Phosphorylation and Signal Transduction Pathways in Translational Control
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Molecular Dynamics and Evolutionary Aspects of the Transition from the Fully Grown Oocyte to Embryo
Downloaded from genesdev.cshlp.org on September 26, 2021 - Published by Cold Spring Harbor Laboratory Press Cracking the egg: molecular dynamics and evolutionary aspects of the transition from the fully grown oocyte to embryo Alexei V. Evsikov,1,5 Joel H. Graber,1 J. Michael Brockman,1,2 Aleš Hampl,3 Andrea E. Holbrook,1 Priyam Singh,1,2 John J. Eppig,1 Davor Solter,1,4 and Barbara B. Knowles1 1The Jackson Laboratory, Bar Harbor, Maine 04609, USA; 2 Bioinformatics Program, Boston University, Boston, Massachusetts 02215, USA; 3Masaryk University Brno and Institute of Experimental Medicine, 625 00 Brno, Czech Republic; 4Max Planck Institute of Immunobiology, 79108 Freiburg, Germany Fully grown oocytes (FGOs) contain all the necessary transcripts to activate molecular pathways underlying the oocyte-to-embryo transition (OET). To elucidate this critical period of development, an extensive survey of the FGO transcriptome was performed by analyzing 19,000 expressed sequence tags of the Mus musculus FGO cDNA library. Expression of 5400 genes and transposable elements is reported. For a majority of genes expressed in mouse FGOs, homologs transcribed in eggs of Xenopus laevis or Ciona intestinalis were found, pinpointing evolutionary conservation of most regulatory cascades underlying the OET in chordates. A large proportion of identified genes belongs to several gene families with oocyte-restricted expression, a likely result of lineage-specific genomic duplications. Gene loss by mutation and expression in female germline of retrotransposed genes specific to M. musculus is documented. These findings indicate rapid diversification of genes involved in female reproduction. Comparison of the FGO and two-cell embryo transcriptomes demarcated the processes important for oogenesis from those involved in OET and identified novel motifs in maternal mRNAs associated with transcript stability. -
A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated. -
4-6 Weeks Old Female C57BL/6 Mice Obtained from Jackson Labs Were Used for Cell Isolation
Methods Mice: 4-6 weeks old female C57BL/6 mice obtained from Jackson labs were used for cell isolation. Female Foxp3-IRES-GFP reporter mice (1), backcrossed to B6/C57 background for 10 generations, were used for the isolation of naïve CD4 and naïve CD8 cells for the RNAseq experiments. The mice were housed in pathogen-free animal facility in the La Jolla Institute for Allergy and Immunology and were used according to protocols approved by the Institutional Animal Care and use Committee. Preparation of cells: Subsets of thymocytes were isolated by cell sorting as previously described (2), after cell surface staining using CD4 (GK1.5), CD8 (53-6.7), CD3ε (145- 2C11), CD24 (M1/69) (all from Biolegend). DP cells: CD4+CD8 int/hi; CD4 SP cells: CD4CD3 hi, CD24 int/lo; CD8 SP cells: CD8 int/hi CD4 CD3 hi, CD24 int/lo (Fig S2). Peripheral subsets were isolated after pooling spleen and lymph nodes. T cells were enriched by negative isolation using Dynabeads (Dynabeads untouched mouse T cells, 11413D, Invitrogen). After surface staining for CD4 (GK1.5), CD8 (53-6.7), CD62L (MEL-14), CD25 (PC61) and CD44 (IM7), naïve CD4+CD62L hiCD25-CD44lo and naïve CD8+CD62L hiCD25-CD44lo were obtained by sorting (BD FACS Aria). Additionally, for the RNAseq experiments, CD4 and CD8 naïve cells were isolated by sorting T cells from the Foxp3- IRES-GFP mice: CD4+CD62LhiCD25–CD44lo GFP(FOXP3)– and CD8+CD62LhiCD25– CD44lo GFP(FOXP3)– (antibodies were from Biolegend). In some cases, naïve CD4 cells were cultured in vitro under Th1 or Th2 polarizing conditions (3, 4). -
Eef2k) Natural Product and Synthetic Small Molecule Inhibitors for Cancer Chemotherapy
