Functional & Integrative Genomics最新文献

{"title":"ALKBH5-mediated NPC2 mRNA m6A demethylation promotes resistance to oxaliplatin in colorectal cancer","authors":"Peng Wan,&nbsp;Yuan Ren,&nbsp;Hong-Tao Li","doi":"10.1007/s10142-025-01651-9","DOIUrl":"10.1007/s10142-025-01651-9","url":null,"abstract":"<div><p>Colorectal cancer (CRC) is the third most common cancer globally and a leading cause of cancer-related death. Oxaliplatin, a key platinum-based chemotherapy, significantly improves outcomes in CRC patients. Nevertheless, oxaliplatin resistance often emerges, leading to worse prognosis. Exploring new biomarkers and mechanisms of resistance is crucial for overcoming oxaliplatin resistance and enhancing therapeutic efficacy. Through bioinformatics analysis, high NPC2 expression was found to be associated with oxaliplatin resistance and poor prognosis in CRC patients. Moreover, NPC2 was highly expressed in CRC tissues, especially in metastatic CRC tissues. Additionally, the expression of N6-methyladenosine (m⁶A) demethylase ALKBH5 was elevated in oxaliplatin-resistant colorectal cancer cells. Mechanically, ALKBH5 promotes m⁶A demethylation of NPC2 mRNA in a YTHDF2-dependent process, thereby enhancing the stability of NPC2 mRNA and making colorectal cancer cells oxaliplatin-resistant. Our results show that by inhibiting NPC2 or ALKBH5, we can re-sensitize resistant CRC cells to oxaliplatin in vitro and in vivo. In summary, ALKBH5-mediated m⁶A demethylation promotes the stability of NPC2 mRNA and plays a key role in promoting oxaliplatin resistance in colorectal cancer. Targeting the ALKBH5/NPC2 axis have important therapeutic potential for patients with oxaliplatin-resistant colorectal cancer.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12274260/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144666805","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":"Phosphoenolpyruvate carboxykinase 2 (PCK2) attenuates bovine adipocyte lipolysis through PNPLA2 activity repression","authors":"Chao Jiang,&nbsp;Hao Wu,&nbsp;Jiupan Zhang,&nbsp;Yaping Song,&nbsp;Xiaoyu Song,&nbsp;Ruopu Jiao,&nbsp;Sayed Haidar Abbas Raza,&nbsp;Sameer Dinkar Pant,&nbsp;Linsen Zan,&nbsp;Dawei Wei","doi":"10.1007/s10142-025-01666-2","DOIUrl":"10.1007/s10142-025-01666-2","url":null,"abstract":"<div>\u0000 \u0000 <p>Marbling traits serve as a key determinant of meat quality in livestock, with interspecies genetic variations significantly influencing intramuscular fat (IMF) deposition. Through systematic analysis of public databases and cross-species RNA sequencing data from cattle, sheep, and pigs exhibiting divergent IMF levels, we identified <i>PCK2</i> as a conserved regulatory factor governing lipogenesis. Functional analyses demonstrated that targeted <i>PCK2</i> silencing significantly impaired cell cycle dynamics and suppressed adipocyte proliferation, which mechanistically involved transcriptional downregulation of key cell cycle regulators (<i>CCNB1</i>, <i>CCND1</i>, <i>CCNE2</i>) and functional inhibition of critical proliferative mediators (<i>CDK1</i>, <i>CDK2</i>, <i>PCNA</i>). Furthermore, <i>PCK2</i> silencing impaired lipid accumulation by decreasing triglyceride synthesis and lipid droplet formation, while down-regulating adipogenic transcription factors <i>PPARγ</i> and <i>C/EBPα</i>, as well as lipid metabolism factors <i>SCD1</i>, <i>ACSL1</i> and <i>FABP4</i>. Integrated multi-omics analysis revealed <i>PCK2</i> modulates bovine IMF deposition through the regulation of lipolysis in adipocyte. Notably, patatin-like phospholipase domain containing 2 (PNPLA2) suppression as a novel mechanism underlying PCK2-mediated enhancement of bovine intramuscular adipocyte differentiation. These findings establish a molecular framework for understanding <i>PCK2</i>’s regulatory role in marbling beef production.