Pub Date : 2023-08-08DOI: 10.1186/s12860-023-00484-3
Ibrahim Khater, Aaya Nassar
Background: Cells can die through a process called apoptosis in both pathological and healthy conditions. Cancer development and progression may result from abnormal apoptosis. The 78-kDa glucose-regulated protein (GRP78) is increased on the surface of cancer cells. Kringle 5, a cell apoptosis agent, is bound to GRP78 to induce cancer cell apoptosis. Kringle 5 was docked to GRP78 using ClusPro 2.0. The interaction between Kringle 5 and GRP78 was investigated.
Results: The interacting amino acids were found to be localized in three areas of Kringle 5. The proposed peptide is made up of secondary structure amino acids that contain Kringle 5 interaction residues. The 3D structure of the peptide model amino acids was created using the PEP-FOLD3 web tool.
Conclusions: The proposed peptide completely binds to the GRP78 binding site on the Kringle 5, signaling that it might be effective in the apoptosis of cancer cells.
{"title":"A computational peptide model induces cancer cells' apoptosis by docking Kringle 5 to GRP78.","authors":"Ibrahim Khater, Aaya Nassar","doi":"10.1186/s12860-023-00484-3","DOIUrl":"10.1186/s12860-023-00484-3","url":null,"abstract":"<p><strong>Background: </strong>Cells can die through a process called apoptosis in both pathological and healthy conditions. Cancer development and progression may result from abnormal apoptosis. The 78-kDa glucose-regulated protein (GRP78) is increased on the surface of cancer cells. Kringle 5, a cell apoptosis agent, is bound to GRP78 to induce cancer cell apoptosis. Kringle 5 was docked to GRP78 using ClusPro 2.0. The interaction between Kringle 5 and GRP78 was investigated.</p><p><strong>Results: </strong>The interacting amino acids were found to be localized in three areas of Kringle 5. The proposed peptide is made up of secondary structure amino acids that contain Kringle 5 interaction residues. The 3D structure of the peptide model amino acids was created using the PEP-FOLD3 web tool.</p><p><strong>Conclusions: </strong>The proposed peptide completely binds to the GRP78 binding site on the Kringle 5, signaling that it might be effective in the apoptosis of cancer cells.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-08-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10408047/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10019395","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Background: Bone morphogenetic protein 9 (BMP9) has been shown to regulate processes such as angiogenesis, endothelial dysfunction, and tumorigenesis. However, the role of BMP9 in preeclampsia (PE) is unclear. The purpose of this study was to investigate the role and mechanism of BMP9 in PE.
Methods: The effects of BMP9 on the viability, migration and invasion of HTR-8/Svneo cells were investigated by CCK-8 assay, wound healing assay and Transwell invasion assay. The effect of BMP9 on apoptosis of HTR-8/Svneo cells was detected by flow cytometry. Plasma levels of BMP9, SDF1 and CXCR4 were detected by ELISA kit. qRT-PCR and Western blot were used to detect the expression levels of each gene in the cells.
Results: Overexpression of BMP9 promoted the proliferation and migration of trophoblast cells and inhibited apoptosis. Knockdown of BMP9 had the opposite effect. The levels of BMP9, SDF1 and CXCR4 in the plasma of PE patients were down-regulated, and BMP9 was positively correlated with the levels of SDF1 and CXCR4. BMP9 also significantly upregulated the mRNA and protein levels of SDF1 and CXCR4 in HTR-8/SVneo cells. Further mechanistic studies found that BMP9 promoted the migration and invasion of HTR-8/SVneo cells and inhibited apoptosis by activating the SDF1/CXCR4 pathway.
Conclusion: We demonstrate for the first time that BMP9 promoted the migration and invasion of HTR-8/SVneo cells and inhibits apoptosis by activating the SDF1/CXCR4 pathway. This suggests that BMP9 may be a biomarker molecule for PE.
