scholarly journals Knockout of the Golgi stacking proteins GRASP55 and GRASP65 impairs Golgi structure and function

2017 ◽  
Vol 28 (21) ◽  
pp. 2833-2842 ◽  
Author(s):  
Michael E. Bekier ◽  
Leibin Wang ◽  
Jie Li ◽  
Haoran Huang ◽  
Danming Tang ◽  
...  
Keyword(s):  
Posttranslational Modifications ◽  
Protein Trafficking ◽  
Critical Role ◽  
Hek293 Cells ◽  
Double Knockout ◽  
Knock Out ◽  
Golgi Stack ◽  
Cellular Processes ◽  
Golgi Structure ◽  
And Function

Golgi reassembly stacking protein of 65 kDa (GRASP65) and Golgi reassembly stacking protein of 55 kDa (GRASP55) were originally identified as Golgi stacking proteins; however, subsequent GRASP knockdown experiments yielded inconsistent results with respect to the Golgi structure, indicating a limitation of RNAi-based depletion. In this study, we have applied the recently developed clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology to knock out GRASP55 and GRASP65, individually or in combination, in HeLa and HEK293 cells. We show that double knockout of GRASP proteins disperses the Golgi stack into single cisternae and tubulovesicular structures, accelerates protein trafficking, and impairs accurate glycosylation of proteins and lipids. These results demonstrate a critical role for GRASPs in maintaining the stacked structure of the Golgi, which is required for accurate posttranslational modifications in the Golgi. Additionally, the GRASP knockout cell lines developed in this study will be useful tools for studying the role of GRASP proteins in other important cellular processes.

scholarly journals Postprenylation CAAX Processing Is Required for Proper Localization of Ras but Not Rho GTPases

2005 ◽  
Vol 16 (4) ◽  
pp. 1606-1616 ◽  
Author(s):  
David Michaelson ◽  
Wasif Ali ◽  
Vi K. Chiu ◽  
Martin Bergo ◽  
Joseph Silletti ◽  
...  
Keyword(s):  
Posttranslational Modifications ◽  
Protein Trafficking ◽  
Rho Gtpases ◽  
Ras Proteins ◽  
Rho Proteins ◽  
Carboxyl Methylation ◽  
C Terminus ◽  
Individual Contributions ◽  
The Individual ◽  
And Function

The CAAX motif at the C terminus of most monomeric GTPases is required for membrane targeting because it signals for a series of three posttranslational modifications that include isoprenylation, endoproteolytic release of the C-terminal– AAX amino acids, and carboxyl methylation of the newly exposed isoprenylcysteine. The individual contributions of these modifications to protein trafficking and function are unknown. To address this issue, we performed a series of experiments with mouse embryonic fibroblasts (MEFs) lacking Rce1 (responsible for removal of the –AAX sequence) or Icmt (responsible for carboxyl methylation of the isoprenylcysteine). In MEFs lacking Rce1 or Icmt, farnesylated Ras proteins were mislocalized. In contrast, the intracellular localizations of geranylgeranylated Rho GTPases were not perturbed. Consistent with the latter finding, RhoGDI binding and actin remodeling were normal in Rce1- and Icmt-deficient cells. Swapping geranylgeranylation for farnesylation on Ras proteins or vice versa on Rho proteins reversed the differential sensitivities to Rce1 and Icmt deficiency. These results suggest that postprenylation CAAX processing is required for proper localization of farnesylated Ras but not geranygeranylated Rho proteins.

scholarly journals N-Cadherin Is Critical for the Survival of Germ Cells, the Formation of Steroidogenic Cells, and the Architecture of Developing Mouse Gonads

Cells ◽  
2019 ◽  
Vol 8 (12) ◽  
pp. 1610 ◽  
Author(s):  
Rafal P. Piprek ◽  
Michal Kolasa ◽  
Dagmara Podkowa ◽  
Malgorzata Kloc ◽  
Jacek Z. Kubiak
Keyword(s):  
Germ Cells ◽  
Hormonal Regulation ◽  
Critical Role ◽  
Somatic Cells ◽  
Gonad Development ◽  
Knock Out ◽  
Gonad Differentiation ◽  
Steroidogenic Cells ◽  
The Individual ◽  
And Function

