scholarly journals MeCP2-E1 isoform is a dynamically expressed, weakly DNA-bound protein with different protein and DNA interactions compared to MeCP2-E2

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Alexia Martínez de Paz ◽  
Leila Khajavi ◽  
Hélène Martin ◽  
Rafael Claveria-Gimeno ◽  
Susanne Tom Dieck ◽  
...  
Keyword(s):  
Rett Syndrome ◽  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Amino Acid Residues ◽  
Proteomics Data ◽  
Specific Expression ◽  
Interacting Protein ◽  
Cellular Processes ◽  
Specific Regulation ◽  
Dynamics Response

Abstract Background MeCP2—a chromatin-binding protein associated with Rett syndrome—has two main isoforms, MeCP2-E1 and MeCP2-E2, differing in a few N-terminal amino acid residues. Previous studies have shown brain region-specific expression of these isoforms which, in addition to their different cellular localization and differential expression during brain development, suggest that they may also have non-overlapping molecular mechanisms. However, differential functions of MeCP2-E1 and E2 remain largely unexplored. Results Here, we show that the N-terminal domains (NTD) of MeCP2-E1 and E2 modulate the ability of the methyl-binding domain (MBD) to interact with DNA as well as influencing the turn-over rates, binding dynamics, response to neuronal depolarization, and circadian oscillations of the two isoforms. Our proteomics data indicate that both isoforms exhibit unique interacting protein partners. Moreover, genome-wide analysis using ChIP-seq provide evidence for a shared as well as a specific regulation of different sets of genes. Conclusions Our study supports the idea that Rett syndrome might arise from simultaneous impairment of cellular processes involving non-overlapping functions of MECP2 isoforms. For instance, MeCP2-E1 mutations might impact stimuli-dependent chromatin regulation, while MeCP2-E2 mutations could result in aberrant ribosomal expression. Overall, our findings provide insight into the functional complexity of MeCP2 by dissecting differential aspects of its two isoforms.

scholarly journals MeCP2-E1 isoform is a dynamically expressed, weakly DNA-bound protein with different protein and DNA interactions compared to MeCP2-E2

2018 ◽  
Author(s):  
Alexia Martínez de Paz ◽  
Leila Khajavi ◽  
Hélène Martin ◽  
Rafael Claveria-Gimeno ◽  
Susanne tom Dieck ◽  
...  
Keyword(s):  
Amino Acid ◽  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Amino Acid Identity ◽  
Turnover Rates ◽  
Proteomics Data ◽  
Specific Expression ◽  
Interacting Protein ◽  
Specific Regulation ◽  
Dynamics Response

AbstractMeCP2 – a chromatin-binding protein associated with Rett syndrome – has two main isoforms, MeCP2-E1 and MeCP2-E2, with 96% amino acid identity differing in a few N-terminal amino acid residues. Previous studies have shown brain region-specific expression of these isoforms which, in addition to their different cellular localization and differential expression during brain development, suggest they may also have non-overlapping molecular mechanisms. However, differential functions of MeCP2-E1 and E2 remain largely unexplored. Here, we show that the N-terminal domains (NTD) of MeCP2-E1 and E2 modulate the ability of the methyl binding domain (MBD) to interact with DNA as well as influencing the turnover rates, binding dynamics, response to nuclear depolarization, and circadian oscillations of the two isoforms. Our proteomics data indicate that both isoforms exhibit unique interacting protein partners. Moreover, genome-wide analysis using ChIP-seq provide evidence for a shared as well as a specific regulation of different sets of genes. Our findings provide insight into the functional complexity of MeCP2 by dissecting differential aspects of its two isoforms.SignificanceWhether the two E1 and E2 isoforms of MeCP2 have different structural and/or functional implications has been highly controversial and is not well known. Here we show that the relatively short N-terminal sequence variation between the two isoforms impinges them with an important DNA binding difference. Moreover, MeCP2-E1 and E2 exhibit a different cellular dynamic behavior and have some distinctive interacting partners. In addition, while sharing genome occupancy they specifically bind to several distinctive genes.