背景:骨形态发生蛋白9 (Bone morphogenetic protein 9, BMP9)已被证明可调节血管生成、内皮功能障碍和肿瘤发生等过程。然而,BMP9在子痫前期(PE)中的作用尚不清楚。本研究的目的是探讨BMP9在PE中的作用和机制。方法:采用CCK-8法、创面愈合法和Transwell侵袭法研究BMP9对HTR-8/Svneo细胞活力、迁移和侵袭的影响。流式细胞术检测BMP9对HTR-8/Svneo细胞凋亡的影响。ELISA试剂盒检测血浆BMP9、SDF1、CXCR4水平。采用qRT-PCR和Western blot检测细胞中各基因的表达水平。结果:BMP9过表达促进滋养细胞增殖和迁移,抑制细胞凋亡。BMP9基因的敲低则有相反的效果。PE患者血浆BMP9、SDF1、CXCR4水平下调,BMP9与SDF1、CXCR4水平呈正相关。BMP9还显著上调HTR-8/SVneo细胞中SDF1和CXCR4的mRNA和蛋白水平。进一步的机制研究发现BMP9通过激活SDF1/CXCR4通路,促进HTR-8/SVneo细胞的迁移和侵袭,抑制细胞凋亡。结论:我们首次证实BMP9通过激活SDF1/CXCR4通路促进HTR-8/SVneo细胞的迁移和侵袭,抑制细胞凋亡。这表明BMP9可能是PE的生物标志物分子。
{"title":"BMP9 maintains the phenotype of HTR-8/Svneo trophoblast cells by activating the SDF1/CXCR4 pathway.","authors":"Xue Yang, Lingling Ren, Xiang Chen, Ying Pang, Baoxia Jia, Jing Sun, Xiaofang Quan","doi":"10.1186/s12860-023-00487-0","DOIUrl":"10.1186/s12860-023-00487-0","url":null,"abstract":"<p><strong>Background: </strong>Bone morphogenetic protein 9 (BMP9) has been shown to regulate processes such as angiogenesis, endothelial dysfunction, and tumorigenesis. However, the role of BMP9 in preeclampsia (PE) is unclear. The purpose of this study was to investigate the role and mechanism of BMP9 in PE.</p><p><strong>Methods: </strong>The effects of BMP9 on the viability, migration and invasion of HTR-8/Svneo cells were investigated by CCK-8 assay, wound healing assay and Transwell invasion assay. The effect of BMP9 on apoptosis of HTR-8/Svneo cells was detected by flow cytometry. Plasma levels of BMP9, SDF1 and CXCR4 were detected by ELISA kit. qRT-PCR and Western blot were used to detect the expression levels of each gene in the cells.</p><p><strong>Results: </strong>Overexpression of BMP9 promoted the proliferation and migration of trophoblast cells and inhibited apoptosis. Knockdown of BMP9 had the opposite effect. The levels of BMP9, SDF1 and CXCR4 in the plasma of PE patients were down-regulated, and BMP9 was positively correlated with the levels of SDF1 and CXCR4. BMP9 also significantly upregulated the mRNA and protein levels of SDF1 and CXCR4 in HTR-8/SVneo cells. Further mechanistic studies found that BMP9 promoted the migration and invasion of HTR-8/SVneo cells and inhibited apoptosis by activating the SDF1/CXCR4 pathway.</p><p><strong>Conclusion: </strong>We demonstrate for the first time that BMP9 promoted the migration and invasion of HTR-8/SVneo cells and inhibits apoptosis by activating the SDF1/CXCR4 pathway. This suggests that BMP9 may be a biomarker molecule for PE.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10405378/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10318932","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-07-25DOI: 10.1186/s12860-023-00486-1
Aysenur Gunaydin-Akyildiz, Rabia Sare Yanikoglu, Meltem Gulec, Gulbahar Ozge Alim-Toraman, Ebru Didem Kuran, Sezen Atasoy, Abdullah Olgun, Gulacti Topcu
Background: Emodin and aloe-emodin are two anthraquinones having positive effects in wound healing. However, their mechanism of action of wound healing is not fully understood. The MAP kinase family, which plays an active role in wound healing, is a well-characterized large family of serine/threonine kinases and regulates processes such as proliferation, oncogenesis, differentiation, and inflammation in the cell. The aim of this study is to comparatively elucidate the mechanisms of action of emodin and aloe-emodin, which are potential agents in wound healing.
Methods: The mechanism of the effects of emodin and aloe-emodin on cell viability and cell migration was examined using the human skin fibroblast (CCD-1079Sk) cell line. The gene expression levels of the MAP kinases (JNK, P38, ERK) in the skin fibroblast cells along with a molecular docking study analyzing their interaction potential were evaluated. Furthermore, the molecules' effects on the lifespan of Caenorhabditis elegans were studied.
Results: Emodin and aloe-emodin inhibited the ATP content of the cells in a concentration dependent manner and accelerated cell migration at the lower concentrations while inhibiting cell migration in the higher concentration treatment groups. The expressions of JNK and P38 were upregulated at the low concentrations and downregulated at the higher concentrations. The molecular docking studies of the molecules gave high docking scores indicating their interaction potential with JNK and P38. C. elegans lifespan under heat stress was observed longer after 75 µM emodin and was significantly reduced after 150 µM aloe-emodin treatment.
Conclusion: Aloe-emodin was found to be more potent on cell viability, cell migration, gene expression levels of the MAP kinases in healthy fibroblastic skin cells, and on the lifespan of C. elegans. This study reveals the functional effects and the biological factors that interact in the wound healing process of emodin and aloe-emodin, and give a possible treatment alternative to shorten the duration of wound care.