International Journal of Molecular Sciences Review Progress in the Development of Eukaryotic Elongation Factor 2 Kinase (eEF2K) Natural Product and Synthetic Small Molecule Inhibitors for Cancer Chemotherapy Bin Zhang 1 , Jiamei Zou 1, Qiting Zhang 2, Ze Wang 1, Ning Wang 2,* , Shan He 1 , Yufen Zhao 2 and C. Benjamin Naman 1,* 1 Li Dak Sum Yip Yio Chin Kenneth Li Marine Biopharmaceutical Research Center, College of Food and Pharmaceutical Sciences, Ningbo University, Ningbo 315800, China; [email protected] (B.Z.); [email protected] (J.Z.); [email protected] (Z.W.); [email protected] (S.H.) 2 Institute of Drug Discovery Technology, Ningbo University, Ningbo 315211, China; [email protected] (Q.Z.); [email protected] (Y.Z.) * Correspondence: [email protected] (N.W.); [email protected] (C.B.N.) Abstract: Eukaryotic elongation factor 2 kinase (eEF2K or Ca2+/calmodulin-dependent protein kinase, CAMKIII) is a new member of an atypical α-kinase family different from conventional protein kinases that is now considered as a potential target for the treatment of cancer. This protein regulates the phosphorylation of eukaryotic elongation factor 2 (eEF2) to restrain activity and inhibit the elongation stage of protein synthesis. Mounting evidence shows that eEF2K regulates the cell cycle, autophagy, apoptosis, angiogenesis, invasion, and metastasis in several types of cancers. The Citation: Zhang, B.; Zou, J.; Zhang, expression of eEF2K promotes survival of cancer cells, and the level of this protein is increased in Q.; Wang, Z.; Wang, N.; He, S.; Zhao, many cancer cells to adapt them to the microenvironment conditions including hypoxia, nutrient Y.; Naman, C.B. -
Mouse Eif4e3 Knockout Project (CRISPR/Cas9)
https://www.alphaknockout.com Mouse Eif4e3 Knockout Project (CRISPR/Cas9) Objective: To create a Eif4e3 knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering. Strategy summary: The Eif4e3 gene (NCBI Reference Sequence: NM_025829 ; Ensembl: ENSMUSG00000093661 ) is located on Mouse chromosome 6. 7 exons are identified, with the ATG start codon in exon 1 and the TAA stop codon in exon 7 (Transcript: ENSMUST00000032151). Exon 3~5 will be selected as target site. Cas9 and gRNA will be co-injected into fertilized eggs for KO Mouse production. The pups will be genotyped by PCR followed by sequencing analysis. Note: Mice homozygous for a knock-out allele exhibit decreased bone trabecula number. Exon 3 starts from about 32.05% of the coding region. Exon 3~5 covers 35.91% of the coding region. The size of effective KO region: ~4602 bp. The KO region does not have any other known gene. Page 1 of 8 https://www.alphaknockout.com Overview of the Targeting Strategy Wildtype allele 5' gRNA region gRNA region 3' 1 3 4 5 7 Legends Exon of mouse Eif4e3 Knockout region Page 2 of 8 https://www.alphaknockout.com Overview of the Dot Plot (up) Window size: 15 bp Forward Reverse Complement Sequence 12 Note: The 2000 bp section upstream of Exon 3 is aligned with itself to determine if there are tandem repeats. No significant tandem repeat is found in the dot plot matrix. So this region is suitable for PCR screening or sequencing analysis. Overview of the Dot Plot (down) Window size: 15 bp Forward Reverse Complement Sequence 12 Note: The 2000 bp section downstream of Exon 5 is aligned with itself to determine if there are tandem repeats. -
Supplementary Information
SUPPLEMENTARY INFORMATION Myeloperoxidase-derived 2-chlorohexadecanal is generated in mouse heart during endotoxemia and induces modification of distinct cardiomyocyte protein subsets in vitro Jürgen Prasch, Eva Bernhart, Helga Reicher, Manfred Kollroser, Gerald N. Rechberger, Chintan N. Koyani, Christopher Trummer, Lavinia Rech, Peter P. Rainer, Astrid Hammer, Ernst Malle, Wolfgang Sattler Table S1: Biological process gene ontology (GO) enrichment analysis. #term ID term description observed background false discovery matching proteins in network (labels) gene count gene count rate GO:0006457 protein folding 10 153 5.21e-09 Cct3,Cct5,Cct8,Fkbp4,Hsp90aa1,Hsp a1l,Hspb1,Pdia3,Pdia6,Tcp1 GO:0007339 binding of sperm to 6 36 4.02e-07 