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Laboratory Investigation; a Journal of... Jul 2019In multiple neurodegenerative diseases, including Alzheimer's disease (AD), a prominent pathological feature is the aberrant aggregation and inclusion formation of the... (Review)
Review
In multiple neurodegenerative diseases, including Alzheimer's disease (AD), a prominent pathological feature is the aberrant aggregation and inclusion formation of the microtubule-associated protein tau. Because of the pathological association, these disorders are often referred to as tauopathies. Mutations in the MAPT gene that encodes tau can cause frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), providing the clearest evidence that tauopathy plays a causal role in neurodegeneration. However, large gaps in our knowledge remain regarding how various FTDP-17-linked tau mutations promote tau aggregation and neurodegeneration, and, more generally, how the tauopathy is linked to neurodegeneration. Herein, we review what is known about how FTDP-17-linked pathogenic MAPT mutations cause disease, with a major focus on the prion-like properties of wild-type and mutant tau proteins. The hypothesized mechanisms by which mutations in the MAPT gene promote tauopathy are quite varied and may not provide definitive insights into how tauopathy arises in the absence of mutation. Further, differences in the ability of tau and mutant tau proteins to support prion-like propagation in various model systems raise questions about the generalizability of this mechanism in various tauopathies. Notably, understanding the mechanisms of tauopathy induction and spread and tau-induced neurodegeneration has important implications for tau-targeting therapeutics.
Topics: Animals; Humans; Microtubules; Mutation; Parkinsonian Disorders; Protein Aggregation, Pathological; Protein Processing, Post-Translational; Protein Splicing; Tauopathies; tau Proteins
PubMed: 30742061
DOI: 10.1038/s41374-019-0197-x -
Molecular Cancer Jan 2021De novo and acquired resistance, which are mainly mediated by genetic alterations, are barriers to effective routine chemotherapy. However, the mechanisms underlying...
De novo and acquired resistance, which are mainly mediated by genetic alterations, are barriers to effective routine chemotherapy. However, the mechanisms underlying gastric cancer (GC) resistance to chemotherapy are still unclear. We showed that the long noncoding RNA CRNDE was related to the chemosensitivity of GC in clinical samples and a PDX model. CRNDE was decreased and inhibited autophagy flux in chemoresistant GC cells. CRNDE directly bound to splicing protein SRSF6 to reduce its protein stability and thus regulate alternative splicing (AS) events. We determined that SRSF6 regulated the PICALM exon 14 skip splice variant and triggered a significant S-to-L isoform switch, which contributed to the expression of the long isoform of PICALM (encoding PICALML). Collectively, our findings reveal the key role of CRNDE in autophagy regulation, highlighting the significance of CRNDE as a potential prognostic marker and therapeutic target against chemoresistance in GC.
Topics: Alternative Splicing; Autophagy; Cell Line, Tumor; Drug Resistance, Neoplasm; Fluorouracil; Humans; Monomeric Clathrin Assembly Proteins; Oxaliplatin; Phosphoproteins; Proteasome Endopeptidase Complex; Proteolysis; RNA, Long Noncoding; Serine-Arginine Splicing Factors; Stomach Neoplasms; Ubiquitination
PubMed: 33397371
DOI: 10.1186/s12943-020-01299-y -
Journal of Cystic Fibrosis : Official... May 2008It is often challenging for the clinician interested in cystic fibrosis (CF) to interpret molecular genetic results, and to integrate them in the diagnostic process. The... (Review)
Review
It is often challenging for the clinician interested in cystic fibrosis (CF) to interpret molecular genetic results, and to integrate them in the diagnostic process. The limitations of genotyping technology, the choice of mutations to be tested, and the clinical context in which the test is administered can all influence how genetic information is interpreted. This paper describes the conclusions of a consensus conference to address the use and interpretation of CF mutation analysis in clinical settings. Although the diagnosis of CF is usually straightforward, care needs to be exercised in the use and interpretation of genetic tests: genotype information is not the final arbiter of a clinical diagnosis of CF or CF transmembrane conductance regulator (CFTR) protein related disorders. The diagnosis of these conditions is primarily based on the clinical presentation, and is supported by evaluation of CFTR function (sweat testing, nasal potential difference) and genetic analysis. None of these features are sufficient on their own to make a diagnosis of CF or CFTR-related disorders. Broad genotype/phenotype associations are useful in epidemiological studies, but CFTR genotype does not accurately predict individual outcome. The use of CFTR genotype for prediction of prognosis in people with CF at the time of their diagnosis is not recommended. The importance of communication between clinicians and medical genetic laboratories is emphasized. The results of testing and their implications should be reported in a manner understandable to the clinicians caring for CF patients.
