LRRK2 activation controls the repair of damaged endomembranes in macrophages
Ontology highlight
ABSTRACT:
SUBMITTER: Dr. Susanne Herbst
PROVIDER: S-SCDT-EMBOJ-2020-104494 | biostudies-other |
REPOSITORIES: biostudies-other
Similar Datasets
Project description:Cells respond to endolysosome damage by either repairing the damage or targeting damaged endolysosomes for degradation via lysophagy. However, the signals regulating the decision for repair or lysophagy are poorly characterised. Here, we show that the Parkinson's disease (PD)-related kinase LRRK2 is activated in macrophages by pathogen- or sterile-induced endomembrane damage. LRRK2 recruits the Rab GTPase Rab8A to damaged endolysosomes as well as the ESCRT-III component CHMP4B, thereby favouring ESCRT-mediated repair. Conversely, in the absence of LRRK2 and Rab8A, damaged endolysosomes are targeted to lysophagy. These observations are recapitulated in macrophages from PD patients where pathogenic LRRK2 gain-of-function mutations result in the accumulation of endolysosomes which are positive for the membrane damage marker Galectin-3. Altogether, this work indicates that LRRK2 regulates endolysosomal homeostasis by controlling the balance between membrane repair and organelle replacement, uncovering an unexpected function for LRRK2, and providing a new link between membrane damage and PD.
| S-EPMC7507578 | biostudies-literature
Project description:Leucine-rich repeat kinase 2 (LRRK2) variants associated with Parkinson's disease (PD) and Crohn's disease lead to increased phosphorylation of its Rab substrates. While it has been recently shown that perturbations in cellular homeostasis including lysosomal damage can increase LRRK2 activity and localization to lysosomes, the molecular mechanisms by which LRRK2 activity is regulated have remained poorly defined. We performed a targeted siRNA screen to identify regulators of LRRK2 activity and identified Rab12 as a novel modulator of LRRK2-dependent phosphorylation of one of its substrates, Rab10. Using a combination of imaging and immunopurification methods to isolate lysosomes, we demonstrated that Rab12 is actively recruited to damaged lysosomes and leads to a local and LRRK2-dependent increase in Rab10 phosphorylation. PD-linked variants, including LRRK2 R1441G and VPS35 D620N, lead to increased recruitment of LRRK2 to the lysosome and a local elevation in lysosomal levels of pT73 Rab10. Together, these data suggest a conserved mechanism by which Rab12, in response to damage or expression of PD-associated variants, facilitates the recruitment of LRRK2 and phosphorylation of its Rab substrate(s) at the lysosome.
| S-EPMC10708889 | biostudies-literature
Project description:Infection by the protozoan parasite Toxoplasma gondii is highly prevalent worldwide and may have serious clinical manifestations in immunocompromised patients. T. gondii is an obligate intracellular parasite that infects almost any cell type in mammalian hosts, including immune cells. The immune cells express purinergic P2 receptors in their membrane--subdivided into P2Y and P2X subfamilies--whose activation is important for infection control. Here, we examined the effect of treatment with UTP and UDP in mouse peritoneal macrophages infected with T. gondii tachyzoites. Treatment with these nucleotides reduced parasitic load by 90%, but did not increase the levels of the inflammatory mediators NO and ROS, nor did it modulate host cell death by apoptosis or necrosis. On the other hand, UTP and UDP treatments induced early egress of tachyzoites from infected macrophages, in a Ca2+-dependent manner, as shown by scanning electron microscopy analysis, and videomicroscopy. In subsequent infections, prematurely egressed parasites had reduced infectivity, and could neither replicate nor inhibit the fusion of lysosomes to the parasitophorous vacuole. The use of selective agonists and antagonists of the receptor subtypes P2Y2 and P2Y4 and P2Y6 showed that premature parasite egress may be mediated by the activation of these receptor subtypes. Our results suggest that the activity of P2Y host cell receptors controls T. gondii infection in macrophages, highlighting the importance of pyrimidinergic signaling for innate immune system response against infection. Finally the P2Y receptors should be considered as new target for the development of drugs against T. gondii infection.