{"title":"Emodin and aloe-emodin, two potential molecules in regulating cell migration of skin cells through the MAP kinase pathway and affecting Caenorhabditis elegans thermotolerance.","authors":"Aysenur Gunaydin-Akyildiz, Rabia Sare Yanikoglu, Meltem Gulec, Gulbahar Ozge Alim-Toraman, Ebru Didem Kuran, Sezen Atasoy, Abdullah Olgun, Gulacti Topcu","doi":"10.1186/s12860-023-00486-1","DOIUrl":"https://doi.org/10.1186/s12860-023-00486-1","url":null,"abstract":"<p><strong>Background: </strong>Emodin and aloe-emodin are two anthraquinones having positive effects in wound healing. However, their mechanism of action of wound healing is not fully understood. The MAP kinase family, which plays an active role in wound healing, is a well-characterized large family of serine/threonine kinases and regulates processes such as proliferation, oncogenesis, differentiation, and inflammation in the cell. The aim of this study is to comparatively elucidate the mechanisms of action of emodin and aloe-emodin, which are potential agents in wound healing.</p><p><strong>Methods: </strong>The mechanism of the effects of emodin and aloe-emodin on cell viability and cell migration was examined using the human skin fibroblast (CCD-1079Sk) cell line. The gene expression levels of the MAP kinases (JNK, P38, ERK) in the skin fibroblast cells along with a molecular docking study analyzing their interaction potential were evaluated. Furthermore, the molecules' effects on the lifespan of Caenorhabditis elegans were studied.</p><p><strong>Results: </strong>Emodin and aloe-emodin inhibited the ATP content of the cells in a concentration dependent manner and accelerated cell migration at the lower concentrations while inhibiting cell migration in the higher concentration treatment groups. The expressions of JNK and P38 were upregulated at the low concentrations and downregulated at the higher concentrations. The molecular docking studies of the molecules gave high docking scores indicating their interaction potential with JNK and P38. C. elegans lifespan under heat stress was observed longer after 75 µM emodin and was significantly reduced after 150 µM aloe-emodin treatment.</p><p><strong>Conclusion: </strong>Aloe-emodin was found to be more potent on cell viability, cell migration, gene expression levels of the MAP kinases in healthy fibroblastic skin cells, and on the lifespan of C. elegans. This study reveals the functional effects and the biological factors that interact in the wound healing process of emodin and aloe-emodin, and give a possible treatment alternative to shorten the duration of wound care.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-07-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10367395/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9875467","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-07-20DOI: 10.1186/s12860-023-00485-2
Li Li, Shunying Wang, Wenming Wang
Background: Renal ischemia/reperfusion (I/R) injury is a major cause of acute kidney injury (AKI). Dysfunction of E74-like ETS transcription factor 4 (ELF4) leads to inflammation. This research intended to look into the function and mechanisms of ELF4 in I/R and oxygen-glucose deprivation/reperfusion (OGD/R) model.
Results: In I/R and OGD/R model, ELF4 expression was downregulated. ELF4 knockout aggravated I/R-induced kidney injury, oxidative stress (OS), endoplasmic reticulum stress (ERS), apoptosis, inflammation, and pyroptosis in mice. In HK-2 cells treated with OGD/R, suppression of ELF4 expression inhibited cell proliferation and promoted cell apoptosis, OS, ERS, inflammation, and pyroptosis. Moreover, ELF4 overexpression led to the opposite results.
Conclusion: ELF4 deficiency aggravated I/R induced AKI, which was involved in apoptosis, OS, ERS, inflammation, and pyroptosis. Targeting ELF4 may be a promising new therapeutic strategy for preventing inflammation after IR-AKI.