</p>\u0000 <span>AbstractSection</span>\u0000 Graphical Abstract\u0000 <div><figure><div><div><picture><source><img></source></picture></div></div></figure></div>\u0000 \u0000 </div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144658037","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":"Integrating multiomics and modern breeding tools for accelerating genetic improvement in Annonas","authors":"Shri Hari Prasad,&nbsp;Grant Bignell,&nbsp;Rhys G.R. Copeland,&nbsp;Vanika Garg,&nbsp;Annapurna Chitikineni,&nbsp;Robert J. Henry,&nbsp;Natalie Dillon,&nbsp;Reyazul Rouf Mir,&nbsp;Rajeev K. Varshney","doi":"10.1007/s10142-025-01653-7","DOIUrl":"10.1007/s10142-025-01653-7","url":null,"abstract":"<div><p>Custard apples (<i>Annona</i> spp.) are among the most important horticultural crops in the world, including Australia. The genus <i>Annona</i> comprises several economically and nutritionally significant species, including atemoya, cherimoya, sugar apple, ilama, soursop, bullock’s heart, and bibra. These fruits are valued for their exotic taste and are popular backyard fruit crops in many countries. While some species are commercially cultivated and exported, the broader potential of these crops remains largely untapped. Despite their historical significance, these <i>Annona</i> species remain neglected or underutilised, with breeding efforts restricted to only a few countries. Extensive genetic resources, including germplasm collections, candidate genotypes, and mapping populations, are available for crop improvement. Traditional breeding methods - such as selection, crossbreeding, and mutation breeding – have been widely applied alongside modern breeding approaches like marker-assisted selection (MAS). However, several challenges, such as a lack of information regarding the crop and a long juvenile period, hinder crop improvement in custard apples. Recent advancements and affordability of sequencing technologies have enabled an increase in the number of multiomics studies, especially genomics and transcriptomics within <i>Annona</i> species. Integrating these data with proteomics, metabolomics, and phenomics will facilitate the genetic dissection of important traits in <i>Annona</i>. This review provides a comprehensive overview of the current advancements and future prospects of multiomics tools and technologies developed and their potential to accelerate custard apple breeding programs.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12254078/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144615675","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":"Methyltransferase ZC3H13 regulates ferroptosis of alveolar macrophages in sepsis-associated acute lung injury via PRDX6/p53/SLC7A11 axis","authors":"Jifang Liang,&nbsp;Zemin Liu,&nbsp;Yajun He,&nbsp;Heihei Li,&nbsp;Weidong Wu","doi":"10.1007/s10142-025-01659-1","DOIUrl":"10.1007/s10142-025-01659-1","url":null,"abstract":"<div><p>Peroxiredoxin 6 (PRDX6) is widely acknowledged as a suppressor of ferroptosis, and recent studies have demonstrated that inhibition of macrophage ferroptosis can alleviate sepsis-associated acute lung injury (SA-ALI). Nonetheless, the specific involvement of PRDX6 in regulating macrophage ferroptosis during SA-ALI remains unexplored. This study aims to elucidate the mechanistic role of PRDX6 in modulating macrophage ferroptosis within the context of SA-ALI. Mouse alveolar macrophages (MH-S cells) were infected with either a PRDX6 overexpression lentivirus or a ZC3H13 knockdown lentivirus prior to lipopolysaccharide (LPS) treatment. In vivo, mice were treated with the same lentiviral constructs and subjected to a SA-ALI model via cecal ligation and puncture (CLP). This study demonstrates that PRDX6 overexpression or ZC3H13 knockdown significantly attenuated LPS-induced ferroptosis in alveolar macrophages and alleviated lung injury in CLP-induced SA-ALI mouse models. However, simultaneous knockdown of both ZC3H13 and PRDX6 abolished the protective effect conferred by ZC3H13 silencing, indicating that PRDX6 mediates the anti-ferroptotic role of ZC3H13 inhibition. Mechanistically, PRDX6 suppresses p53 expression, thereby upregulating SLC7A11 and inhibiting ferroptosis. Additionally, ZC3H13 promotes the m6A modification of PRDX6 mRNA, which facilitates its degradation in a YTHDF2-dependent manner, ultimately leading to reduced PRDX6 expression. Overall, these findings demonstrate that the methyltransferase ZC3H13 modulates PRDX6 expression by elevating the m6A methylation level of PRDX6 mRNA in a YTHDF2-dependent manner, thereby influencing the p53/SLC7A11 axis and promoting ferroptosis in alveolar macrophages, ultimately contributing to the progression of SA-ALI.