| S-EPMC4507979 | biostudies-literature
Project description:BackgroundAdipose-derived mesenchymal stromal cells (ADSCs) are multipotent stromal cells. The cells secrete a number of cytokines and growth factors and show immunoregulatory and proangiogenic properties. Their properties may be used to repair damaged tissues. The aim of our work is to explain the muscle damage repair mechanism with the utilization of the human adipose-derived mesenchymal stromal cells (hADSCs).MethodsFor the hADSCs isolation, we used the subcutaneous adipose tissue collected during the surgery. The murine hind limb ischemia was used as a model. The unilateral femoral artery ligation was performed on 10-12-week-old male C57BL/6NCrl and NOD SCID mice. The mice received PBS- (controls) or 1 × 106 hADSCs. One, 3, 7, 14 and 21 days after the surgery, we collected the gastrocnemius muscles for the immunohistochemical analysis. The results were analyzed with relevant tests using the Statistica software.ResultsThe retention time of hADSCs in the limb lasted about 14 days. In the mice receiving hADSCs, the improvement in the functionality of the damaged limb occurred faster than in the control mice. More new blood vessels were formed in the limbs of the mice receiving hADSCs than in limbs of the control mice. hADSCs also increased the infiltration of the macrophages with the M2 phenotype (7-AAD-/CD45+/F4/80+/CD206+) into the ischemic limbs. hADSCs introduced into the limb of mice secreted interleukin-6. This cytokine stimulates the emergence of the proangiogenic M2 macrophages, involved, among others, in the repair of a damaged tissue. Both macrophage depletion and IL-6 blockage suppressed the therapeutic effect of hADSCs. In the mice treated with hADSCs and liposomes with clodronate (macrophages depletion), the number of capillaries formed was lower than in the mice treated with hADSCs alone. Administration of hADSCs to the mice that received siltuximab (human IL-6 blocker) did not cause an influx of the M2 macrophages, and the number of capillaries formed was at the level of the control group, as in contrast to the mice that received only the hADSCs.ConclusionsThe proposed mechanism for the repair of the damaged muscle using hADSCs is based on the activity of IL-6. In our opinion, the cytokine, secreted by the hADSCs, stimulates the M2 macrophages responsible for repairing damaged muscle and forming new blood vessels.
| S-EPMC6417195 | biostudies-literature
Project description:Given that few ecosystems on the Earth have been unaffected by humans, restoring them holds great promise for stemming the biodiversity crisis and ensuring ecosystem services are provided to humanity. Nonetheless, few studies have documented the recovery of ecosystems globally or the rates at which ecosystems recover. Even fewer have addressed the added benefit of actively restoring ecosystems versus allowing them to recover without human intervention following the cessation of a disturbance. Our meta-analysis of 400 studies worldwide that document recovery from large-scale disturbances, such as oil spills, agriculture and logging, suggests that though ecosystems are progressing towards recovery following disturbances, they rarely recover completely. This result reinforces conservation of intact ecosystems as a key strategy for protecting biodiversity. Recovery rates slowed down with time since the disturbance ended, suggesting that the final stages of recovery are the most challenging to achieve. Active restoration did not result in faster or more complete recovery than simply ending the disturbances ecosystems face. Our results on the added benefit of restoration must be interpreted cautiously, because few studies directly compared different restoration actions in the same location after the same disturbance. The lack of consistent value added of active restoration following disturbance suggests that passive recovery should be considered as a first option; if recovery is slow, then active restoration actions should be better tailored to overcome specific obstacles to recovery and achieve restoration goals. We call for a more strategic investment of limited restoration resources into innovative collaborative efforts between scientists, local communities and practitioners to develop restoration techniques that are ecologically, economically and socially viable.
| S-EPMC5832705 | biostudies-literature
Project description:Many bacteria contain an RNA repair operon, encoding the RtcB RNA ligase and the RtcA RNA cyclase, that is regulated by the RtcR transcriptional activator. Although RtcR contains a divergent version of the CARF (CRISPR-associated Rossman fold) oligonucleotide-binding regulatory domain, both the specific signal that regulates operon expression and the substrates of the encoded enzymes are unknown. We report that tRNA fragments activate operon expression. Using a genetic screen in Salmonella enterica serovar Typhimurium, we find that the operon is expressed in the presence of mutations that cause tRNA fragments to accumulate. RtcA, which converts RNA phosphate ends to 2', 3'-cyclic phosphate, is also required. Operon expression and tRNA fragment accumulation also occur upon DNA damage. The CARF domain binds 5' tRNA fragments ending in cyclic phosphate, and RtcR oligomerizes upon binding these ligands, a prerequisite for operon activation. Our studies reveal a signaling pathway involving broken tRNAs and implicate the operon in tRNA repair.
| S-EPMC7790460 | biostudies-literature
Project description:Missense mutations in leucine-rich repeat kinase 2 (LRRK2) cause familial Parkinson's disease (PD). However, a potential role of wild-type LRRK2 in idiopathic PD (iPD) remains unclear. Here, we developed proximity ligation assays to assess Ser1292 phosphorylation of LRRK2 and, separately, the dissociation of 14-3-3 proteins from LRRK2. Using these proximity ligation assays, we show that wild-type LRRK2 kinase activity was selectively enhanced in substantia nigra dopamine neurons in postmortem brain tissue from patients with iPD and in two different rat models of the disease. We show that this occurred through an oxidative mechanism, resulting in phosphorylation of the LRRK2 substrate Rab10 and other downstream consequences including abnormalities in mitochondrial protein import and lysosomal function. Our study suggests that, independent of mutations, wild-type LRRK2 plays a role in iPD. LRRK2 kinase inhibitors may therefore be useful for treating patients with iPD who do not carry LRRK2 mutations.