Normal gonad development assures the fertility of the individual. The properly functioning gonads must contain a sufficient number of the viable germ cells, possess a correct architecture and tissue structure, and assure the proper hormonal regulation. This is achieved by the interplay between the germ cells and different types of somatic cells. N-cadherin coded by the Cdh2 gene plays a critical role in this interplay. To gain an insight into the role of N-cadherin in the development of mouse gonads, we used the Cre-loxP system to knock out N-cadherin separately in two cell lines: the SF1+ somatic cells and the OCT4+ germ cells. We observed that N-cadherin plays a key role in the survival of both female and male germ cells. However, the N-cadherin is not necessary for the differentiation of the Sertoli cells or the initiation of the formation of testis cords or ovigerous cords. In the later stages of gonad development, N-cadherin is important for the maintenance of testis cord structure and is required for the formation of steroidogenic cells. In the ovaries, N-cadherin is necessary for the formation of the ovarian follicles. These results indicate that N-cadherin plays a major role in gonad differentiation, structuralization, and function.

scholarly journals Hypoxia. 4. Hypoxia and ion channel function

2011 ◽  
Vol 300 (5) ◽  
pp. C951-C967 ◽  
Author(s):  
Larissa A. Shimoda ◽  
Jan Polak
Keyword(s):  
Ion Channels ◽  
Cell Function ◽  
Critical Role ◽  
Cardiac Contractility ◽  
Cell Types ◽  
Cellular Processes ◽  
Oxygen Sensitive ◽  
Sense And Respond ◽  
And Function ◽  
And Control

The ability to sense and respond to oxygen deprivation is required for survival; thus, understanding the mechanisms by which changes in oxygen are linked to cell viability and function is of great importance. Ion channels play a critical role in regulating cell function in a wide variety of biological processes, including neuronal transmission, control of ventilation, cardiac contractility, and control of vasomotor tone. Since the 1988 discovery of oxygen-sensitive potassium channels in chemoreceptors, the effect of hypoxia on an assortment of ion channels has been studied in an array of cell types. In this review, we describe the effects of both acute and sustained hypoxia (continuous and intermittent) on mammalian ion channels in several tissues, the mode of action, and their contribution to diverse cellular processes.

scholarly journals Innate Immune Signaling in the Pathogenesis of Necrotizing Enterocolitis

2013 ◽  
Vol 2013 ◽  
pp. 1-10 ◽  
Author(s):  
David J. Hackam ◽  
Amin Afrazi ◽  
Misty Good ◽  
Chhinder P. Sodhi
Keyword(s):  
Necrotizing Enterocolitis ◽  
Critical Role ◽  
Mucosal Injury ◽  
Term Infant ◽  
Tlr4 Signaling ◽  
Cellular Processes ◽  
Innate Immune Signaling ◽  
Proinflammatory Responses ◽  
And Function ◽  
Extracellular Environment

Necrotizing enterocolitis (NEC) is a challenging disease to treat, and caring for patients afflicted by it remains both frustrating and difficult. While NEC may develop quickly and without warning, it may also develop slowly, insidiously, and appear to take the caregiver by surprise. In seeking to understand the molecular and cellular processes that lead to NEC development, we have identified a critical role for the receptor for bacterial lipopolysaccharide (LPS) toll like receptor 4 (TLR4) in the pathogenesis of NEC, as its activation within the intestinal epithelium of the premature infant leads to mucosal injury and reduced epithelial repair. The expression and function of TLR4 were found to be particularly elevated within the intestinal mucosa of the premature as compared with the full-term infant, predisposing to NEC development. Importantly, factors within both the enterocyte itself, such as heat shock protein 70 (Hsp70), and in the extracellular environment, such as amniotic fluid, can curtail the extent of TLR4 signaling and reduce the propensity for NEC development. This review will highlight the critical TLR4-mediated steps that lead to NEC development, with a focus on the proinflammatory responses of TLR4 signaling that have such devastating consequences in the premature host.

scholarly journals MOG1 restores the expression and function of SCN5A-p.R104W through sec23a-mediated forward trafficking

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
L Luo ◽  
Y Wang ◽  
Y Du ◽  
C Dong ◽  
A Ma ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Sodium Current ◽  
Site Directed Mutagenesis ◽  
Hek293 Cells ◽  
Funding Source ◽  
Inherited Disease ◽  
Patch Clamp Recording ◽  
Knock Out ◽  
Cardiac Sodium Channel ◽  
And Function