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144615676","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":"Integration of scRNA-seq and bulk tissue RNA-seq data to identify cancer-associated fibroblast-related gene RGMA as a potential treatment target for esophageal cancer","authors":"Yanqing Gao,&nbsp;Shuguang Bao,&nbsp;Bao Wen,&nbsp;Haoyuan Li,&nbsp;Qiang Guo,&nbsp;Ao Li,&nbsp;Luri Bao,&nbsp;Meitao Li,&nbsp;Bateer Han","doi":"10.1007/s10142-025-01660-8","DOIUrl":"10.1007/s10142-025-01660-8","url":null,"abstract":"<div><p>Cancer-associated fibroblasts (CAFs) serve as key stromal components within tumor microenvironment (TME), playing a significant role in the development and outcome of esophageal cancer (EC). There is an urgent need to identify genes related to CAFs to improve treatment strategies. The scRNA-sequencing dataset GSE196756 were used to identify fibroblast-related genes. Additionally, a WGCNA analysis was also conducted to identify modules related to CAFs within the TCGA-esophageal carcinoma (ESCA) cohort. By taking the intersection of identified genes of these two sections, CAF-related genes were identified. Expression of RGMA in EC samples compared to normal controls was assessed by RT-qPCR and western blot. In vitro and in vivo experiments were conducted to assess the impact of RGMA on EC cell growth. Compared to adjacent normal tissues, the levels of RGMA were notably reduced in EC tissues. Reduced RGMA levels were linked to a poor prognosis for EC patients. Furthermore, RGMA was found to have a positive correlation with the expression of fibroblast-related gene DCN, and showed a negative correlation with the expression of tumor-promoting chemokines, CXCL1, CXCL3 and CXCL8. Functionally, RGMA overexpression strongly prevented ECA109 cell viability, proliferation and migration, as well as suppresses tumor growth in vivo, suggesting that RGMA may function as a tumor suppressor in EC. Additionally, RGMA levels were also remarkably decreased in human esophageal CAFs relative to esophageal fibroblast cells (NFs). Importantly, the downregulation of RGMA may facilitate the transdifferentiation of NFs into CAFs by activating Akt signaling or upregulating CXCL1, CXCL3, and CXCL8, subsequently contributing to ECA109 cell proliferation. Collectively, RGMA may serve as a prognostic marker and a potential therapeutic target for EC.</p><p>\u0000 <b>Clinical trial number</b></p><p>\u0000 Not applicable.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144607005","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":"SPP1 promotes malignant characteristics and drug resistance in hepatocellular carcinoma by activating fatty acid metabolic pathway","authors":"Zhijiang Wang,&nbsp;Chengfang Wang","doi":"10.1007/s10142-025-01664-4","DOIUrl":"10.1007/s10142-025-01664-4","url":null,"abstract":"<div><p>Hepatocellular carcinoma (HCC) progression and prognosis are influenced by various molecular markers. This study aimed to identify the hub gene associated with HCC clinical characteristics and its role in HCC progression. Differentially expressed genes (DEGs) between HCC tumor and normal tissues, as well as between stage I/II and stage III/IV, were analyzed. Machine learning algorithms were used to pinpoint three critical hub genes (SPP1, ADH4, and ANXA10). A prognostic risk model was constructed and evaluated using Kaplan-Meier curves, COX regression, and decision curve analysis, which could effectively predict HCC survival. Among the three hub genes, SPP1 was significantly associated with the overall survival (OS) of HCC patients and effectively predicted prognosis. More importantly, SPP1 was upregulated in HCC tumor tissues and cells, and its overexpression enhanced HCC cell proliferation, migration, invasion, and drug resistance. It also promoted fatty acid metabolism in HCC cells, with malignant characteristics and drug resistance induced by SPP1 being mitigated by fatty acid oxidation inhibition. In vivo experiments showed that SPP1 knockdown inhibited tumor growth and fatty acid metabolism of HCC mice. In conclusion, SPP1 is a pivotal gene that influences HCC prognosis by enhancing malignancy and drug resistance through fatty acid metabolism.