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MBio Aug 2023In the apicomplexans, endocytosed cargos (e.g., hemoglobin) are trafficked to a specialized organelle for digestion. This follows a unique endocytotic process at the...
In the apicomplexans, endocytosed cargos (e.g., hemoglobin) are trafficked to a specialized organelle for digestion. This follows a unique endocytotic process at the micropore/cytostome in these parasites. However, the mechanism underlying endocytic trafficking remains elusive, due to the repurposing of classical endocytic proteins for the biogenesis of apical organelles. To resolve this issue, we have exploited the genetic tractability of the model apicomplexan , which ingests host cytosolic materials (e.g., green fluorescent protein[GFP]). We determined an association between protein prenylation and endocytic trafficking, and using an alkyne-labeled click chemistry approach, the prenylated proteome was characterized. Genome editing, using clustered regularly interspaced short palindromic repaet/CRISPR-associated nuclease 9 (CRISPR/Cas9), was efficiently utilized to generate genetically modified lines for the functional screening of 23 prenylated candidates. This identified four of these proteins that regulate the trafficking of endocytosed GFP vesicles. Among these proteins, Rab1B and YKT6.1 are highly conserved but are non-classical endocytic proteins in eukaryotes. Confocal imaging analysis showed that Rab1B and Ras are substantially localized to both the trans-Golgi network and the endosome-like compartments in the parasite. Conditional knockdown of Rab1B caused a rapid defect in secretory trafficking to the rhoptry bulb, suggesting a trafficking intersection role for the key regulator Rab1B. Further experiments confirmed a critical role for protein prenylation in regulating the stability/activity of these proteins (i.e., Rab1B and YKT6.1) in the parasite. Our findings define the molecular basis of endocytic trafficking and reveal a potential intersection function of Rab1B on membrane trafficking in . This might extend to other related protists, including the malarial parasites. IMPORTANCE The protozoan establishes a permissive niche, in host cells, that allows parasites to acquire large molecules such as proteins. Numerous studies have demonstrated that the parasite repurposes the classical endocytic components for secretory sorting to the apical organelles, leaving the question of endocytic transport to the lysosome-like compartment unclear. Recent studies indicated that endocytic trafficking is likely to associate with protein prenylation in malarial parasites. This information promoted us to examine this association in the model apicomplexan and to identify the key components of the prenylated proteome that are involved. By exploiting the genetic tractability of and a host GFP acquisition assay, we reveal four non-classical endocytic proteins that regulate the transport of endocytosed cargos (e.g., GFP) in . Thus, we extend the principle that protein prenylation regulates endocytic trafficking and elucidate the process of non-classical endocytosis in and potentially in other related protists.
Topics: Toxoplasma; Proteome; Protozoan Proteins; Protein Transport; Endosomes; Green Fluorescent Proteins
PubMed: 37548452
DOI: 10.1128/mbio.01309-23 -
Archivum Immunologiae Et Therapiae... Apr 2021The review discusses a new approach to the prevention and treatment of viral infections based on the use of pine needles polyprenyl phosphate (PPP) and associated with... (Review)
Review
The review discusses a new approach to the prevention and treatment of viral infections based on the use of pine needles polyprenyl phosphate (PPP) and associated with the infringement of prenylation process-the attachment of farnesol or geranyl geraniol to the viral protein. Currently, prenylation has been detected in type 1 adenovirus, hepatitis C virus, several herpes viruses, influenza virus, HIV. However, this list is far from complete, given that prenylated proteins play an extremely important role in the activity of the virus. We assume that the interferon produced in response to PPP may suppress expression of the SREBP2 transcription factor. As a result, the mevalonic acid pathway is violated and, as a result, the formation of early polyprenols precursors (geraniol, geranyl geraniol, farnesol), which are necessary for the prenylation of viral proteins, is blocked and the formation of mature, virulent virus particles is broken. As a consequence, the maturation of viral particles is inhibited, and defective particles are formed. Polyprenol was extracted from greenery (pine, fir and spruce needles, mulberry leaves, etc.), purified by chromatography, phosphorylated and identified by HPLC and NMR. Obtained PPP was used as antiviral in some experimental models in vitro and in vivo. During numerous studies, it was found that PPP manifested versatile antiviral effects, both in vitro and in vivo. The maximum effect was observed with viruses in which the presence of prenylated proteins was established, namely influenza A virus, HIV-1, tick-borne encephalitis virus, hepatitis A and C viruses, herpes simplex viruses type 1 and 2, some coronavirus. The available data obtained both in the experimental conditions and during clinical trials allow us to regard PPPs as safe and effective medicine for prevention and treatment of viral diseases.
