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International Journal of Molecular... Feb 2021Ribosome-inactivating proteins (RIPs) are a class of cytotoxic enzymes that can inhibit protein translation by depurinating rRNA. Most plant RIPs are synthesized with a...
Ribosome-inactivating proteins (RIPs) are a class of cytotoxic enzymes that can inhibit protein translation by depurinating rRNA. Most plant RIPs are synthesized with a leader sequence that sequesters the proteins to a cell compartment away from the host ribosomes. However, several rice RIPs lack these signal peptides suggesting they reside in the cytosol in close proximity to the plant ribosomes. This paper aims to elucidate the physiological function of two nucleocytoplasmic RIPs from rice, in particular, the type 1 RIP referred to as OsRIP1 and a presumed type 3 RIP called nuRIP. Transgenic rice lines overexpressing these RIPs were constructed and studied for developmental effects resulting from this overexpression under greenhouse conditions. In addition, the performance of transgenic seedlings in response to drought, salt, abscisic acid and methyl jasmonate treatment was investigated. Results suggest that both RIPs can affect methyl jasmonate mediated stress responses.
Topics: Abscisic Acid; Acetates; Cyclopentanes; Cytosol; Gene Expression Regulation, Plant; Green Fluorescent Proteins; Oryza; Oxylipins; Phenotype; Plant Proteins; Plants, Genetically Modified; Protein Biosynthesis; Ribosomes; Salts; Saporins; Seedlings; Stress, Physiological
PubMed: 33535383
DOI: 10.3390/ijms22031434 -
Toxins Jan 2021Ribosome-inactivating proteins (RIPs) are plant toxins that irreversibly damage ribosomes and other substrates, thus causing cell death. RIPs are classified in type 1...
Ribosome-inactivating proteins (RIPs) are plant toxins that irreversibly damage ribosomes and other substrates, thus causing cell death. RIPs are classified in type 1 RIPs, single-chain enzymatic proteins, and type 2 RIPs, consisting of active A chains, similar to type 1 RIPs, linked to lectin B chains, which enable the rapid internalization of the toxin into the cell. For this reason, many type 2 RIPs are very cytotoxic, ricin, volkensin and stenodactylin being the most toxic ones. From the caudex of (Mast.) Engl., a new type 2 RIP, named kirkiin, was purified by affinity chromatography on acid-treated Sepharose CL-6B and gel filtration. The lectin, with molecular weight of about 58 kDa, agglutinated erythrocytes and inhibited protein synthesis in a cell-free system at very low concentrations. Moreover, kirkiin was able to depurinate mammalian and yeast ribosomes, but it showed little or no activity on other nucleotide substrates. In neuroblastoma cells, kirkiin inhibited protein synthesis and induced apoptosis at doses in the pM range. The biological characteristics of kirkiin make this protein a potential candidate for several experimental pharmacological applications both alone for local treatments and as component of immunoconjugates for systemic targeting in neurodegenerative studies and cancer therapy.
Topics: Antineoplastic Agents, Phytogenic; Cell Line, Tumor; Cell Survival; Erythrocyte Aggregation; Humans; Molecular Weight; Neuroblastoma; Passifloraceae; Protein Biosynthesis; Protein Synthesis Inhibitors; Ribosome Inactivating Proteins, Type 2; Ribosomes
PubMed: 33499082
DOI: 10.3390/toxins13020081 -
Ecology and Evolution Jan 2021Ancient DNA research has developed rapidly over the past few decades due to improvements in PCR and next-generation sequencing (NGS) technologies, but challenges still...