{"title":"Knockdown of ELF4 aggravates renal injury in ischemia/reperfusion mice through promotion of pyroptosis, inflammation, oxidative stress, and endoplasmic reticulum stress.","authors":"Li Li, Shunying Wang, Wenming Wang","doi":"10.1186/s12860-023-00485-2","DOIUrl":"10.1186/s12860-023-00485-2","url":null,"abstract":"<p><strong>Background: </strong>Renal ischemia/reperfusion (I/R) injury is a major cause of acute kidney injury (AKI). Dysfunction of E74-like ETS transcription factor 4 (ELF4) leads to inflammation. This research intended to look into the function and mechanisms of ELF4 in I/R and oxygen-glucose deprivation/reperfusion (OGD/R) model.</p><p><strong>Results: </strong>In I/R and OGD/R model, ELF4 expression was downregulated. ELF4 knockout aggravated I/R-induced kidney injury, oxidative stress (OS), endoplasmic reticulum stress (ERS), apoptosis, inflammation, and pyroptosis in mice. In HK-2 cells treated with OGD/R, suppression of ELF4 expression inhibited cell proliferation and promoted cell apoptosis, OS, ERS, inflammation, and pyroptosis. Moreover, ELF4 overexpression led to the opposite results.</p><p><strong>Conclusion: </strong>ELF4 deficiency aggravated I/R induced AKI, which was involved in apoptosis, OS, ERS, inflammation, and pyroptosis. Targeting ELF4 may be a promising new therapeutic strategy for preventing inflammation after IR-AKI.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-07-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10360327/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9911606","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-06-20DOI: 10.1186/s12860-023-00482-5
Amir Zareifard, Francis Beaudry, Kalidou Ndiaye
Janus kinase 3 (JAK3) is a member of the JAK family of tyrosine kinase proteins involved in cytokine receptor-mediated intracellular signal transduction through the JAK/STAT signaling pathway. JAK3 was previously shown as differentially expressed in granulosa cells (GC) of bovine pre-ovulatory follicles suggesting that JAK3 could modulate GC function and activation/inhibition of downstream targets. We used JANEX-1, a JAK3 inhibitor, and FSH treatments and analyzed proliferation markers, steroidogenic enzymes and phosphorylation of target proteins including STAT3, CDKN1B/p27Kip1 and MAPK8IP3/JIP3. Cultured GC were treated with or without FSH in the presence or not of JANEX-1. Expression of steroidogenic enzyme CYP11A1, but not CYP19A1, was upregulated in GC treated with FSH and both were significantly decreased when JAK3 was inhibited. Proliferation markers CCND2 and PCNA were reduced in JANEX-1-treated GC and upregulated by FSH. Western blots analyses showed that JANEX-1 treatment reduced pSTAT3 amounts while JAK3 overexpression increased pSTAT3. Similarly, FSH treatment increased pSTAT3 even in JANEX-1-treated GC. UHPLC-MS/MS analyses revealed phosphorylation of specific amino acid residues within JAK3 as well as CDKN1B and MAPK8IP3 suggesting possible activation or inhibition post-FSH or JANEX-1 treatments. We show that FSH activates JAK3 in GC, which could phosphorylate target proteins and likely modulate other signaling pathways involving CDKN1B and MAPK8IP3, therefore controlling GC proliferation and steroidogenic activity.
{"title":"Janus Kinase 3 phosphorylation and the JAK/STAT pathway are positively modulated by follicle-stimulating hormone (FSH) in bovine granulosa cells.","authors":"Amir Zareifard, Francis Beaudry, Kalidou Ndiaye","doi":"10.1186/s12860-023-00482-5","DOIUrl":"https://doi.org/10.1186/s12860-023-00482-5","url":null,"abstract":"<p><p>Janus kinase 3 (JAK3) is a member of the JAK family of tyrosine kinase proteins involved in cytokine receptor-mediated intracellular signal transduction through the JAK/STAT signaling pathway. JAK3 was previously shown as differentially expressed in granulosa cells (GC) of bovine pre-ovulatory follicles suggesting that JAK3 could modulate GC function and activation/inhibition of downstream targets. We used JANEX-1, a JAK3 inhibitor, and FSH treatments and analyzed proliferation markers, steroidogenic enzymes and phosphorylation of target proteins including STAT3, CDKN1B/p27<sup>Kip1</sup> and MAPK8IP3/JIP3. Cultured GC were treated with or without FSH in the presence or not of JANEX-1. Expression of steroidogenic enzyme CYP11A1, but not CYP19A1, was upregulated in GC treated with FSH and both were significantly decreased when JAK3 was inhibited. Proliferation markers CCND2 and PCNA were reduced in JANEX-1-treated GC and upregulated by FSH. Western blots analyses showed that JANEX-1 treatment reduced pSTAT3 amounts while JAK3 overexpression increased pSTAT3. Similarly, FSH treatment increased pSTAT3 even in JANEX-1-treated GC. UHPLC-MS/MS analyses revealed phosphorylation of specific amino acid residues within JAK3 as well as CDKN1B and MAPK8IP3 suggesting possible activation or inhibition post-FSH or JANEX-1 treatments. We show that FSH activates JAK3 in GC, which could phosphorylate target proteins and likely modulate other signaling pathways involving CDKN1B and MAPK8IP3, therefore controlling GC proliferation and steroidogenic activity.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-06-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10280845/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9708113","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-26DOI: 10.1186/s12860-023-00483-4
Andrea Bieder, Gayathri Chandrasekar, Arpit Wason, Steffen Erkelenz, Jay Gopalakrishnan, Juha Kere, Isabel Tapia-Páez
Background: DYX1C1 (DNAAF4) and DCDC2 are two of the most replicated dyslexia candidate genes in genetic studies. They both have demonstrated roles in neuronal migration, in cilia growth and function and they both are cytoskeletal interactors. In addition, they both have been characterized as ciliopathy genes. However, their exact molecular functions are still incompletely described. Based on these known roles, we asked whether DYX1C1 and DCDC2 interact on the genetic and the protein level.