Topics: Animals; Antiviral Agents; Clinical Trials as Topic; Disease Models, Animal; Gene Expression Regulation; Humans; Interferons; Microscopy, Electron; Pinus; Polyisoprenyl Phosphates; Protein Prenylation; Sterol Regulatory Element Binding Protein 2; Treatment Outcome; Viral Proteins; Virion; Virus Diseases; Virus Replication
PubMed: 33811524
DOI: 10.1007/s00005-021-00613-w -
International Journal of Environmental... Jul 2022The cholesterol biosynthesis represents a crucial metabolic pathway for cellular homeostasis. The end products of this pathway are sterols, such as cholesterol, which... (Review)
Review
The cholesterol biosynthesis represents a crucial metabolic pathway for cellular homeostasis. The end products of this pathway are sterols, such as cholesterol, which are essential components of cell membranes, precursors of steroid hormones, bile acids, and other molecules such as ubiquinone. Furthermore, some intermediates of this metabolic system perform biological activity in specific cellular compartments, such as isoprenoid molecules that can modulate different signal proteins through the prenylation process. The defects of prenylation represent one of the main causes that promote the activation of inflammation. In particular, this mechanism, in association with oxidative stress, induces a dysfunction of the mitochondrial activity. The purpose of this review is to describe the pleiotropic role of prenylation in neuroinflammation and to highlight the consequence of the defects of prenylation.
Topics: Cholesterol; Humans; Mevalonic Acid; Neuroinflammatory Diseases; Oxidative Stress; Prenylation
PubMed: 35897423
DOI: 10.3390/ijerph19159061 -
Angewandte Chemie (International Ed. in... Aug 2022In nature, prenylation and geranylation are two important metabolic processes for the creation of hemiterpenoids and monoterpenoids under enzyme catalysis. Herein, we...
In nature, prenylation and geranylation are two important metabolic processes for the creation of hemiterpenoids and monoterpenoids under enzyme catalysis. Herein, we have demonstrated bioinspired unnatural prenylation and geranylation of oxindoles using the basic industrial feedstock isoprene through ligand regulation under Pd catalysis. Pentenylated oxindoles (with C added) were attained with high selectivity when using a bisphosphine ligand, whereas upon switching to a monophosphine ligand, selectivity toward geranylated oxindoles (with C added) was achieved. Moreover, the head-to-head product could be further isomerized to an internal skipped diene under Pd-H catalysis. No stoichiometric by-product was formed in the process.
Topics: Butadienes; Catalysis; Hemiterpenes; Ligands; Oxindoles; Palladium; Prenylation
PubMed: 35650687
DOI: 10.1002/anie.202207202 -
The Enzymes 2020The reversible (de)carboxylation of unsaturated carboxylic acids is carried out by the UbiX-UbiD system, ubiquitously present in microbes. The biochemical basis of this...
The reversible (de)carboxylation of unsaturated carboxylic acids is carried out by the UbiX-UbiD system, ubiquitously present in microbes. The biochemical basis of this challenging reaction has recently been uncovered by the discovery of the UbiD cofactor, prenylated FMN (prFMN). This heavily modified flavin is synthesized by the flavin prenyltransferase UbiX, which catalyzes the non-metal dependent prenyl transfer from dimethylallyl(pyro)phosphate (DMAP(P)) to the flavin N5 and C6 positions, creating a fourth non-aromatic ring. Following prenylation, prFMN undergoes oxidative maturation to form the iminium species required for UbiD activity. prFMN acts as a prostethic group and is bound via metal ion mediated interactions between UbiD and the prFMN phosphate moiety. The modified isoalloxazine ring is place adjacent to the E(D)-R-E UbiD signature sequent motif. The fungal ferulic acid decarboxylase Fdc from Aspergillus niger has emerged as a UbiD-model system, and has yielded atomic level insight into the prFMN mediated (de)carboxylation. A wealth of data now supports a mechanism reliant on reversible 1,3 dipolar cycloaddition between substrate and cofactor for this enzyme. This poses the intriguing question whether a similar mechanism is used by all UbiD enzymes, especially those that act as carboxylases on inherently more difficult substrates such as phenylphosphate or benzene/naphthalene. Indeed, considerable variability in terms of oligomerization, domain motion and active site structure is now reported for the UbiD family.
