutic inventions.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-06-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141449246","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Navigating the intricacies of RT-qPCR data analysis in gene expression studies","authors":"Linah Wetthasinghe,&nbsp;Hien Fuh Ng,&nbsp;Yun Fong Ngeow,&nbsp;Kee Seang Chew,&nbsp;Way Seah Lee","doi":"10.1007/s10142-024-01393-0","DOIUrl":"10.1007/s10142-024-01393-0","url":null,"abstract":"","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141441923","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Reviving resilience: MEcPP-mediated ASK1-IMPα-9-TRP2 stress-responsive module","authors":"Hamza Sohail,&nbsp;Iqra Noor,&nbsp;Xuewen Xu,&nbsp;Rahat Sharif,&nbsp;Xuehao Chen,&nbsp;Xiaodong Yang","doi":"10.1007/s10142-024-01395-y","DOIUrl":"10.1007/s10142-024-01395-y","url":null,"abstract":"<div><p>Chloroplasts are not only critical photosynthesis sites in plants, but they also participate in plastidial retrograde signaling in response to developmental and environmental signals. MEcPP (2-C-Methyl-D-erythritol-2,4-cyclopyrophosphate) is an intermediary in the methylerythritol phosphate (MEP) pathway in chloroplasts. It is a critical precursor for the synthesis of isoprenoids and terpenoid derivatives, which play crucial roles in plant growth and development, photosynthesis, reproduction, and defense against environmental constraints. Accumulation of MEcPP under stressful conditions triggers the expression of IMPα-9 and TPR2, contributing to the activation of abiotic stress-responsive genes. In this correspondence, we discuss plastidial retrograde signaling in support of a recently published paper in Molecular Plant (Zeng et al. 2024). We hope that it can shed more insight on the retrograde signaling cascade.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141441924","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Pharmacological inhibition of RUNX1 reduces infarct size after acute myocardial infarction in rats and underlying mechanism revealed by proteomics implicates repressed cathepsin levels","authors":"Hengshu Chen,&nbsp;Si Wang,&nbsp;Xiaoling Zhang,&nbsp;Xing Hua,&nbsp;Meng Liu,&nbsp;Yanan Wang,&nbsp;Simiao Wu,&nbsp;Weihong He","doi":"10.1007/s10142-024-01391-2","DOIUrl":"10.1007/s10142-024-01391-2","url":null,"abstract":"<div><p>Myocardial infarction (MI) results in prolonged ischemia and the subsequent cell death leads to heart failure which is linked to increased deaths or hospitalizations. New therapeutic targets are urgently needed to prevent cell death and reduce infarct size among patients with MI. Runt-related transcription factor-1 (RUNX1) is a master-regulator transcription factor intensively studied in the hematopoietic field. Recent evidence showed that RUNX1 has a critical role in cardiomyocytes post-MI. The increased RUNX1 expression in the border zone of the infarct heart contributes to decreased cardiac contractile function and can be therapeutically targeted to protect against adverse cardiac remodelling. This study sought to investigate whether pharmacological inhibition of RUNX1 function has an impact on infarct size following MI. In this work we demonstrate that inhibiting RUNX1 with a small molecule inhibitor (Ro5-3335) reduces infarct size in an in vivo rat model of acute MI. Proteomics study using data-independent acquisition method identified increased cathepsin levels in the border zone myocardium following MI, whereas heart samples treated by RUNX1 inhibitor present decreased cathepsin levels. Cathepsins are lysosomal proteases which have been shown to orchestrate multiple cell death pathways. Our data illustrate that inhibition of RUNX1 leads to reduced infarct size which is associated with the suppression of cathepsin expression. This study demonstrates that pharmacologically antagonizing RUNX1 reduces infarct size in a rat model of acute MI and unveils a link between RUNX1 and cathepsin-mediated cell death, suggesting that RUNX1 is a novel therapeutic target that could be exploited clinically to limit infarct size after an acute MI.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-06-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11166773/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141305058","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Iodine-125 seed inhibits proliferation and promotes apoptosis of cholangiocarcinoma cells by inducing the ROS/p53 axis","authors":"Fuping Kang,&nbsp;Jing Wu,&nbsp;Li Hong,&nbsp;Peng Zhang,&nbsp;Jianjun Song","doi":"10.1007/s10142-024-01392-1","DOIUrl":"10.1007/s10142-024-01392-1","url":null,"abstract":"<div><p>With advances in radioactive particle implantation in clinical practice, Iodine-125 (¹²⁵I) seed brachytherapy has emerged as a promising treatment for cholangiocarcinoma (CCA), showing good prognosis; however, the underlying molecular mechanism of the therapeutic effect of ¹²⁵I seed is unclear. To study the effects of ¹²⁵I seed on the proliferation and apoptosis of CCA cells. CCA cell lines, RBE and HCCC-9810, were treated with reactive oxygen species (ROS) scavenger acetylcysteine (NAC) or the p53 functional inhibitor, pifithrin-α hydrobromide (PFTα). Cell counting kit-8 (CCK-8) assay, 5-bromo-2-deoxy-uridine (BrdU) staining, and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay and flow cytometry assay were performed to test the radiation-sensitivity of ¹²⁵I seed toward CCA cells at different radiation doses (0.4 mCi and 0.8 mCi). 2,7-dichlorofluorescein diacetate (DCF-DA) assay, real-time quantitative polymerase chain reaction (RT-qPCR), and western blot analysis were performed to assess the effect of ¹²⁵I seed on the ROS/p53 axis. A dose-dependent inhibitory effect of ¹²⁵I seeds on the proliferation of CCA cells was observed. The ¹²⁵I seed promoted apoptosis of CCA cells and induced the activation of the ROS/p53 pathway in a dose-dependent manner. NAC or PFTα treatment effectively reversed the stimulatory effect of ¹²⁵I seed on the proliferation of CCA cells. NAC or PFTα suppressed apoptosis and p53 protein expression induced by the ¹²⁵I seed. ¹²⁵I seed can inhibit cell growth mainly through the apoptotic pathway. The mechanism may involve the activation of p53 and its downstream apoptotic pathway by up-regulating the level of ROS in cells.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2024-06-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11166828/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141305057","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}