Downregulation of cilia-localized Il-6Rα by 17β-estradiol in mouse and human fallopian tubes

2009 ◽  
Vol 297 (1) ◽  
pp. C140-C151 ◽  
Author(s):  
Ruijin Shao ◽  
Magdalena Nutu ◽  
Linda Karlsson-Lindahl ◽  
Anna Benrick ◽  
Birgitta Weijdegård ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Fallopian Tube ◽  
Knockout Mice ◽  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Fallopian Tubes ◽  
Female Reproduction ◽  
Specific Expression ◽  
17Β Estradiol ◽  
Specific Regulation

The action of interleukin-6 (IL-6) impacts female reproduction. Although IL-6 was recently shown to inhibit cilia activity in human fallopian tubes in vitro, the molecular mechanisms underlying IL-6 signaling to tubal function remain elusive. Here, we investigate the cellular localization, regulation, and possible function of two IL-6 receptors (IL-6Rα and gp130) in mouse and human fallopian tubes in vivo. We show that IL-6Rα is restricted to the cilia of epithelial cells in both mouse and human fallopian tubes. Exogenous 17β-estradiol (E2), but not progesterone (P4), causes a time-dependent decrease in IL-6Rα expression, which is blocked by the estrogen receptor (ER) antagonist ICI-182,780. Exposure of different ER-selective agonists propyl-(1H)-pyrazole-1,3,5-triyl-trisphenol or 2,3-bis-(4-hydroxyphenyl)-propionitrile demonstrated an ER subtype-specific regulation of IL-6Rα in mouse fallopian tubes. In contrast to IL-6Rα, gp130 was detected in tubal epithelial cells in mice but not in humans. In humans, gp130 was found in the muscle cells and was decreased in the periovulatory and luteal phases during the reproductive cycles, indicating a species-specific expression and regulation of gp130 in the fallopian tube. Expression of tubal IL-6Rα and gp130 in IL-6 knockout mice was found to be normal; however, E2 treatment increased IL-6Rα, but not gp130, in IL-6 knockout mice when compared with wild-type mice. Furthermore, expression levels of IL-6Rα, but not gp130, decreased in parallel with estrogenic accelerated oocyte-cumulus complex (OCC) transport in mouse fallopian tubes. Our findings open the posibility that cilia-specific IL-6Rα may play a role in the regulation of OCC transport and suggest an estrogen-regulatory pathway of IL-6Rα in the fallopian tube.

scholarly journals A Concerted Action of UBA5 C-Terminal Unstructured Regions Is Important for Transfer of Activated UFM1 to UFC1

2021 ◽  
Vol 22 (14) ◽  
pp. 7390
Author(s):  
Nicole Wesch ◽  
Frank Löhr ◽  
Natalia Rogova ◽  
Volker Dötsch ◽  
Vladimir V. Rogov
Keyword(s):  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Biochemical Characterization ◽  
Effective Interaction ◽  
Regulatory Region ◽  
Titration Calorimetry ◽  
Enzymatic Reactions ◽  
Cellular Processes ◽  
Mechanism Of Interaction

Ubiquitin fold modifier 1 (UFM1) is a member of the ubiquitin-like protein family. UFM1 undergoes a cascade of enzymatic reactions including activation by UBA5 (E1), transfer to UFC1 (E2) and selective conjugation to a number of target proteins via UFL1 (E3) enzymes. Despite the importance of ufmylation in a variety of cellular processes and its role in the pathogenicity of many human diseases, the molecular mechanisms of the ufmylation cascade remains unclear. In this study we focused on the biophysical and biochemical characterization of the interaction between UBA5 and UFC1. We explored the hypothesis that the unstructured C-terminal region of UBA5 serves as a regulatory region, controlling cellular localization of the elements of the ufmylation cascade and effective interaction between them. We found that the last 20 residues in UBA5 are pivotal for binding to UFC1 and can accelerate the transfer of UFM1 to UFC1. We solved the structure of a complex of UFC1 and a peptide spanning the last 20 residues of UBA5 by NMR spectroscopy. This structure in combination with additional NMR titration and isothermal titration calorimetry experiments revealed the mechanism of interaction and confirmed the importance of the C-terminal unstructured region in UBA5 for the ufmylation cascade.

scholarly journals Determining rewiring effects of alternatively spliced isoforms on protein-protein interactions using a computational approach