Topics: Aspergillus niger; Catalytic Domain; Dimethylallyltranstransferase; Flavin Mononucleotide; Fungal Proteins; Prenylation
PubMed: 32951834
DOI: 10.1016/bs.enz.2020.05.013 -
Frontiers in Molecular Biosciences 2021Newly synthesized small GTPases in the Ras and Rho families are prenylated by cytosolic prenyltransferases and then escorted by chaperones to membranes, the nucleus, and... (Review)
Review
Newly synthesized small GTPases in the Ras and Rho families are prenylated by cytosolic prenyltransferases and then escorted by chaperones to membranes, the nucleus, and other sites where the GTPases participate in a variety of signaling cascades. Understanding how prenylation and trafficking are regulated will help define new therapeutic strategies for cancer and other disorders involving abnormal signaling by these small GTPases. A growing body of evidence indicates that splice variants of SmgGDS (gene name RAP1GDS1) are major regulators of the prenylation, post-prenylation processing, and trafficking of Ras and Rho family members. SmgGDS-607 binds pre-prenylated small GTPases, while SmgGDS-558 binds prenylated small GTPases. This review discusses the history of SmgGDS research and explains our current understanding of how SmgGDS splice variants regulate the prenylation and trafficking of small GTPases. We discuss recent evidence that mutant forms of RabL3 and Rab22a control the release of small GTPases from SmgGDS, and review the inhibitory actions of DiRas1, which competitively blocks the binding of other small GTPases to SmgGDS. We conclude with a discussion of current strategies for therapeutic targeting of SmgGDS in cancer involving splice-switching oligonucleotides and peptide inhibitors.
PubMed: 34222337
DOI: 10.3389/fmolb.2021.685135 -
Current Protein & Peptide Science 2022Clodronate, a non-nitrogen-containing bisphosphonate (non-NBP), is intracellularly converted into non-hydrolyzable ATP analogs. Clodronate and its analogs impair normal...
BACKGROUND
Clodronate, a non-nitrogen-containing bisphosphonate (non-NBP), is intracellularly converted into non-hydrolyzable ATP analogs. Clodronate and its analogs impair normal cell functions, including the exocytosis process. However, how this occurs in mast cells is still not well characterized.
OBJECTIVE
To summarize the possible mechanisms of clodronate-mediated exocytosis inhibition in mast cells.
RESULTS
Non-NBPs display several possible mechanisms of exocytosis inhibition in various cell types, including vesicular nucleotide transporter (VNUT) and purinergic receptor inhibition. Inhibition of purinergic receptors has been shown in mast cells, but VNUT inhibition remains to be confirmed. Inhibition of protein prenylation by non-NBPs has also been shown; however, direct evidence of non-NBPs in prenylated exocytosis proteins is still contradictory. Finally, non-NBPs may inhibit mast cell exocytosis via impairment of protein pyrophosphorylation. This mechanism is less studied, and direct evidence of the involvement of pyrophosphorylated proteins in exocytosis is still lacking.
CONCLUSION
Non-NBPs may affect mast cell exocytosis by interacting with purinergic receptors or VNUT or by preventing post-translational modifications of exocytosis protein(s), i.e., prenylation and pyrophosphorylation. The latter needs further investigation to provide direct evidence of a role for non- NBPs.
Topics: Diphosphonates; Clodronic Acid; Exocytosis; Mast Cells
PubMed: 35726425
DOI: 10.2174/1389203723666220620164024 -
BioRxiv : the Preprint Server For... Jul 2023Prenylation is a universal and irreversible post-translational modification that supports membrane interactions of proteins involved in various cellular processes,...
Prenylation is a universal and irreversible post-translational modification that supports membrane interactions of proteins involved in various cellular processes, including migration, proliferation, and survival. Thus, dysregulation of prenylation contributes to multiple disorders, including cancers, vascular diseases, and neurodegenerative diseases. During prenylation, prenyltransferase enzymes tether metabolically produced isoprenoid lipids to proteins via a thioether linkage. Pharmacological inhibition of the lipid synthesis pathway by statins has long been a therapeutic approach to control hyperlipidemia. Building on our previous finding that statins inhibit membrane association of G protein γ (Gγ) in a subtype-dependent manner, we investigated the molecular reasoning for this differential. We examined the prenylation efficacy of carboxy terminus (Ct) mutated Gγ in cells exposed to Fluvastatin and prenyl transferase inhibitors and monitored the subcellular localization of fluorescently tagged Gγ subunits and their mutants using live-cell confocal imaging. Reversible optogenetic unmasking-masking of Ct residues was used to probe their contribution to the prenylation process and membrane interactions of the prenylated proteins. Our findings suggest that specific Ct residues regulate membrane interactions of the Gγ polypeptide statin sensitivity, and prenylation efficacy. Our results also show that a few hydrophobic and charged residues at the Ct are crucial determinants of a protein's prenylation ability, especially under suboptimal conditions. Given the cell and tissue-specific expression of different Gγ subtypes, our findings explain how and why statins differentially perturb heterotrimeric G protein signaling in specific cells and tissues. Our results may provide molecular reasoning for repurposing statins as Ras oncogene inhibitors and the failure of using prenyltransferase inhibitors in cancer treatment.