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144607006","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":"USP6NL knockdown suppresses colorectal cancer progression by inducing CASP9-Mediated apoptosis and disrupting FOXC2/SNAI1-Driven EMT and angiogenesis","authors":"Mohammed Ahmed Mohammed,&nbsp;Ahmed Abdelmagied Soffar,&nbsp;Amany I. Yousef,&nbsp;Mohammed Salama,&nbsp;Fawziya A. R. Ibrahim,&nbsp;Tarek El-Sewedy,&nbsp;Alaa Elmetwalli","doi":"10.1007/s10142-025-01663-5","DOIUrl":"10.1007/s10142-025-01663-5","url":null,"abstract":"&lt;div&gt;&lt;h3&gt;Background&lt;/h3&gt;&lt;p&gt;Colorectal cancer (CRC) remains a leading cause of cancer-related mortality worldwide, with tumor progression often driven by dysregulated oncogenic pathways. USP6NL, a known regulator of endocytic trafficking, has recently been implicated in tumorigenesis. However, its precise role in CRC remains unclear, and more studies are still needed to deepen our understanding of underlying mechanisms implicated in its oncogenic role. Therefore, silencing USP6NL could provide a novel therapeutic strategy by concurrently disrupting several oncogenic mechanisms, creating a new avenue for CRC management, particularly in patients who develop resistance against conventional therapies. This study investigates the impact of USP6NL knockdown on CRC cell morphology, proliferation, apoptosis, migration, angiogenesis, and metabolic adaptation, providing mechanistic insights into its oncogenic functions.&lt;/p&gt;&lt;h3&gt;Methods&lt;/h3&gt;&lt;p&gt;HCT116 colorectal cancer cells were transfected with USP6NL-specific siRNA. Immunocytochemistry was used to confirm successful silencing, functional assays were performed to assess changes in cell morphology using phase-contrast and scanning electron microscopy, and colony formation and wound healing assays were performed to assess cell clonogenic capacity and migration, respectively, in addition to apoptosis assay via flow cytometry, and RT²-Profiler PCR array to measure variation in gene expression of 84 cancer-related genes. Statistical analyses were performed to evaluate significant differences between control and USP6NL-silenced groups.&lt;/p&gt;&lt;h3&gt;Results&lt;/h3&gt;&lt;p&gt;USP6NL depletion led to profound morphological changes, including membrane blebbing, cell shrinkage, and loss of adhesion, reflecting late apoptotic features of cells. These findings were further supported by flow cytometry, which confirmed increased apoptosis, with a higher proportion of late apoptotic cells (20.99% in USP6NL knockdown vs. 2.69% in control, &lt;i&gt;p&lt;/i&gt; = 0.042). Colony formation assays revealed a significant reduction in the clonogenic potential, suggesting a critical role of USP6NL in promoting CRC cell proliferation (&lt;i&gt;p&lt;/i&gt; ≤ 0.05). The wound healing assay demonstrated impaired migration in USP6NL-silenced cells, with a marked delay in wound closure (&lt;i&gt;p&lt;/i&gt; = 0.0201 at 48 h). Gene expression analysis revealed a significant downregulation of VEGFC (-8.62-fold) and ANGPT2 (-4.03-fold), impairing angiogenesis and suppressing FOXC2, SNAI1, and SNAI2, indicating EMT inhibition. Additionally, CASP9, APAF1, and BCL2L11 were upregulated, confirming the activation of intrinsic apoptosis, while metabolic regulators HIF1A and LDHA were downregulated, suggesting impaired tumor hypoxic adaptation.