2018 ◽  
Author(s):  
Oleksandr Narykov ◽  
Nathan Johnson ◽  
Dmitry Korkin
Keyword(s):  
Protein Interactions ◽  
Molecular Mechanisms ◽  
Critical Role ◽  
Computational Approach ◽  
Superior Performance ◽  
Large Set ◽  
Protein Protein Interactions ◽  
Specific Expression ◽  
Alternatively Spliced ◽  
Specific Regulation

AbstractThe critical role of alternative splicing (AS) in cell functioning has recently become apparent, whether in studying tissue-or cell-specific regulation, or understanding molecular mechanisms governing a complex disorder. Studying the rewiring, or edgetic, effects of alternatively spliced isoforms on protein interactome can provide system-wide insights into these questions. Unfortunately, high-throughput experiments for such studies are expensive and time-consuming, hence the need to develop an in-silico approach. Here, we formulated the problem of characterization the edgetic effects of AS on protein-protein interactions (PPIs) as a binary classification problem and introduced a first computational approach to solve it. We first developed a supervised feature-based classifier that benefited from the traditional features describing a PPI, the problem-specific features that characterized the difference between the reference and alternative isoforms, and a novel domain interaction potential that allowed pinpointing the domains employed during a specific PPI. We then expanded this approach by including a large set of unlabeled interactomics data and developing a semi-supervised learning method. Our method called AS-IN (Alternatively Splicing INteraction prediction) Tool was compared with the state-of-the-art PPI prediction tools and showed a superior performance, achieving 0.92 in precision and recall. We demonstrated the utility of AS-IN Tool by applying it to the transcriptomic data obtained from the brain and liver tissues of a healthy mouse and western diet fed mouse that developed type two diabetes. We showed that the edgetic effects of differentially expressed transcripts associated with the disease condition are system-wide and unlikely to be detected by looking only at the gene-specific expression levels.

scholarly journals Regulation of Root Nutrient Transporters by CIPK23: ‘One Kinase to Rule Them All’

2020 ◽  
Author(s):  
Reyes Ródenas ◽  
Grégory Vert
Keyword(s):  
Protein Kinases ◽  
Molecular Mechanisms ◽  
Potassium Nitrate ◽  
Nutrient Transporters ◽  
Interacting Protein ◽  
Cellular Processes ◽  
Iron Metal ◽  
Nutrient Imbalances ◽  
Root Responses ◽  
Phosphate Groups

Abstract Protein kinases constitute essential regulatory components in the majority of cellular processes in eukaryotic cells. The CBL-INTERACTING PROTEIN KINASE (CIPK) family of plant protein kinases functions in calcium (Ca2+)-related signaling pathways and is therefore involved in the response to a wide variety of signals in plants. By covalently linking phosphate groups to their target proteins, CIPKs regulate the activity of downstream targets, their localization, their stability and their ability to interact with other proteins. In Arabidopsis, the CIPK23 kinase has emerged as a major hub driving root responses to diverse environmental stresses, including drought, salinity and nutrient imbalances, such as potassium, nitrate and iron deficiencies, as well as ammonium, magnesium and non-iron metal toxicities. This review will chiefly report on the prominent roles of CIPK23 in the regulation of plant nutrient transporters and on the underlying molecular mechanisms. We will also discuss the different scenarios explaining how a single promiscuous kinase, such as CIPK23, may convey specific responses to a myriad of signals.

scholarly journals SUMOylation Regulates TDP-43 Splicing Activity and Nucleocytoplasmic Distribution

2021 ◽  
Author(s):  
AnnaMaria Maraschi ◽  
Valentina Gumina ◽  
Jessica Dragotto ◽  
Claudia Colombrita ◽  
Miguel Mompeán ◽  
...  
Keyword(s):  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Exon Skipping ◽  
Nucleocytoplasmic Transport ◽  
Post Translational Modifications ◽  
Cellular Processes ◽  
Lysine Residues ◽  
Regulatory Effects ◽  
Nucleocytoplasmic Distribution