Aldoa,Cct3,Cct5,Cct8,Hspa1l,Tcp1 zona pellucida GO:0061077 chaperone-mediated 6 60 2.67e-06 Cct3,Cct5,Cct8,Fkbp4,Hspb1,Tcp1 protein folding GO:0017144 drug metabolic process 11 494 4.06e-06 Aldh2,Aldoa,Eno1,Gapdh,Hsp90aa1,I dh3a,Ldha,Ndufs2,Pgam1,Phgdh,Uq crc1 GO:2000573 positive regulation of 6 69 4.16e-06 Cct3,Cct5,Cct8,Ddx39b,Hsp90aa1,Tc DNA biosynthetic p1 process GO:0009987 cellular process 47 12459 4.22e-06 Alad,Alb,Aldh2,Aldoa,Cct3,Cct5,Cct8, Dctn2,Ddx39,Ddx39b,Des,Eef1g,Eef 2,Eif3f,Eif4a2,Eno1,Fdps,Fkbp4,Gap dh,Hnrnpl,Hsp90aa1,Hspa1l,Hspb1,I dh3a,Ldha,Lmna,Lyz1,Ndufs2,Pcna, Pdia3,Pdia6,Pgam1,Phgdh,Prph,Psm d13,Rpsa,Ruvbl2,Tcp1,Tuba3b,Tubal 3,Tubb3,Tubb6,Uap1l1,Uqcrc1,Uqcrc 2,Vim,Ywhab GO:1904851 positive regulation of 4 10 4.22e-06 Cct3,Cct5,Cct8,Tcp1 establishment of protein localization to telomere GO:0046031 -
Molecular Pharmacology of Cancer Therapy in Human Colorectal Cancer by Gene Expression Profiling1,2
[CANCER RESEARCH 63, 6855–6863, October 15, 2003] Molecular Pharmacology of Cancer Therapy in Human Colorectal Cancer by Gene Expression Profiling1,2 Paul A. Clarke,3 Mark L. George, Sandra Easdale, David Cunningham, R. Ian Swift, Mark E. Hill, Diana M. Tait, and Paul Workman Cancer Research UK Centre for Cancer Therapeutics, Institute of Cancer Research, Sutton, Surrey SM2 5NG [P. A. C., M. L. G., S. E., P. W.]; Department of Gastrointestinal Oncology, Royal Marsden Hospital, Sutton, Surrey [D. C., M. E. H., D. M. T.]; and Department of Surgery, Mayday Hospital, Croydon, Surrey [M. L. G., R. I. S.], United Kingdom ABSTRACT ment with a single dose of MMC4 and during a continuous infusion of 5FU. In this study, we report for the first time gene expression Global gene expression profiling has potential for elucidating the com- profiling in cancer patients before, and critically, during the period of plex cellular effects and mechanisms of action of novel targeted anticancer exposure to chemotherapy. We have demonstrated that the approach agents or existing chemotherapeutics for which the precise molecular is feasible, and we have detected a novel molecular response that mechanism of action may be unclear. In this study, decreased expression would not have been predicted from in vitro studies and that would of genes required for RNA and protein synthesis, and for metabolism were have otherwise been missed by conventional approaches. The results detected in rectal cancer biopsies taken from patients during a 5-fluorou- also suggest a possible new therapeutic approach. Overall our obser- racil infusion. Our observations demonstrate that this approach is feasible and can detect responses that may have otherwise been missed by con- vations suggest that gene expression profiling in response to treatment ventional methods. -
RNA-Seq Transcriptome Reveals Different Molecular Responses
Zhao et al. BMC Genomics (2020) 21:475 https://doi.org/10.1186/s12864-020-06885-4 RESEARCH ARTICLE Open Access RNA-Seq transcriptome reveals different molecular responses during human and mouse oocyte maturation and fertilization Zheng-Hui Zhao1,2, Tie-Gang Meng1,3, Ang Li1, Heide Schatten4, Zhen-Bo Wang1,2* and Qing-Yuan Sun1,3* Abstract Background: Female infertility is a worldwide concern and the etiology of infertility has not been thoroughly demonstrated. Although the mouse is a good model system to perform functional studies, the differences between mouse and human also need to be considered. The objective of this study is to elucidate the different molecular mechanisms underlying oocyte maturation and fertilization between human and mouse. Results: A comparative transcriptome analysis was performed to identify the differentially expressed genes and associated biological processes between human and mouse oocytes. In total, 8513 common genes, as well as 15, 165 and 6126 uniquely expressed genes were detected in human and mouse MII oocytes, respectively. Additionally, the ratios of non-homologous genes in human and mouse MII oocytes were 37 and 8%, respectively. Functional