Results: Here, we report the physical protein-protein interaction of DYX1C1 and DCDC2 as well as their respective interactions with the centrosomal protein CPAP (CENPJ) on exogenous and endogenous levels in different cell models including brain organoids. In addition, we show a synergistic genetic interaction between dyx1c1 and dcdc2b in zebrafish exacerbating the ciliary phenotype. Finally, we show a mutual effect on transcriptional regulation among DYX1C1 and DCDC2 in a cellular model.
Conclusions: In summary, we describe the physical and functional interaction between the two genes DYX1C1 and DCDC2. These results contribute to the growing understanding of the molecular roles of DYX1C1 and DCDC2 and set the stage for future functional studies.
{"title":"Genetic and protein interaction studies between the ciliary dyslexia candidate genes DYX1C1 and DCDC2.","authors":"Andrea Bieder, Gayathri Chandrasekar, Arpit Wason, Steffen Erkelenz, Jay Gopalakrishnan, Juha Kere, Isabel Tapia-Páez","doi":"10.1186/s12860-023-00483-4","DOIUrl":"https://doi.org/10.1186/s12860-023-00483-4","url":null,"abstract":"<p><strong>Background: </strong>DYX1C1 (DNAAF4) and DCDC2 are two of the most replicated dyslexia candidate genes in genetic studies. They both have demonstrated roles in neuronal migration, in cilia growth and function and they both are cytoskeletal interactors. In addition, they both have been characterized as ciliopathy genes. However, their exact molecular functions are still incompletely described. Based on these known roles, we asked whether DYX1C1 and DCDC2 interact on the genetic and the protein level.</p><p><strong>Results: </strong>Here, we report the physical protein-protein interaction of DYX1C1 and DCDC2 as well as their respective interactions with the centrosomal protein CPAP (CENPJ) on exogenous and endogenous levels in different cell models including brain organoids. In addition, we show a synergistic genetic interaction between dyx1c1 and dcdc2b in zebrafish exacerbating the ciliary phenotype. Finally, we show a mutual effect on transcriptional regulation among DYX1C1 and DCDC2 in a cellular model.</p><p><strong>Conclusions: </strong>In summary, we describe the physical and functional interaction between the two genes DYX1C1 and DCDC2. These results contribute to the growing understanding of the molecular roles of DYX1C1 and DCDC2 and set the stage for future functional studies.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-05-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10224228/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9564251","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-16DOI: 10.1186/s12860-023-00481-6
Kehuan Wang, Natalia Papadopoulos, Anahita Hamidi, Johan Lennartsson, Carl-Henrik Heldin
Background: The platelet-derived growth factor (PDGF) family of ligands exerts their cellular effects by binding to α- and β-tyrosine kinase receptors (PDGFRα and PDGFRβ, respectively). SUMOylation is an important posttranslational modification (PTM) which regulates protein stability, localization, activation and protein interactions. A mass spectrometry screen has demonstrated SUMOylation of PDGFRα. However, the functional role of SUMOylation of PDGFRα has remained unknown.
Results: In the present study, we validated that PDGFRα is SUMOylated on lysine residue 917 as was previously reported using a mass spectrometry approach. Mutation of lysine residue 917 to arginine (K917R) in PDGFRα substantially decreased SUMOylation, indicating that this amino acid residue is a major SUMOylation site. Whereas no difference in the stability of wild-type and mutant receptor was observed, the K917R mutant PDGFRα was less ubiquitinated than wild-type PDGFRα. The internalization and trafficking of the receptor to early and late endosomes were not affected by the mutation, neither was the localization of the PDGFRα to Golgi. However, the K917R mutant PDGFRα showed delayed activation of PLC-γ and enhanced activation of STAT3. Functional assays showed that the mutation of K917 of PDGFRα decreased cell proliferation in response to PDGF-BB stimulation.
Conclusions: SUMOylation of PDGFRα decreases ubiquitination of the receptor and affects ligand-induced signaling and cell proliferation.