AbstractThe nuclear RNA-binding protein TDP-43 forms abnormal cytoplasmic aggregates in the brains of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) patients and several molecular mechanisms promoting TDP-43 cytoplasmic mislocalization and aggregation have been proposed, including defects in nucleocytoplasmic transport, stress granules (SG) disassembly and post-translational modifications (PTM). SUMOylation is a PTM which regulates a variety of cellular processes and, similarly to ubiquitination, targets lysine residues. To investigate the possible regulatory effects of SUMOylation on TDP-43 activity and trafficking, we first assessed that TDP-43 is SUMO-conjugated in the nuclear compartment both covalently and non-covalently in the RRM1 domain at the predicted lysine 136 and SUMO-interacting motif (SIM, 106–110 residues), respectively. By using the SUMO-mutant TDP-43 K136R protein, we demonstrated that SUMOylation modifies TDP-43 splicing activity, specifically exon skipping, and influences its sub-cellular localization and recruitment to SG after oxidative stress. When promoting deSUMOylation by SENP1 enzyme over-expression or by treatment with the cell-permeable SENP1 peptide TS-1, the cytoplasmic localization of TDP-43 increased, depending on its SUMOylation. Moreover, deSUMOylation by TS-1 peptide favoured the formation of small cytoplasmic aggregates of the C-terminal TDP-43 fragment p35, still containing the SUMO lysine target 136, but had no effect on the already formed p25 aggregates. Our data suggest that TDP-43 can be post-translationally modified by SUMOylation which may regulate its splicing function and trafficking, indicating a novel and druggable mechanism to explore as its dysregulation may lead to TDP-43 pathological aggregation in ALS and FTD.

scholarly journals SUMOylation Regulates TDP-43 Splicing Activity and Nucleocytoplasmic Distribution

2021 ◽  
Author(s):  
AnnaMaria Maraschi ◽  
Valentina Gumina ◽  
Jessica Dragotto ◽  
Claudia Colombrita ◽  
Miguel Monpeán ◽  
...  
Keyword(s):  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Exon Skipping ◽  
Nucleocytoplasmic Transport ◽  
Post Translational Modifications ◽  
Cellular Processes ◽  
Lysine Residues ◽  
Regulatory Effects ◽  
Nucleocytoplasmic Distribution

Abstract The nuclear RNA-binding protein TDP-43 forms abnormal cytoplasmic aggregates in the brains of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) patients and several molecular mechanisms promoting TDP-43 cytoplasmic mislocalization and aggregation have been proposed, including defects in nucleocytoplasmic transport, stress granules (SG) disassembly and post-translational modifications (PTM). SUMOylation is a PTM which regulates a variety of cellular processes and, similarly to ubiquitination, targets lysine residues. To investigate the possible regulatory effects of SUMOylation on TDP-43 activity and trafficking, we first assessed that TDP-43 is SUMO-conjugated in the nuclear compartment both covalently and non-covalently in the RRM1 domain at the predicted lysine 136 and SUMO-interacting motif (SIM, 106–110 residues), respectively. By using the SUMO-mutant TDP-43 K136R protein, we demonstrated that SUMOylation modifies TDP-43 splicing activity, specifically exon skipping, and influences its sub-cellular localization and recruitment to SG after oxidative stress. When promoting deSUMOylation by SENP1 enzyme over-expression or by treatment with the cell-permeable SENP1 peptide TS-1, the cytoplasmic localization of TDP-43 increased, depending on its SUMOylation. Moreover, deSUMOylation by TS-1 peptide favoured the formation of small cytoplasmic aggregates of the C-terminal TDP-43 fragment p35, still containing the SUMO lysine target 136, but had no effect on the already formed p25 aggregates. Our data suggest that TDP-43 can be post-translationally modified by SUMOylation which may regulate its splicing function and trafficking, indicating a novel and druggable mechanism to explore as its dysregulation may lead to TDP-43 pathological aggregation in ALS and FTD.

Targeting of Rab GTPases to cellular membranes

2005 ◽  
Vol 33 (4) ◽  
pp. 652-656 ◽  
Author(s):  
B.R. Ali ◽  
M.C. Seabra
Keyword(s):  
Membrane Trafficking ◽  
Cellular Localization ◽  
Molecular Mechanisms ◽  
Current Knowledge ◽  
Small Gtpases ◽  
Rab Proteins ◽  
Rab Gtpases ◽  
Cellular Membranes ◽  
Structural Determinants ◽  
Cellular Processes

Rab proteins are members of the superfamily of Ras-like small GTPases and are involved in several cellular processes relating to membrane trafficking and organelle mobility throughout the cell. Like other small GTPases, Rab proteins are initially synthesized as soluble proteins and for membrane attachment they require the addition of lipid moiety(ies) to specific residues of their polypeptide chain. Despite their well-documented roles in regulating cellular trafficking, Rab proteins own trafficking is still poorly understood. We still need to elucidate the molecular mechanisms of their recruitment to cellular membranes and the structural determinants for their specific cellular localization. Recent results indicate that Rab cellular targeting might be Rab-dependent, and this paper briefly reviews our current knowledge of this process.