categorization analysis of the human MII non-homologous genes revealed that cAMP-mediated signaling, sister chromatid cohesin, and cell recognition were the major enriched biological processes. Interestingly, we couldn’t detect any GO categories in mouse non-homologous genes. Conclusions: This study demonstrates that human and mouse oocytes exhibit significant differences in gene expression profiles during oocyte maturation, which probably deciphers the differential molecular responses to oocyte maturation and fertilization. The significant differences between human and mouse oocytes limit the generalizations from mouse to human oocyte maturation. -
Drosophila and Human Transcriptomic Data Mining Provides Evidence for Therapeutic
Drosophila and human transcriptomic data mining provides evidence for therapeutic mechanism of pentylenetetrazole in Down syndrome Author Abhay Sharma Institute of Genomics and Integrative Biology Council of Scientific and Industrial Research Delhi University Campus, Mall Road Delhi 110007, India Tel: +91-11-27666156, Fax: +91-11-27662407 Email: [email protected] Nature Precedings : hdl:10101/npre.2010.4330.1 Posted 5 Apr 2010 Running head: Pentylenetetrazole mechanism in Down syndrome 1 Abstract Pentylenetetrazole (PTZ) has recently been found to ameliorate cognitive impairment in rodent models of Down syndrome (DS). The mechanism underlying PTZ’s therapeutic effect is however not clear. Microarray profiling has previously reported differential expression of genes in DS. No mammalian transcriptomic data on PTZ treatment however exists. Nevertheless, a Drosophila model inspired by rodent models of PTZ induced kindling plasticity has recently been described. Microarray profiling has shown PTZ’s downregulatory effect on gene expression in fly heads. In a comparative transcriptomics approach, I have analyzed the available microarray data in order to identify potential mechanism of PTZ action in DS. I find that transcriptomic correlates of chronic PTZ in Drosophila and DS counteract each other. A significant enrichment is observed between PTZ downregulated and DS upregulated genes, and a significant depletion between PTZ downregulated and DS dowwnregulated genes. Further, the common genes in PTZ Nature Precedings : hdl:10101/npre.2010.4330.1 Posted 5 Apr 2010 downregulated and DS upregulated sets show enrichment for MAP kinase pathway. My analysis suggests that downregulation of MAP kinase pathway may mediate therapeutic effect of PTZ in DS. Existing evidence implicating MAP kinase pathway in DS supports this observation. -
Relevance of Translation Initiation in Diffuse Glioma Biology and Its
cells Review Relevance of Translation Initiation in Diffuse Glioma Biology and its Therapeutic Potential Digregorio Marina 1, Lombard Arnaud 1,2, Lumapat Paul Noel 1, Scholtes Felix 1,2, Rogister Bernard 1,3 and Coppieters Natacha 1,* 1 Laboratory of Nervous System Disorders and Therapy, GIGA-Neurosciences Research Centre, University of Liège, 4000 Liège, Belgium; [email protected] (D.M.); [email protected] (L.A.); [email protected] (L.P.N.); [email protected] (S.F.); [email protected] (R.B.) 2 Department of Neurosurgery, CHU of Liège, 4000 Liège, Belgium 3 Department of Neurology, CHU of Liège, 4000 Liège, Belgium * Correspondence: [email protected] Received: 18 October 2019; Accepted: 26 November 2019; Published: 29 November 2019 Abstract: Cancer cells are continually exposed to environmental stressors forcing them to adapt their protein production to survive. The translational machinery can be recruited by malignant cells to synthesize proteins required to promote their survival, even in times of high physiological and pathological stress. This phenomenon has been described in several cancers including in gliomas. Abnormal regulation of translation has encouraged the development of new therapeutics targeting the protein synthesis pathway. This approach could be meaningful for glioma given the fact that the median survival following diagnosis of the highest grade of glioma remains short despite current therapy. The identification of new targets for the development of novel therapeutics is therefore needed in order to improve this devastating overall survival rate. This review discusses current literature on translation in gliomas with a focus on the initiation step covering both the cap-dependent and cap-independent modes of initiation. -