{"title":"SUMOylation of PDGF receptor α affects signaling via PLCγ and STAT3, and cell proliferation.","authors":"Kehuan Wang, Natalia Papadopoulos, Anahita Hamidi, Johan Lennartsson, Carl-Henrik Heldin","doi":"10.1186/s12860-023-00481-6","DOIUrl":"https://doi.org/10.1186/s12860-023-00481-6","url":null,"abstract":"<p><strong>Background: </strong>The platelet-derived growth factor (PDGF) family of ligands exerts their cellular effects by binding to α- and β-tyrosine kinase receptors (PDGFRα and PDGFRβ, respectively). SUMOylation is an important posttranslational modification (PTM) which regulates protein stability, localization, activation and protein interactions. A mass spectrometry screen has demonstrated SUMOylation of PDGFRα. However, the functional role of SUMOylation of PDGFRα has remained unknown.</p><p><strong>Results: </strong>In the present study, we validated that PDGFRα is SUMOylated on lysine residue 917 as was previously reported using a mass spectrometry approach. Mutation of lysine residue 917 to arginine (K917R) in PDGFRα substantially decreased SUMOylation, indicating that this amino acid residue is a major SUMOylation site. Whereas no difference in the stability of wild-type and mutant receptor was observed, the K917R mutant PDGFRα was less ubiquitinated than wild-type PDGFRα. The internalization and trafficking of the receptor to early and late endosomes were not affected by the mutation, neither was the localization of the PDGFRα to Golgi. However, the K917R mutant PDGFRα showed delayed activation of PLC-γ and enhanced activation of STAT3. Functional assays showed that the mutation of K917 of PDGFRα decreased cell proliferation in response to PDGF-BB stimulation.</p><p><strong>Conclusions: </strong>SUMOylation of PDGFRα decreases ubiquitination of the receptor and affects ligand-induced signaling and cell proliferation.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-05-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10186711/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9540912","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-05-15DOI: 10.1186/s12860-023-00478-1
Aijia Cai, Paul Schneider, Zeng-Ming Zheng, Justus P Beier, Marcus Himmler, Dirk W Schubert, Volker Weisbach, Raymund E Horch, Andreas Arkudas
Background: For the purpose of skeletal muscle engineering, primary myoblasts (Mb) and adipogenic mesenchymal stem cells (ADSC) can be co-cultured and myogenically differentiated. Electrospun composite nanofiber scaffolds represent suitable matrices for tissue engineering of skeletal muscle, combining both biocompatibility and stability Although growth differentiation factor 11 (GDF11) has been proposed as a rejuvenating circulating factor, restoring skeletal muscle function in aging mice, some studies have also described a harming effect of GDF11. Therefore, the aim of the study was to analyze the effect of GDF11 on co-cultures of Mb and ADSC on poly-ε-caprolactone (PCL)-collagen I-polyethylene oxide (PEO)-nanofibers.
Results: Human Mb were co-cultured with ADSC two-dimensionally (2D) as monolayers or three-dimensionally (3D) on aligned PCL-collagen I-PEO-nanofibers. Differentiation media were either serum-free with or without GDF11, or serum containing as in a conventional differentiation medium. Cell viability was higher after conventional myogenic differentiation compared to serum-free and serum-free + GDF11 differentiation as was creatine kinase activity. Immunofluorescence staining showed myosine heavy chain expression in all groups after 28 days of differentiation without any clear evidence of more or less pronounced expression in either group. Gene expression of myosine heavy chain (MYH2) increased after serum-free + GDF11 stimulation compared to serum-free stimulation alone.
Conclusions: This is the first study analyzing the effect of GDF11 on myogenic differentiation of Mb and ADSC co-cultures under serum-free conditions. The results of this study show that PCL-collagen I-PEO-nanofibers represent a suitable matrix for 3D myogenic differentiation of Mb and ADSC. In this context, GDF11 seems to promote myogenic differentiation of Mb and ADSC co-cultures compared to serum-free differentiation without any evidence of a harming effect.
{"title":"Myogenic differentiation of human myoblasts and Mesenchymal stromal cells under GDF11 on NPoly-ɛ-caprolactone-collagen I-Polyethylene-nanofibers.","authors":"Aijia Cai, Paul Schneider, Zeng-Ming Zheng, Justus P Beier, Marcus Himmler, Dirk W Schubert, Volker Weisbach, Raymund E Horch, Andreas Arkudas","doi":"10.1186/s12860-023-00478-1","DOIUrl":"https://doi.org/10.1186/s12860-023-00478-1","url":null,"abstract":"<p><strong>Background: </strong>For the purpose of skeletal muscle engineering, primary myoblasts (Mb) and adipogenic mesenchymal stem cells (ADSC) can be co-cultured and myogenically differentiated. Electrospun composite nanofiber scaffolds represent suitable matrices for tissue engineering of skeletal muscle, combining both biocompatibility and stability Although growth differentiation factor 11 (GDF11) has been proposed as a rejuvenating circulating factor, restoring skeletal muscle function in aging mice, some studies have also described a harming effect of GDF11. Therefore, the aim of the study was to analyze the effect of GDF11 on co-cultures of Mb and ADSC on poly-ε-caprolactone (PCL)-collagen I-polyethylene oxide (PEO)-nanofibers.</p><p><strong>Results: </strong>Human Mb were co-cultured with ADSC two-dimensionally (2D) as monolayers or three-dimensionally (3D) on aligned PCL-collagen I-PEO-nanofibers. Differentiation media were either serum-free with or without GDF11, or serum containing as in a conventional differentiation medium. Cell viability was higher after conventional myogenic differentiation compared to serum-free and serum-free + GDF11 differentiation as was creatine kinase activity. Immunofluorescence staining showed myosine heavy chain expression in all groups after 28 days of differentiation without any clear evidence of more or less pronounced expression in either group. Gene expression of myosine heavy chain (MYH2) increased after serum-free + GDF11 stimulation compared to serum-free stimulation alone.</p><p><strong>Conclusions: </strong>This is the first study analyzing the effect of GDF11 on myogenic differentiation of Mb and ADSC co-cultures under serum-free conditions. The results of this study show that PCL-collagen I-PEO-nanofibers represent a suitable matrix for 3D myogenic differentiation of Mb and ADSC. In this context, GDF11 seems to promote myogenic differentiation of Mb and ADSC co-cultures compared to serum-free differentiation without any evidence of a harming effect.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-05-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10184409/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9497697","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Background: Cytochrome P450 4F2 (CYP4F2) enzyme is a member of the CYP4 family responsible for the metabolism of fatty acids, therapeutic drugs, and signaling molecules such as arachidonic acid, tocopherols, and vitamin K. Several reports have demonstrated that the missense variant CYP4F2*3 (V433M) causes decreased activity of CYP4F2 and inter-individual variations in warfarin dose in different ethnic groups. However, the molecular pathogenicity mechanism of missense V433M in CYP4F2 at the atomic level has not yet been completely elucidated.