&lt;/p&gt;&lt;h3&gt;Conclusion&lt;/h3&gt;&lt;p&gt;This study establishes USP6NL as a key modulator of CRC progression, regulating proliferation, apoptosis, migration, angiogenesis, and metabolic pathways. The loss of USP6NL leads to EMT suppression, apoptosis induction, and reduced tu","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144607007","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":"Genome-wide characterization of invertases in Arabica coffee and its progenitors reveals putative genes involved in fruit development","authors":"Gabriela Ester Ferraz,&nbsp;Robert Márquez Gutiérrez,&nbsp;Tiago Yukio Inoue,&nbsp;Gabriel de Campos Rume,&nbsp;Thais Aparecida Sales,&nbsp;Teodorico Castro Ramalho,&nbsp;Evandro Novaes,&nbsp;Dapeng Zhang,&nbsp;Antonio Chalfun-Junior,&nbsp;Flavia Maria Avelar Gonçalves","doi":"10.1007/s10142-025-01662-6","DOIUrl":"10.1007/s10142-025-01662-6","url":null,"abstract":"<div><p>In plant physiological processes, invertases act by irreversibly degrading sucrose into hexoses, which actively contribute to plant growth and development. Few invertase genes have been characterized in <i>Coffea</i>, and consequently, the role of these enzymes in coffee plant development is not well understood. The present study conducted a genome-wide characterization of invertases in Arabica coffee and its diploid progenitors. A total of 65 invertase genes were identified, with 28 in <i>C. arabica</i> (<i>CaINV</i>), 18 in <i>C. canephora</i> (<i>CcINV</i>), and 19 in <i>C. eugenioides</i> (<i>CeINV</i>), with polyploidization being the primary cause of the expansion of the <i>CaINV</i> gene family. Synteny analyses and chromosomal mapping showed that the <i>CaINV</i> genes are mainly derived from <i>C. eugenioides</i>. The cis-acting elements suggested the involvement of invertase genes in three main categories: growth and development, responses to biotic and abiotic stress, and hormonal response. Transcriptome analyses showed that, except for the neutral/alkaline invertase gene <i>CaN/AINV1</i>, all invertase genes of <i>C. arabica</i> and <i>C. canephora</i> were expressed at different stages of fruit development, some constitutively and others conditionally. Although <i>CaN/AINV1</i> was not expressed in the evaluated tissues, the presence of two GCN4 cis-elements suggests its involvement in specific processes in tissues or stages not analyzed in this study. Our results open many avenues for future studies in <i>Coffea</i>, especially for <i>C. arabica</i> and <i>C. canephora</i>, which are economically most significant and exhibit particularities regarding disease resistance, tolerance to abiotic stresses, and fruit chemical composition.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144607004","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":"Efficient gene editing of BMP15, GDF9, and MSTN—but not the imprinted CLPG gene—in goat embryos via electrotransfection and handmade cloning","authors":"Mehdi Hajian,&nbsp;Ahmad Pirali,&nbsp;Seyed Hossein Hosseini Moghaddam,&nbsp;Reza Moradi-Hajidavaloo,&nbsp;Shiva Rouhollahi Varnosfaderani,&nbsp;Mina Jozi,&nbsp;Tayebeh Izadi,&nbsp;Nima Tanhaie-vash,&nbsp;Wilfried Kues,&nbsp;Mohammad-Hossein Nasr-Esfahani,&nbsp;Farnoosh Jafarpour,&nbsp;Shahin Eghbalsaied","doi":"10.1007/s10142-025-01644-8","DOIUrl":"10.1007/s10142-025-01644-8","url":null,"abstract":"<div><p>CRISPR/Cas9 technology represents a powerful tool for advancing livestock breeding by enabling precise, on-target gene edits without the genomic mixing associated with traditional introgression methods. In this study, we employed a dual gRNA-based CRISPR/Cas9 strategy to induce targeted deletions and indel mutations in both reproductive and growth-related genes. These included the metacentric genes bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9), which are associated with increased ovulation rate and litter size, as well as the telomeric genes myostatin (MSTN) and callipyge (CLPG), which are linked to muscle development and enhanced meat production. We employed an optimized electrotransfection protocol consisting of 10–20 µg of each plasmid DNA, 250 µL OptiMEM-GlutaMAX, and one million goat fibroblast cells. The electroporation was performed using a Bio-Rad system in a 4-mm cuvette, with two 