Eukaryotic Initiation Factor 4A2 Promotes Experimental Metastasis
Chen et al. Journal of Experimental & Clinical Cancer Research (2019) 38:196 https://doi.org/10.1186/s13046-019-1178-z RESEARCH Open Access Eukaryotic initiation factor 4A2 promotes experimental metastasis and oxaliplatin resistance in colorectal cancer Zhan-Hong Chen1,2†, Jing-Jing Qi1†, Qi-Nian Wu3†, Jia-Huan Lu1†, Ze-Xian Liu1, Yun Wang1, Pei-Shan Hu1, Ting Li1, Jin-Fei Lin1, Xiang-Yuan Wu2, Lei Miao1, Zhao-Lei Zeng1, Dan Xie3, Huai-Qiang Ju1, Rui-Hua Xu1* and Feng Wang1* Abstract Background: Deregulation of protein translation control is a hallmark of cancers. Eukaryotic initiation factor 4A2 (EIF4A2) is required for mRNA binding to ribosome and plays an important role in translation initiation. However, little is known about its functions in colorectal cancer (CRC). Methods: Analysis of CRC transcriptome data from TCGA identified that EIF4A2 was associated with poor prognosis. Immunohistochemistry study of EIF4A2 was carried out in 297 paired colorectal tumor and adjacent normal tissue samples. In vitro and in vivo cell-biological assays were performed to study the biological functions of EIF4A2 on experimental metastasis and sensitivity to oxaliplatin treatment. Bioinformatic prediction, chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assay were carried out to unveil the transcription factor of EIF4A2 regulation. Results: EIF4A2 Expression is significantly higher in colorectal tumors. Multivariate analysis suggests EIF4A2 as an independent predictor of overall, disease-free and progression-free survival. Dysfunction of EIF4A2 by genetic knock-down or small-molecule inhibitor silvestrol dramatically inhibited CRC invasion and migration, sphere formation and enhanced sensitivity to oxaliplatin treatment in vitro and in vivo. -
Regulation of Host Translational Machinery by African Swine Fever Virus
Regulation of Host Translational Machinery by African Swine Fever Virus Alfredo Castello´ ¤, Ana Quintas, Elena G. Sa´nchez, Prado Sabina, Marisa Nogal, Luis Carrasco, Yolanda Revilla* Centro de Biologı´a Molecular Severo Ochoa, CSIC-UAM, Universidad Auto´noma de Madrid, Madrid, Spain Abstract African swine fever virus (ASFV), like other complex DNA viruses, deploys a variety of strategies to evade the host’s defence systems, such as inflammatory and immune responses and cell death. Here, we analyse the modifications in the translational machinery induced by ASFV. During ASFV infection, eIF4G and eIF4E are phosphorylated (Ser1108 and Ser209, respectively), whereas 4E-BP1 is hyperphosphorylated at early times post infection and hypophosphorylated after 18 h. Indeed, a potent increase in eIF4F assembly is observed in ASFV-infected cells, which is prevented by rapamycin treatment. Phosphorylation of eIF4E, eIF4GI and 4E-BP1 is important to enhance viral protein production, but is not essential for ASFV infection as observed in rapamycin- or CGP57380-treated cells. Nevertheless, eIF4F components are indispensable for ASFV protein synthesis and virus spread, since eIF4E or eIF4G depletion in COS-7 or Vero cells strongly prevents accumulation of viral proteins and decreases virus titre. In addition, eIF4F is not only activated but also redistributed within the viral factories at early times of infection, while eIF4G and eIF4E are surrounding these areas at late times. In fact, other components of translational machinery such as eIF2a, eIF3b, eIF4E, eEF2 and ribosomal P protein are enriched in areas surrounding ASFV factories. Notably, the mitochondrial network is polarized in ASFV-infected cells co-localizing with ribosomes.