Ancient DNA research has developed rapidly over the past few decades due to improvements in PCR and next-generation sequencing (NGS) technologies, but challenges still exist. One major challenge in relation to ancient DNA research is to recover genuine endogenous ancient DNA sequences from raw sequencing data. This is often difficult due to degradation of ancient DNA and high levels of contamination, especially homologous contamination that has extremely similar genetic background with that of the real ancient DNA. In this study, we collected whole-genome sequencing (WGS) data from 6 ancient samples to compare different mapping algorithms. To further explore more effective methods to separate endogenous DNA from homologous contaminations, we attempted to recover reads based on ancient DNA specific characteristics of deamination, depurination, and DNA fragmentation with different parameters. We propose a quick and improved pipeline for separating endogenous ancient DNA while simultaneously decreasing homologous contaminations to very low proportions. Our goal in this research was to develop useful recommendations for ancient DNA mapping and for separation of endogenous DNA to facilitate future studies of ancient DNA.
PubMed: 33437437
DOI: 10.1002/ece3.7056 -
SLAS Discovery : Advancing Life... Mar 2021Saporin, a type I ribosome-inactivating protein from soapwort plant, is a potent protein synthesis inhibitor. Catalytically, saporin is a characteristic -glycosidase,...
Saporin, a type I ribosome-inactivating protein from soapwort plant, is a potent protein synthesis inhibitor. Catalytically, saporin is a characteristic -glycosidase, and it depurinates a specific adenine residue from a universally conserved loop of the major ribosomal RNA (rRNA) of eukaryotic cells. It is well-known that saporin induces apoptosis through different pathways, including ribotoxic stress response, cell signal transduction, genomic DNA fragmentation and RNA abasic lyase (RAlyase) activity, and NAD depletion by poly-(ADP)-ribose polymerase hyperactivation. Saporin's high enzymatic activity, high stability, and resistance to conjugation procedures make it a well-suited tool for immunotherapy approaches.In the present study, we focus on saporin-based targeted toxins that may be efficacious therapeutic agents for the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Our discussed points suggest that saporin may be a strategic molecule for therapeutic knockout treatments and a powerful candidate for novel drugs in the struggle against coronavirus 2019 (COVID-19).
Topics: Antiviral Agents; Apoptosis; Humans; Immunotoxins; NAD; Poly(ADP-ribose) Polymerases; Saporins; Signal Transduction; COVID-19 Drug Treatment
PubMed: 33155515
DOI: 10.1177/2472555220970911 -
Scientific Reports Sep 2020Ribosome inactivating proteins (RIPs) are RNA N-glycosidases that depurinate a specific adenine residue in the conserved sarcin/ricin loop of 28S rRNA. These enzymes...
Ribosome inactivating proteins (RIPs) are RNA N-glycosidases that depurinate a specific adenine residue in the conserved sarcin/ricin loop of 28S rRNA. These enzymes are widely distributed among plants and bacteria. Previously, we have described for the first time RIP genes in mosquitoes belonging to the Culicidae family. We showed that these genes are derived from a single event of horizontal gene transfer (HGT) from a prokaryotic donor. Mosquito RIP genes are evolving under purifying selection, strongly suggesting that these toxins have acquired a functional role. In this work, we show the existence of two RIP encoding genes in the genome of the whitefly Bemisia tabaci, a hemiptera species belonging to the Aleyrodidae family distantly related to mosquitoes. Contamination artifacts were ruled out analyzing three independent B. tabaci genome databases. In contrast to mosquito RIPs, whitefly genes harbor introns and according to transcriptomic evidence are transcribed and spliced. Phylogeny and the taxonomic distribution strongly support that whitefly RIP genes are derived from an independent HGT event from a plant source. These results, along with our previous description of RIPs in Diptera, suggest that the acquired genes are functional in these insects and confer some fitness advantage.
Topics: Animals; Gene Expression Profiling; Gene Transfer, Horizontal; Genes, Insect; Genes, Plant; Genome, Insect; Hemiptera; Phylogeny; RNA, Ribosomal, 28S; Ribosome Inactivating Proteins; Selection, Genetic; Sequence Alignment; Sequence Analysis, DNA
PubMed: 32968092
DOI: 10.1038/s41598-020-72267-1 -
Journal of the American Chemical Society Sep 2020Solid-phase oligonucleotide synthesis (SPOS) based on phosphoramidite chemistry is currently the most widespread technique for DNA and RNA synthesis but suffers from...