10-millisecond pulses at 270 volts, separated by a 10-second interval. This protocol enabled efficient genome editing of goat embryonic fibroblast cells, which were subsequently used to generate cloned embryos via handmade somatic cell nuclear transfer (SCNT), involving manual enucleation and cell–oocyte fusion steps. Sequencing revealed high mutation rates (78–97%) and a predominance of biallelic edits in <i>BMP15</i>, <i>GDF9</i>, and <i>MSTN</i>. Notably, <i>MSTN</i> gRNAs with a 7-bp overlapping sequence at their 3′ ends showed a high editing efficiency. In contrast, the imprinted <i>CLPG</i> gene exhibited a significantly lower mutation rate (~ 30%), likely due to epigenetic constraints. While overall mutation rates did not differ significantly between metacentric and telomeric genes, on-target deletions were more frequent in metacentric genes (43%) than in telomeric ones (20%). Embryo development rates from gene-edited cells were comparable to those from non-edited controls. These findings underscore the utility of combining electrotransfection with SCNT for efficient editing of non-imprinted genes and highlight the need for improved strategies to overcome barriers in editing imprinted loci.</p></div>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144599058","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 Nanopore and short-read RNA sequencing identifies dysregulation of METTL3- m6A modifications in endocrine therapy- sensitive and resistant breast cancer cells.","authors":"Belinda J Petri, Kellianne M Piell, Bailey L Avila-Valdes, Christian G Stanley, Laura J Winkler, Johnny Tyler Brown, Rye Ulett, Gabriela Sanchez, Julia H Chariker, Eric C Rouchka, Carolyn M Klinge","doi":"10.1007/s10142-025-01658-2","DOIUrl":"10.1007/s10142-025-01658-2","url":null,"abstract":"<p><p>The role of epitranscriptomic changes in the development of acquired endocrine therapy (ET)- resistance in estrogen receptor α (ER) expressing breast cancer (BC) is unknown. We tested the hypothesis that inhibition of METTL3, the methyltransferase responsible for the mRNA modification N-6 methyladenosine (m6A), alters m6A modifications and differentially regulates the abundance of mRNA transcripts in ET-sensitive MCF-7 versus resistant LCC9 ER + human BC cells. Differential m6A modifications were identified using direct mRNA sequencing (DRS) performed on five replicates for each cell line ± 1 µM STM2457, a selective METTL3 inhibitor, using Nanopore MinION long read RNA-seq. Parallel short read Illumina RNA-seq quantified differential transcript abundance in the same samples. Selected results were validated by RT-qPCR, m6A-RIP-qPCR, reporter assays, and western blot analysis. Statistical analysis combined m6Anet, a machine-learning algorithm designed to call m6A modified bases, with a generalized linear model following a binomial distribution analysis to identify significant differential m6A modification ratios (DMR). Distinct METTL3 dependent m6A modification patterns in LCC9 and MCF-7 cells were observed in differentially expressed genes (DEG) associated with ET-resistance, including EEF1A2, ACTB, FLNA, PDIA6, AMIGO2, TPT1, XBP1, and CITED4. Select results were validated in additional ET-resistant BC cell lines. m6A-RIP-RT-qPCR validated specific m6A sites. We examined the proximity of m6A sites to estrogen receptor α (ER α)-mRNA binding sites reported in MCF-7 cells. ACTB, PDIA6, and XBP1 demonstrated a short-range proximity, with m6A sites located within 100 bp of ERα binding sites, suggesting a role for m6A in influencing ERα-mRNA binding. Our work provides a framework for integrating DRS and DEG omics data. Our results suggest a role for dysregulation of m6A modifications in pathways implicated in ET resistance in BC.</p>","PeriodicalId":574,"journal":{"name":"Functional & Integrative Genomics","volume":"25 1","pages":"149"},"PeriodicalIF":3.9,"publicationDate":"2025-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12241209/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144590178","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}