PubMed: 37461501
DOI: 10.1101/2023.07.04.547731 -
Frontiers in Pharmacology 2019Over the last decade, several studies demonstrated that prenylation of flavonoids enhances various biological activities as compared to the respective nonprenylated... (Review)
Review
Over the last decade, several studies demonstrated that prenylation of flavonoids enhances various biological activities as compared to the respective nonprenylated compounds. In line with this, the natural prenylated isoflavonoid alpinumisoflavone (AIF) has been explored for a number of biological and pharmacological effects (therapeutic potential). In this review, we summarize the current information on health-promoting properties of AIF. Reported data evidenced that AIF has a multitherapeutic potential with antiosteoporotic, antioxidant and anti-inflammatory, antimicrobial, anticancer, estrogenic and antiestrogenic, antidiabetic, and neuroprotective properties. However, research on these aspects of AIF is not sufficient and needs to be reevaluated using more appropriate methods and methodology. Further series of studies are needed to confirm these pharmacological effects, and this review should lay the basis for the design of respective investigations. Overall, despite the drawbacks of studies recorded, AIF exhibits a potential as drug candidate.
PubMed: 31551770
DOI: 10.3389/fphar.2019.00952 -
Respiratory Research Dec 2022Chronic obstructive pulmonary disease (COPD) is a progressive disorder that causes airway obstruction and lung inflammation. The first-line treatment of COPD is the...
Nebulization of risedronate alleviates airway obstruction and inflammation of chronic obstructive pulmonary diseases via suppressing prenylation-dependent RAS/ERK/NF-κB and RhoA/ROCK1/MLCP signaling.
BACKGROUND
Chronic obstructive pulmonary disease (COPD) is a progressive disorder that causes airway obstruction and lung inflammation. The first-line treatment of COPD is the bronchodilators of β2-agonists and antimuscarinic drugs, which can help control the airway obstruction, but the long-term use might render the drug tolerance. Bisphosphonates are widely used in osteoclast-mediated bone diseases treatment for decades. For drug repurposing, can delivery of a third generation of nitrogen-containing bisphosphonate, risedronate (RIS) ameliorate the progression of COPD?
METHODS
COPD rats or mice models have been established through cigarette-smoking and elastase injection, and then the animals are received RIS treatment via nebulization. Lung deposition of RIS was primarily assessed by high-performance liquid chromatography (HPLC). The respiratory parameters of airway obstruction in COPD rats and mice were documented using plethysmography method and resistance-compliance system.
RESULTS
High lung deposition and bioavailability of RIS was monitored with 88.8% of RIS input dose. We found that RIS could rescue the lung function decline of airspace enlargement and mean linear intercept in the COPD lung. RIS could curb the airway obstruction by suppressing 60% of the respiratory resistance and elevating the airway's dynamic compliance, tidal volume and mid-expiratory flow. As an inhibitor of farnesyl diphosphate synthase (FDPS), RIS suppresses FDPS-mediated RAS and RhoA prenylation to obstruct its membrane localization in airway smooth muscle cells (ASMCs), leading to the inhibition of downstream ERK-MLCK and ROCK1-MLCP pathway to cause ASMCs relaxation. Additionally, RIS nebulization impeded pro-inflammatory cell accumulation, particularly macrophages infiltration in alveolar parenchyma. The NF-κB, tumor necrosis factor-alpha, IL-1β, IL-8, and IL-6 declined in microphages following RIS nebulization. Surprisingly, nebulization of RIS could overcome the tolerance of β2-agonists in COPD-rats by increasing the expression of β2 receptors.
CONCLUSIONS
Nebulization of RIS could alleviate airway obstruction and lung inflammation in COPD, providing a novel strategy for treating COPD patients, even those with β2-agonists tolerance.
Topics: Rats; Mice; Animals; NF-kappa B; Risedronic Acid; Pulmonary Disease, Chronic Obstructive; Lung; Airway Obstruction; Inflammation; Prenylation; rho-Associated Kinases
PubMed: 36575527
DOI: 10.1186/s12931-022-02274-5