Topics: Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; DNA Mutational Analysis; Humans; Nutritional Status; Polymorphism, Genetic; Prognosis; Protein Splicing; Quality Control; Respiratory Function Tests; Terminology as Topic
PubMed: 18456578
DOI: 10.1016/j.jcf.2008.03.009 -
Nature Communications Oct 2019Selectable markers are widely used in transgenesis and genome editing for selecting engineered cells with a desired genotype but the variety of markers is limited. Here...
Selectable markers are widely used in transgenesis and genome editing for selecting engineered cells with a desired genotype but the variety of markers is limited. Here we present split selectable markers that each allow for selection of multiple "unlinked" transgenes in the context of lentivirus-mediated transgenesis as well as CRISPR-Cas-mediated knock-ins. Split marker gene segments fused to protein splicing elements called "inteins" can be separately co-segregated with different transgenic vectors, and rejoin via protein trans-splicing to reconstitute a full-length marker protein in host cells receiving all intended vectors. Using a lentiviral system, we create and validate 2-split Hygromycin, Puromycin, Neomycin and Blasticidin resistance genes as well as mScarlet fluorescent proteins. By combining split points, we create 3- and 6-split Hygromycin resistance genes, demonstrating that higher-degree split markers can be generated by a "chaining" design. We adapt the split marker system for selecting biallelically engineered cells after CRISPR gene editing. Future engineering of split markers may allow selection of a higher number of genetic modifications in target cells.
Topics: CRISPR-Cas Systems; Cell Line, Tumor; Cinnamates; Drug Resistance, Bacterial; Gene Editing; Gene Transfer Techniques; Genetic Engineering; Genetic Vectors; HEK293 Cells; HeLa Cells; Humans; Hygromycin B; Induced Pluripotent Stem Cells; Inteins; Lentivirus; Luminescent Proteins; Neomycin; Nucleosides; Protein Splicing; Puromycin; Trans-Splicing; Transgenes
PubMed: 31672965
DOI: 10.1038/s41467-019-12891-2 -
Current Protein & Peptide Science 2019Protein splicing domains, also called inteins, have become a powerful biotechnological tool for applications involving molecular biology and protein engineering. Early... (Review)
Review
Protein splicing domains, also called inteins, have become a powerful biotechnological tool for applications involving molecular biology and protein engineering. Early applications of inteins focused on self-cleaving affinity tags, generation of recombinant polypeptide α-thioesters for the production of semisynthetic proteins and backbone cyclized polypeptides. The discovery of naturallyoccurring split-inteins has allowed the development of novel approaches for the selective modification of proteins both in vitro and in vivo. This review gives a general introduction to protein splicing with a focus on their role in expanding the applications of intein-based technologies in protein engineering and chemical biology.
Topics: Binding Sites; Biocatalysis; Biosensing Techniques; Biotechnology; Inteins; Peptides; Protein Binding; Protein Conformation; Protein Engineering; Protein Splicing; Proteins; Recombinant Proteins
PubMed: 30734675
DOI: 10.2174/1389203720666190208110416 -
Frontiers in Microbiology 2023Protein splicing is a posttranslational process in which an intein segment excises itself from two flanking peptides, referred to as exteins. In the native context,... (Review)
Review
Protein splicing is a posttranslational process in which an intein segment excises itself from two flanking peptides, referred to as exteins. In the native context, protein splicing results in two separate protein products coupled to the activation of the intein-containing host protein. Inteins are generally described as either full-length inteins, mini-inteins or split inteins, which are differentiated by their genetic structure and features. Inteins can also be divided into three classes based on their splicing mechanisms, which differ in the location of conserved residues that mediate the splicing pathway. Although inteins were once thought to be selfish genetic elements, recent evidence suggests that inteins may confer a genetic advantage to their host cells through posttranslational regulation of their host proteins. Finally, the ability of modified inteins to splice and cleave their fused exteins has enabled many new applications in protein science and synthetic biology. In this review, we briefly cover the mechanisms of protein splicing, evidence for some inteins as environmental sensors, and intein-based applications in protein engineering.