Wilms tumor suppressor, Wt1, is a transcriptional activator of the erythropoietin gene

Blood ◽  
2006 ◽  
Vol 107 (11) ◽  
pp. 4282-4290 ◽  
Author(s):  
Christof Dame ◽  
Karin M. Kirschner ◽  
Katharina V. Bartz ◽  
Thomas Wallach ◽  
Christiane S. Hussels ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Molecular Mechanisms ◽  
Wilms Tumor ◽  
Transcriptional Activator ◽  
Mobility Shift ◽  
Specific Expression ◽  
Tissue Specific ◽  
Hek 293 ◽  
Mobility Shift Assay ◽  
Specific Regulation

AbstractMolecular mechanisms for the developmental stage and tissue-specific regulation of the erythropoietin (EPO) gene are poorly understood. Recent findings indicate a role of the Wilms tumor suppressor, Wt1, in the formation of the hematopoietic system. Herein, we tested the hypothesis that Wt1 is a transcriptional regulator of the EPO gene. Binding of the transcriptionally competent Wt1(–KTS) isoform to the minimal EPO promoter was demonstrated by electrophoretic mobility shift assay and chromatin immunoprecipitation. Under normoxia, EPO expression was significantly increased in HEK 293 and HepG2 cells with forced expression of Wt1(–KTS). A reporter construct harboring the 117-bp minimal human EPO promoter was activated up to 20-fold by transient cotransfection of Wt1(–KTS) in different cell lines. Mutation of the Wt1 binding site in the EPO promoter abrogated this stimulatory effect of the Wt1(–KTS) protein. Hepatic Epo mRNA expression was significantly reduced in embryonic mice with homozygous Wt1 deletion. Furthermore, Wt1 and EPO were colocalized in hepatocytes of the liver and in neuronal cells of the dorsal root ganglia in developing mice. Both proteins were also detected in Sertoli cells of the adult murine testis. In conclusion, we identified Wt1(–KTS) as a novel transcriptional activator for the tissue-specific expression of the EPO gene.

scholarly journals Cell-specific expression of ENACα gene by FOXA1 in the glucocorticoid receptor pathway

2020 ◽  
Vol 34 ◽  
pp. 205873842094619 ◽  
Author(s):  
Young Sun Chung ◽  
Hong Lan Jin ◽  
Kwang Won Jeong
Keyword(s):  
Gene Expression ◽  
Glucocorticoid Receptor ◽  
Molecular Mechanisms ◽  
Regulation Of Gene Expression ◽  
A549 Cells ◽  
Rna Seq ◽  
Cell Type ◽  
Specific Expression ◽  
Cell Type Specific ◽  
Specific Regulation

Introduction: The glucocorticoid receptor (GR) is one of the most widely studied ligand-dependent nuclear receptors. The combination of transcriptional regulatory factors required for the expression of individual genes targeted by GR varies across cell types; however, the mechanisms underlying this cell type–specific regulation of gene expression are not yet clear. Methods: Here, we investigated genes regulated by GR in two different cell lines, A549 and ARPE-19, and examined how gene expression varied according to the effect of pioneer factors using RNA-seq and RT-qPCR. Results: Our RNA-seq results identified 19 and 63 genes regulated by GR that are ARPE-19-specific and A549-specific, respectively, suggesting that GR induces the expression of different sets of genes in a cell type–specific manner. RT-qPCR confirmed that the epithelial sodium channel ( ENACα) gene is an ARPE-19 cell-specific GR target gene, whereas the FK506 binding protein 5 ( FKBP5) gene was A549 cell-specific. There was a significant decrease in ENACα expression in FOXA1-deficient ARPE-19 cells, suggesting that FOXA1 might function as a pioneer factor enabling the selective expression of ENACα in ARPE-19 cells but not in A549 cells. Conclusion: These findings indicate that ENACα expression in ARPE-19 cells is regulated by FOXA1 and provide insights into the molecular mechanisms of cell type–specific expression of GR-regulated genes.

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