Solid-phase oligonucleotide synthesis (SPOS) based on phosphoramidite chemistry is currently the most widespread technique for DNA and RNA synthesis but suffers from scalability limitations and high reagent consumption. Liquid-phase oligonucleotide synthesis (LPOS) uses soluble polymer supports and has the potential of being scalable. However, at present, LPOS requires 3 separate reaction steps and 4-5 precipitation steps per nucleotide addition. Moreover, long acid exposure times during the deprotection step degrade sequences with high A content (adenine) due to depurination and chain cleavage. In this work, we present the first one-pot liquid-phase DNA synthesis technique which allows the addition of one nucleotide in a one-pot reaction of sequential coupling, oxidation, and deprotection followed by a single precipitation step. Furthermore, we demonstrate how to suppress depurination during the addition of adenine nucleotides. We showcase the potential of this technique to prepare high-purity 4-arm PEG-T (T = thymine) and 4-arm PEG-A building blocks in multigram scale. Such complementary 4-arm PEG-DNA building blocks reversibly self-assemble into supramolecular model network hydrogels and facilitate the elucidation of bond lifetimes. These model network hydrogels exhibit new levels of mechanical properties (storage modulus, bond lifetimes) in DNA bonds at room temperature (melting at 44 °C) and thus open up pathways to next-generation DNA materials programmable through sequence recognition and available for macroscale applications.
Topics: DNA; Hydrogels; Models, Molecular; Molecular Structure; Polyethylene Glycols
PubMed: 32902960
DOI: 10.1021/jacs.0c05488 -
Frontiers in Immunology 2020The integrity of the genome is under constant threat of environmental and endogenous agents that cause DNA damage. Endogenous damage is particularly pervasive, occurring... (Review)
Review
The integrity of the genome is under constant threat of environmental and endogenous agents that cause DNA damage. Endogenous damage is particularly pervasive, occurring at an estimated rate of 10,000-30,000 per cell/per day, and mostly involves chemical DNA base lesions caused by oxidation, depurination, alkylation, and deamination. The base excision repair (BER) pathway is primary responsible for removing and repairing these small base lesions that would otherwise lead to mutations or DNA breaks during replication. Next to preventing DNA mutations and damage, the BER pathway is also involved in mutagenic processes in B cells during immunoglobulin (Ig) class switch recombination (CSR) and somatic hypermutation (SHM), which are instigated by uracil (U) lesions derived from activation-induced cytidine deaminase (AID) activity. BER is required for the processing of AID-induced lesions into DNA double strand breaks (DSB) that are required for CSR, and is of pivotal importance for determining the mutagenic outcome of uracil lesions during SHM. Although uracils are generally efficiently repaired by error-free BER, this process is surprisingly error-prone at the loci in proliferating B cells. Breakdown of this high-fidelity process outside of the loci has been linked to mutations observed in B-cell tumors and DNA breaks and chromosomal translocations in activated B cells. Next to its role in preventing cancer, BER has also been implicated in immune tolerance. Several defects in BER components have been associated with autoimmune diseases, and animal models have shown that BER defects can cause autoimmunity in a B-cell intrinsic and extrinsic fashion. In this review we discuss the contribution of BER to genomic integrity in the context of immune receptor diversification, cancer and autoimmune diseases.
Topics: Animals; B-Lymphocytes; DNA; DNA Breaks, Double-Stranded; DNA Damage; DNA Repair; Humans; Immune System; Immunoglobulin Class Switching; Somatic Hypermutation, Immunoglobulin
PubMed: 32547565
DOI: 10.3389/fimmu.2020.01084 -
Saudi Journal of Biological Sciences Jun 2020Plant ribosome-inactivating proteins (RIPs) are N-glycosidases which inhibit protein synthesis through depurination of the ribosomal RNA sequence. Type II RIPs are...