Abstract Background Brugada syndrome (BrS) is an inherited disease which causes fatal arrhythmias and sudden cardiac death. Mutations in SCN5A gene, which encoding cardiac sodium channel (NaV1.5), are the most common genotype of BrS patients. Some SCN5A-related variants were reported to retain NaV1.5 in endoplasmic reticulum (ER) due to trafficking deficiency. MOG1 was previously reported to interact with NaV1.5 and increased sodium current (INa) through enhancing the trafficking. However, its molecular mechanisms are still unclear. Coat protein complex II (COPII) is responsible for the ER to Golgi transport. Sec23 forms the inner coat of COPII and participates in cargo proteins selection. Purpose To demonstrate that MOG1 rescues SCN5A-related variants by enhancing the forward trafficking through Sec23a-NaV1.5 interaction. Methods Site directed mutagenesis, immunofluorescence staining, biotinylation assay, Western blot analysis and whole-cell patch clamp recording were used. CRISPR/Cas9 was used to knock out Sec23a expression in HEK293 cells. Results We found that SCN5A-p.R104W was characterized as reduced NaV1.5 level and lack of INa. The variant SCN5A-p.R104W was mainly distributed in ER. MOG1 could rescue the total and surface expression of SCN5A-p.R104W but could not restore INa (Figure 1a). Considering that most patients are heterozygous, co-transfection of SCN5A-WT and SCN5A-p.R104W were obtained. We found MOG1 could increase both NaV1.5 level and INa of heterozygous expressed SCN5A-p.R104W. We further revealed an interaction between NaV1.5 and Sec23a by co-immunoprecipitation (Co-IP) assay. The interaction between NaV1.5 and Sec23a was increased by MOG1, which indicates that Sec23a participates in MOG1-mediated increase in NaV1.5 level (Figure 1b). Knockout of Sec23a reduced cell surface, but not total, NaV1.5 level (Figure 1c and 1d). Next, the Sec23a knockout HEK293 cells were co-transfected with SCN5A-p.R104W and pcDNA3 or MOG1. MOG1 could not increase SCN5A-p.R104W protein level in Sec23a knockout cells. Conclusion Our data demonstrated a novel mechanism that MOG1 restores the expression and function of SCN5A-p.R104W by enhancing its forward trafficking through Sec23a-NaV1.5 interaction. Figure 1 Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Natural Science Foundation of China

scholarly journals Posttranslational modifications in proteins: resources, tools and prediction methods

Database ◽  
2021 ◽  
Author(s):  
Shahin Ramazi ◽  
Javad Zahiri
Keyword(s):  
Computational Methods ◽  
Posttranslational Modifications ◽  
Side Chain ◽  
Amino Acid Side Chain ◽  
Cellular Processes ◽  
Online Databases ◽  
Different Types ◽  
Protein Functions ◽  
And Function ◽  
Molecular Regulatory Mechanisms

Abstract Posttranslational modifications (PTMs) refer to amino acid side chain modification in some proteins after their biosynthesis. There are more than 400 different types of PTMs affecting many aspects of protein functions. Such modifications happen as crucial molecular regulatory mechanisms to regulate diverse cellular processes. These processes have a significant impact on the structure and function of proteins. Disruption in PTMs can lead to the dysfunction of vital biological processes and hence to various diseases. High-throughput experimental methods for discovery of PTMs are very laborious and time-consuming. Therefore, there is an urgent need for computational methods and powerful tools to predict PTMs. There are vast amounts of PTMs data, which are publicly accessible through many online databases. In this survey, we comprehensively reviewed the major online databases and related tools. The current challenges of computational methods were reviewed in detail as well.

scholarly journals Microtubule Regulation and Function during Virus Infection

2017 ◽  
Vol 91 (16) ◽  
Author(s):  
Mojgan H. Naghavi ◽  
Derek Walsh
Keyword(s):  
Posttranslational Modifications ◽  
Intracellular Transport ◽  
Spatial Organization ◽  
Virus Particles ◽  
Cellular Processes ◽  
Wide Range ◽  
Associated Proteins ◽  
And Function ◽  
Induced Changes ◽  
Growth Pause

ABSTRACT Microtubules (MTs) form a rapidly adaptable network of filaments that radiate throughout the cell. These dynamic arrays facilitate a wide range of cellular processes, including the capture, transport, and spatial organization of cargos and organelles, as well as changes in cell shape, polarity, and motility. Nucleating from MT-organizing centers, including but by no means limited to the centrosome, MTs undergo rapid transitions through phases of growth, pause, and catastrophe, continuously exploring and adapting to the intracellular environment. Subsets of MTs can become stabilized in response to environmental cues, acquiring distinguishing posttranslational modifications and performing discrete functions as specialized tracks for cargo trafficking. The dynamic behavior and organization of the MT array is regulated by MT-associated proteins (MAPs), which include a subset of highly specialized plus-end-tracking proteins (+TIPs) that respond to signaling cues to alter MT behavior. As pathogenic cargos, viruses require MTs to transport to and from their intracellular sites of replication. While interactions with and functions for MT motor proteins are well characterized and extensively reviewed for many viruses, this review focuses on MT filaments themselves. Changes in the spatial organization and dynamics of the MT array, mediated by virus- or host-induced changes to MT regulatory proteins, not only play a central role in the intracellular transport of virus particles but also regulate a wider range of processes critical to the outcome of infection.