Methods and results: In the current study, we evaluated the effect of the V433M substitution on CYP4F2 using 14 different bioinformatics tools. Further molecular dynamics (MD) simulations were performed to assess the impact of the V433M mutation on the CYP4F2 protein structure, stability, and dynamics. In addition, molecular docking was used to illustrate the effect of V433M on its interaction with vitamin K1. Based on our results, the CYP4F2*3 variant was a damaging amino acid substitution with a destabilizing nature. The simulation results showed that missense V433M affects the dynamics and stability of CYP4F2 by reducing its compactness and stability, which means that it tends to change the overall structural conformation and flexibility of CYP4F2. The docking results showed that the CYP4F2*3 variant decreased the binding affinity between vitamin K1 and CYP4F2, which reduced the activity of CYP4F2*3 compared to native CYP4F2.
Conclusions: This study determined the molecular pathogenicity mechanism of the CYP4F2*3 variant on the human CYP4F2 protein and provided new information for understanding the structure-function relationship of CYP4F2 and other CYP4 enzymes. These findings will aid in the development of effective drugs and treatment options.
{"title":"Computational analysis of missense variant CYP4F2*3 (V433M) in association with human CYP4F2 dysfunction: a functional and structural impact.","authors":"Mahvash Farajzadeh-Dehkordi, Ladan Mafakher, Fatemeh Samiee-Rad, Babak Rahmani","doi":"10.1186/s12860-023-00479-0","DOIUrl":"https://doi.org/10.1186/s12860-023-00479-0","url":null,"abstract":"<p><strong>Background: </strong>Cytochrome P450 4F2 (CYP4F2) enzyme is a member of the CYP4 family responsible for the metabolism of fatty acids, therapeutic drugs, and signaling molecules such as arachidonic acid, tocopherols, and vitamin K. Several reports have demonstrated that the missense variant CYP4F2*3 (V433M) causes decreased activity of CYP4F2 and inter-individual variations in warfarin dose in different ethnic groups. However, the molecular pathogenicity mechanism of missense V433M in CYP4F2 at the atomic level has not yet been completely elucidated.</p><p><strong>Methods and results: </strong>In the current study, we evaluated the effect of the V433M substitution on CYP4F2 using 14 different bioinformatics tools. Further molecular dynamics (MD) simulations were performed to assess the impact of the V433M mutation on the CYP4F2 protein structure, stability, and dynamics. In addition, molecular docking was used to illustrate the effect of V433M on its interaction with vitamin K1. Based on our results, the CYP4F2*3 variant was a damaging amino acid substitution with a destabilizing nature. The simulation results showed that missense V433M affects the dynamics and stability of CYP4F2 by reducing its compactness and stability, which means that it tends to change the overall structural conformation and flexibility of CYP4F2. The docking results showed that the CYP4F2*3 variant decreased the binding affinity between vitamin K1 and CYP4F2, which reduced the activity of CYP4F2*3 compared to native CYP4F2.</p><p><strong>Conclusions: </strong>This study determined the molecular pathogenicity mechanism of the CYP4F2*3 variant on the human CYP4F2 protein and provided new information for understanding the structure-function relationship of CYP4F2 and other CYP4 enzymes. These findings will aid in the development of effective drugs and treatment options.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-05-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10170697/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9505584","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-04-17DOI: 10.1186/s12860-023-00475-4
Huan Ting Ong, Cecilia M Prêle, Rodney J Dilley
Background: Hypoxic culture conditions have been used to study the impact of oxygen deprivation has on gene expression in a number of disease models. However, hypoxia response elements present in the promoter regions of some commonly used housekeeping genes, such as GAPDH and PGK1, can confound the relative gene expression analysis. Thus, there is ongoing debate as to which housekeeping gene is appropriate for studies investigating hypoxia-induced cell responses. Specifically, there is still contradicting information for which housekeeping genes are stable in hypoxia cultures of mesenchymal stem cells. In this study, candidate housekeeping genes curated from the literature were matched to RNAseq data of normoxic and hypoxic human adipose-derived stem cell cultures to determine if gene expression was modulated by hypoxia or not. Expression levels of selected candidates were used to calculate coefficient of variation. Then, accounting for the mean coefficient of variation, and normalised log twofold change, genes were ranked and shortlisted, before validating with qRT-PCR. Housekeeping gene suitability were then determined using GeNorm, NormFinder, BestKeeper, comparative[Formula: see text], RefFinder, and the Livak method.