Plant ribosome-inactivating proteins (RIPs) are N-glycosidases which inhibit protein synthesis through depurination of the ribosomal RNA sequence. Type II RIPs are heterodimer proteins which can bind to cell surfaces. The cytotoxicity of these RIPs is different. spp. are a rich source of RIP proteins with different properties. In the present study, a type II RIP was isolated from plant that grows widely in the north of Iran, and different bioinformatics tools were used for the evaluation of physicochemical, functional and 3D protein characteristics. The results showed significant differences among isolated RIP and other Sambucus RIP proteins. The study of these differences can not only expand our insight into the functioning mechanisms of plant RIPs but also provide information about a novel RIP protein with potential biological applications.
PubMed: 32489302
DOI: 10.1016/j.sjbs.2020.02.009 -
Frontiers in Pharmacology 2020Stenodactylin, a highly toxic type 2 ribosome-inactivating protein purified from the caudex of Harms, is a potential anticancer drug candidate. Previous studies...
Stenodactylin, a highly toxic type 2 ribosome-inactivating protein purified from the caudex of Harms, is a potential anticancer drug candidate. Previous studies demonstrated that stenodactylin induces apoptosis and necroptosis in treated cells, involving the production of reactive oxygen species. We analyzed the effect of stenodactylin on Raji and Ramos (Human Burkitt's lymphoma cells) and MOLM-13 (acute myeloid leukemia cells). Moreover, we focused on the early events in MOLM-13 cells that characterize the cellular response to the toxin by whole-genome microarray analysis of gene expression. Treatment with stenodactylin induced the depurination of 28S rRNA within 4 h and increased the phosphorylation of p38 and JNK. A time-dependent activation of caspase 1, 2, 8, 9, 3/7 was also observed. Genome-wide gene expression microarray analysis revealed early changes in the expression of genes involved in the regulation of cell death, inflammation and stress response. After 4 h, a significant increase of transcript level was detectable for ATF3, BTG2, DUSP1, EGR1, and JUN. Increased upstream JUN signaling was also confirmed at protein level. The early response to stenodactylin treatment involves inflammatory and apoptotic signaling compatible with the activation of multiple cell death pathways. Because of the above described properties toward acute myeloid leukemia cells, stenodactylin may be a promising candidate for the design of new immunoconjugates for experimental cancer treatment.
PubMed: 32457623
DOI: 10.3389/fphar.2020.00630 -
Toxins Mar 2020Ribosome-inactivating proteins (RIPs) are N-glycosidases, which depurinate a specific adenine residue in the conserved α-sarcin/ricin loop (α-SRL) of rRNA. This loop... (Review)
Review
Ribosome-inactivating proteins (RIPs) are N-glycosidases, which depurinate a specific adenine residue in the conserved α-sarcin/ricin loop (α-SRL) of rRNA. This loop is important for anchoring elongation factor (EF-G for prokaryote or eEF2 for eukaryote) in mRNA translocation. Translation is inhibited after the attack. RIPs therefore may have been applied for anti-cancer, and anti-virus and other therapeutic applications. The main obstacles of treatment with RIPs include short plasma half-life, non-selective cytotoxicity and antigenicity. This review focuses on the strategies used to improve the pharmacological properties of RIPs on human immunodeficiency virus (HIV) and cancers. Coupling with polyethylene glycol (PEG) increases plasma time and reduces antigenicity. RIPs conjugated with antibodies to form immunotoxins increase the selective toxicity to target cells. The prospects for future development on the engineering of RIPs for improving their pharmacological properties are also discussed.
Topics: Animals; Anti-HIV Agents; Antineoplastic Agents; Dextrans; Humans; Polyethylene Glycols; Protein Engineering; Ribosome Inactivating Proteins
PubMed: 32182799
DOI: 10.3390/toxins12030167