scholarly journals FADD in Cancer: Mechanisms of Altered Expression and Function, and Clinical Implications

Cancers ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 1462 ◽  
Author(s):  
José L Marín-Rubio ◽  
Laura Vela-Martín ◽  
José Fernández-Piqueras ◽  
María Villa-Morales
Keyword(s):  
Posttranslational Modifications ◽  
Cell Cycle Control ◽  
Regulation Of Gene Expression ◽  
Death Receptor ◽  
Clinical Implications ◽  
Altered Expression ◽  
Cellular Processes ◽  
Potential Biomarker ◽  
And Function ◽  
And Control

FADD was initially described as an adaptor molecule for death receptor-mediated apoptosis, but subsequently it has been implicated in nonapoptotic cellular processes such as proliferation and cell cycle control. During the last decade, FADD has been shown to play a pivotal role in most of the signalosome complexes, such as the necroptosome and the inflammasome. Interestingly, various mechanisms involved in regulating FADD functions have been identified, essentially posttranslational modifications and secretion. All these aspects have been thoroughly addressed in previous reviews. However, FADD implication in cancer is complex, due to pleiotropic effects. It has been reported either as anti- or protumorigenic, depending on the cell type. Regulation of FADD expression in cancer is a complex issue since both overexpression and downregulation have been reported, but the mechanisms underlying such alterations have not been fully unveiled. Posttranslational modifications also constitute a relevant mechanism controlling FADD levels and functions in tumor cells. In this review, we aim to provide detailed, updated information on alterations leading to changes in FADD expression and function in cancer. The participation of FADD in various biological processes is recapitulated, with a mention of interesting novel functions recently proposed for FADD, such as regulation of gene expression and control of metabolic pathways. Finally, we gather all the available evidence regarding the clinical implications of FADD alterations in cancer, especially as it has been proposed as a potential biomarker with prognostic value.

scholarly journals Regulation of Deubiquitinating Enzymes by Post-Translational Modifications

2020 ◽  
Vol 21 (11) ◽  
pp. 4028 ◽  
Author(s):  
Tanuza Das ◽  
Sang Chul Shin ◽  
Eun Joo Song ◽  
Eunice EunKyeong Kim
Keyword(s):  
Catalytic Activity ◽  
Posttranslational Modifications ◽  
Critical Role ◽  
Clear Understanding ◽  
Deubiquitinating Enzymes ◽  
Post Translational Modifications ◽  
Cellular Processes ◽  
Multiple Factors ◽  
Regulatory Processes ◽  
Partner Proteins

Ubiquitination and deubiquitination play a critical role in all aspects of cellular processes, and the enzymes involved are tightly regulated by multiple factors including posttranslational modifications like most other proteins. Dysfunction or misregulation of these enzymes could have dramatic physiological consequences, sometimes leading to diseases. Therefore, it is important to have a clear understanding of these regulatory processes. Here, we have reviewed the posttranslational modifications of deubiquitinating enzymes and their consequences on the catalytic activity, stability, abundance, localization, and interaction with the partner proteins.

Research progress in protein posttranslational modification site prediction

2018 ◽  
Vol 18 (4) ◽  
pp. 220-229 ◽  
Author(s):  
Wenying He ◽  
Leyi Wei ◽  
Quan Zou
Keyword(s):  
Posttranslational Modifications ◽  
Cell Biology ◽  
Research Progress ◽  
Future Research ◽  
Cellular Processes ◽  
Machine Learning Methods ◽  
Bioinformatics Tools ◽  
Glycosylation Sites ◽  
Almost All ◽  
And Function

AbstractPosttranslational modifications (PTMs) play an important role in regulating protein folding, activity and function and are involved in almost all cellular processes. Identification of PTMs of proteins is the basis for elucidating the mechanisms of cell biology and disease treatments. Compared with the laboriousness of equivalent experimental work, PTM prediction using various machine-learning methods can provide accurate, simple and rapid research solutions and generate valuable information for further laboratory studies. In this review, we manually curate most of the bioinformatics tools published since 2008. We also summarize the approaches for predicting ubiquitination sites and glycosylation sites. Moreover, we discuss the challenges of current PTM bioinformatics tools and look forward to future research possibilities.

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