Results: Gene expression levels of 78 candidate genes identified in the literature were analysed in the RNAseq dataset generated from hADSC cultured under Nx and Hx conditions. From the dataset, 15 candidates with coefficient of variation ≤ 0.15 were identified, where differential expression analysis results further shortlisted 8 genes with least variation in expression levels. The top 4 housekeeping gene candidates, ALAS1, RRP1, GUSB, and POLR2B, were chosen for qRT-PCR validation. Additionally, 18S, a ribosomal RNA commonly used as housekeeping gene but not detected in the RNAseq method, was added to the list of housekeeping gene candidates to validate. From qRT-PCR results, 18S and RRP1 were determined to be stably expressed in cells cultured under hypoxic conditions.
Conclusions: We have demonstrated that 18S and RRP1 are suitable housekeeping genes for use in hypoxia studies with human adipose-derived stem cell and should be used in combination. Additionally, these data shown that the commonly used GAPDH and PGK1 are not suitable housekeeping genes for investigations into the effect of hypoxia in human adipose-derived stem cell.
{"title":"Using RNA-seq to identify suitable housekeeping genes for hypoxia studies in human adipose-derived stem cells.","authors":"Huan Ting Ong, Cecilia M Prêle, Rodney J Dilley","doi":"10.1186/s12860-023-00475-4","DOIUrl":"https://doi.org/10.1186/s12860-023-00475-4","url":null,"abstract":"<p><strong>Background: </strong>Hypoxic culture conditions have been used to study the impact of oxygen deprivation has on gene expression in a number of disease models. However, hypoxia response elements present in the promoter regions of some commonly used housekeeping genes, such as GAPDH and PGK1, can confound the relative gene expression analysis. Thus, there is ongoing debate as to which housekeeping gene is appropriate for studies investigating hypoxia-induced cell responses. Specifically, there is still contradicting information for which housekeeping genes are stable in hypoxia cultures of mesenchymal stem cells. In this study, candidate housekeeping genes curated from the literature were matched to RNAseq data of normoxic and hypoxic human adipose-derived stem cell cultures to determine if gene expression was modulated by hypoxia or not. Expression levels of selected candidates were used to calculate coefficient of variation. Then, accounting for the mean coefficient of variation, and normalised log twofold change, genes were ranked and shortlisted, before validating with qRT-PCR. Housekeeping gene suitability were then determined using GeNorm, NormFinder, BestKeeper, comparative[Formula: see text], RefFinder, and the Livak method.</p><p><strong>Results: </strong>Gene expression levels of 78 candidate genes identified in the literature were analysed in the RNAseq dataset generated from hADSC cultured under Nx and Hx conditions. From the dataset, 15 candidates with coefficient of variation ≤ 0.15 were identified, where differential expression analysis results further shortlisted 8 genes with least variation in expression levels. The top 4 housekeeping gene candidates, ALAS1, RRP1, GUSB, and POLR2B, were chosen for qRT-PCR validation. Additionally, 18S, a ribosomal RNA commonly used as housekeeping gene but not detected in the RNAseq method, was added to the list of housekeeping gene candidates to validate. From qRT-PCR results, 18S and RRP1 were determined to be stably expressed in cells cultured under hypoxic conditions.</p><p><strong>Conclusions: </strong>We have demonstrated that 18S and RRP1 are suitable housekeeping genes for use in hypoxia studies with human adipose-derived stem cell and should be used in combination. Additionally, these data shown that the commonly used GAPDH and PGK1 are not suitable housekeeping genes for investigations into the effect of hypoxia in human adipose-derived stem cell.</p>","PeriodicalId":9099,"journal":{"name":"BMC Molecular and Cell Biology","volume":null,"pages":null},"PeriodicalIF":2.8,"publicationDate":"2023-04-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10108514/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9323366","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}