| S-EPMC6344941 | biostudies-literature
Project description:To provide an overview on the present understanding of roles of oxidative DNA damage repair in cell signaling underlying bronchoconstriction common to, but not restricted to various forms of asthma and chronic obstructive pulmonary disease.Bronchoconstriction is a tightening of smooth muscle surrounding the bronchi and bronchioles with consequent wheezing and shortness of breath. Key stimuli include air pollutants, viral infections, allergens, thermal and osmotic changes, and shear stress of mucosal epithelium, triggering a wide range of cellular, vascular, and neural events. Although activation of nerve fibers, the role of G-proteins, protein kinases and Ca++, and molecular interaction within contracting filaments of muscle are well defined, the overarching mechanisms by which a wide range of stimuli initiate these events are not fully understood. Many, if not all, stimuli increase levels of reactive oxygen species, which are signaling and oxidatively modifying macromolecules, including DNA. The primary reactive oxygen species target in DNA is guanine, and 8-oxoguanine is one of the most abundant base lesions. It is repaired by 8-oxoguanine DNA glycosylase1 during base excision repair processes. The product, free 8-oxo-7,8-dihydro-2'-deoxyguanosine base, is bound by 8-oxoguanine DNA glycosylase1 with high affinity, and the complex then functions as an activator of small guanosine triphosphatases, triggering pathways for inducing gene expression and contraction of intracellular filaments in mast and smooth muscle cells.Oxidative DNA damage repair-mediated cell activation signaling result in gene expression that 'primes' the mucosal epithelium and submucosal tissues to generate mediators of airway smooth muscle contractions.
| S-EPMC4940044 | biostudies-literature
Project description:Male germ cell DNA damage, after exposure to radiation, exogenous chemicals, or chemotherapeutic agents, is a major cause of male infertility. DNA-damaged spermatozoa can fertilize oocytes; this is of concern because there is limited information on the capacity of early embryos to repair a damaged male genome or on the fate of these embryos if repair is inadequate. We hypothesized that the early activation of DNA damage response in the early embryo is a critical determinant of its fate. The objective of this study was to assess the DNA damage response and mitochondrial function as a measure of the energy supply for DNA repair and general health in cleavage-stage embryos sired by males chronically exposed to an anticancer alkylating agent, cyclophosphamide. Male rats were treated with saline or cyclophosphamide (6 mg/kg/day) for 4 weeks and mated to naturally cycling females. Pronuclear two- and eight-cell embryos were collected for immunofluorescence analysis of mitochondrial function and biomarkers of the DNA damage response: ?H2AX foci, 53BP1 reactivity, and poly(ADP-ribose) polymer formation. Mitochondrial activities did not differ between embryos sired by control- and cyclophosphamide-exposed males. At the two-cell stage, there was no treatment-related increase in DNA double-strand breaks; by the eight-cell stage, a significant increase was noted, as indicated by increased medium and large ?H2AX foci. This was accompanied by a dampened DNA repair response, detected as a decrease in the nuclear intensity of poly(ADP-ribose) polymers. The micronuclei formed in cyclophosphamide-sired embryos contained large ?H2AX foci and enhanced poly(ADP-ribose) polymer and 53BP1 reactivity compared with their nuclear counterparts. Thus, paternal cyclophosphamide exposure activated a DNA damage response in cleavage-stage embryos. Furthermore, this damage response may be useful in assessing embryo quality and developmental competence.
| S-EPMC3355317 | biostudies-literature
Project description:Dysfunction of the immune system and mitochondrial metabolism has been associated with Parkinson's disease (PD) pathology. Mutations and increased kinase activity of leucine-rich repeat kinase 2 (LRRK2) are linked to both idiopathic and familial PD. However, the function of LRRK2 in the immune cells under inflammatory conditions is contradictory. Our results showed that lipopolysaccharide (LPS) stimulation increased the kinase activity of LRRK2 in parental RAW 264.7 (WT) cells. In addition to this, LRRK2 deletion in LRRK2 KO RAW 264.7 (KO) cells altered cell morphology following LPS stimulation compared to the WT cells, as shown by an increase in the cell impedance as observed by the xCELLigence measurements. LPS stimulation caused an increase in the cellular reactive oxygen species (ROS) levels in both WT and KO cells. However, WT cells displayed a higher ROS level compared to the KO cells. Moreover, LRRK2 deletion led to a reduction in interleukin-6 (IL-6) inflammatory cytokine and cyclooxygenase-2 (COX-2) expression and an increase in lactate production after LPS stimulation compared to the WT cells. These data illustrate that LRRK2 has an effect on inflammatory processes in RAW macrophages upon LPS stimulation.
| S-EPMC9865464 | biostudies-literature