PubMed: 38029209
DOI: 10.3389/fmicb.2023.1305848 -
Biosensors Apr 2022Cell-based sensors provide a flexible platform for screening biologically active targets and for monitoring their interactions in live cells. Their applicability extends... (Review)
Review
Cell-based sensors provide a flexible platform for screening biologically active targets and for monitoring their interactions in live cells. Their applicability extends across a vast array of biological research and clinical applications. Particularly, cell-based sensors are becoming a potent tool in drug discovery and cell-signaling studies by allowing function-based screening of targets in biologically relevant environments and enabling the in vivo visualization of cellular signals in real-time with an outstanding spatiotemporal resolution. In this review, we aim to provide a clear view of current cell-based sensor technologies, their limitations, and how the recent improvements were using intein-mediated protein engineering. We first discuss the characteristics of cell-based sensors and present several representative examples with a focus on their design strategies, which differentiate cell-based sensors from in vitro analytical biosensors. We then describe the application of intein-mediated protein engineering technology for cell-based sensor fabrication. Finally, we explain the characteristics of intein-mediated reactions and present examples of how the intein-mediated reactions are used to improve existing methods and develop new approaches in sensor cell fabrication to address the limitations of current technologies.
Topics: Biosensing Techniques; Inteins; Protein Engineering; Signal Transduction
PubMed: 35624584
DOI: 10.3390/bios12050283 -
Microorganisms Dec 2020Inteins are mobile genetic elements that apply standard enzymatic strategies to excise themselves post-translationally from the precursor protein via protein splicing.... (Review)
Review
Inteins are mobile genetic elements that apply standard enzymatic strategies to excise themselves post-translationally from the precursor protein via protein splicing. Since their discovery in the 1990s, recent advances in intein technology allow for them to be implemented as a modern biotechnological contrivance. Radical improvement in the structure and catalytic framework of - and -splicing inteins devised the development of engineered inteins that contribute to various efficient downstream techniques. Previous literature indicates that implementation of intein-mediated splicing has been extended to in vivo systems. Besides, the homing endonuclease domain also acts as a versatile biotechnological tool involving genetic manipulation and control of monogenic diseases. This review orients the understanding of inteins by sequentially studying the distribution and evolution pattern of intein, thereby highlighting a role in genetic mobility. Further, we include an in-depth summary of specific applications branching from protein purification using self-cleaving tags to protein modification, post-translational processing and labelling, followed by the development of intein-based biosensors. These engineered inteins offer a disruptive approach towards research avenues like biomaterial construction, metabolic engineering and synthetic biology. Therefore, this linear perspective allows for a more comprehensive understanding of intein function and its diverse applications.
PubMed: 33339089
DOI: 10.3390/microorganisms8122004 -
The Journal of Biological Chemistry May 2014Protein splicing is a posttranslational modification where intervening proteins (inteins) cleave themselves from larger precursor proteins and ligate their flanking... (Review)
Review
Protein splicing is a posttranslational modification where intervening proteins (inteins) cleave themselves from larger precursor proteins and ligate their flanking polypeptides (exteins) through a multistep chemical reaction. First thought to be an anomaly found in only a few organisms, protein splicing by inteins has since been observed in microorganisms from all domains of life. Despite this broad phylogenetic distribution, all inteins share common structural features such as a horseshoe-like pseudo two-fold symmetric fold, several canonical sequence motifs, and similar splicing mechanisms. Intriguingly, the splicing efficiencies and substrate specificity of different inteins vary considerably, reflecting subtle changes in the chemical mechanism of splicing, linked to their local structure and dynamics. As intein chemistry has widespread use in protein chemistry, understanding the structural and dynamical aspects of inteins is crucial for intein engineering and the improvement of intein-based technologies.
Topics: Amino Acids; Catalytic Domain; Inteins; Models, Molecular; Mutation; Protein Folding; Protein Precursors; Protein Splicing; Protein Structure, Tertiary; Proteins
PubMed: 24695731
DOI: 10.1074/jbc.R113.540302 -
Biological Chemistry Mar 2019Synthetic biologists aim at engineering controllable biological parts such as DNA, RNA and proteins in order to steer biological activities using external inputs.... (Review)
Review
Synthetic biologists aim at engineering controllable biological parts such as DNA, RNA and proteins in order to steer biological activities using external inputs. Proteins can be controlled in several ways, for instance by regulating the expression of their encoding genes with small molecules or light. However, post-translationally modifying pre-existing proteins to regulate their function or localization leads to faster responses. Conditional splicing of internal protein domains, termed inteins, is an attractive methodology for this purpose. Here we discuss methods to control intein activity with a focus on those compatible with applications in living cells.
Topics: Animals; Humans; Inteins; Protein Processing, Post-Translational; Protein Splicing; Proteins
PubMed: 30226200
DOI: 10.1515/hsz-2018-0309