Extracellular pectin-RALF phase separation mediates FERONIA global signaling function.

The FERONIA (FER)-LLG1 co-receptor and its peptide ligand RALF regulate myriad processes for plant growth and survival. Focusing on signal-induced cell surface responses, we discovered that intrinsically disordered RALF triggers clustering and endocytosis of its cognate receptors and FER- and LLG1-dependent endocytosis of non-cognate regulators of diverse processes, thus capable of broadly impacting downstream responses. RALF, however, remains extracellular. We demonstrate that RALF binds the cell wall polysaccharide pectin. They phase separate and recruit FER and LLG1 into pectin-RALF-FER-LLG1 condensates to initiate RALF-triggered cell surface responses. We show further that two frequently encountered environmental challenges, elevated salt and temperature, trigger RALF-pectin phase separation, promiscuous receptor clustering and massive endocytosis, and that this process is crucial for recovery from stress-induced growth attenuation. Our results support that RALF-pectin phase separation mediates an exoskeletal mechanism to broadly activate FER-LLG1-dependent cell surface responses to mediate the global role of FER in plant growth and survival.

Integrin Mechano-chemical Signaling Generates Plasma Membrane Nanodomains that Promote Cell Spreading.

Glycosylphosphatidylinositol-anchored proteins (GPI-APs) are a major class of lipid-anchored plasma membrane proteins. GPI-APs form nanoclusters generated by cortical acto-myosin activity. While our understanding of the physical principles governing this process is emerging, the molecular machinery and functional relevance of GPI-AP nanoclustering are unknown. Here, we first show that a membrane receptor signaling pathway directs nanocluster formation. Arg-Gly-Asp motif-containing ligands bound to the ß1-integrin receptor activate src and focal adhesion kinases, resulting in RhoA signaling. This cascade triggers actin-nucleation via specific formins, which, along with myosin activity, drive the nanoclustering of membrane proteins with actin-binding domains. Concurrently, talin-mediated activation of the mechano-transducer vinculin is required for the coupling of the acto-myosin machinery to inner-leaflet lipids, thereby generating GPI-AP nanoclusters. Second, we show that these nanoclusters are functional; disruption of their formation either in GPI-anchor remodeling mutants or in vinculin mutants impairs cell spreading and migration, hallmarks of integrin function.

To each its own: Mechanisms of cross-talk between GPI biosynthesis and cAMP-PKA signaling in Candida albicans versus Saccharomyces cerevisiae.

Candida albicans is an opportunistic fungal pathogen that can switch between yeast and hyphal morphologies depending on the environmental cues it receives. The switch to hyphal form is crucial for the establishment of invasive infections. The hyphal form is also characterized by the cell surface expression of hyphae-specific proteins, many of which are GPI-anchored and important determinants of its virulence. The coordination between hyphal morphogenesis and the expression of GPI-anchored proteins is made possible by an interesting cross-talk between GPI biosynthesis and the cAMP-PKA signaling cascade in the fungus; a parallel interaction is not found in its human host. On the other hand, in the nonpathogenic yeast, Saccharomyces cerevisiae, GPI biosynthesis is shut down when filamentation is activated and vice versa. This too is achieved by a cross-talk between GPI biosynthesis and cAMP-PKA signaling. How are diametrically opposite effects obtained from the cross-talk between two reasonably well-conserved pathways present ubiquitously across eukarya? This Review attempts to provide a model to explain these differences. In order to do so, it first provides an overview of the two pathways for the interested reader, highlighting the similarities and differences that are observed in C. albicans versus the well-studied S. cerevisiae model, before going on to explain how the different mechanisms of regulation are effected. While commonalities enable the development of generalized theories, it is hoped that a more nuanced approach, that takes into consideration species-specific differences, will enable organism-specific understanding of these processes and contribute to the development of targeted therapies.

COBRA-LIKE4 Modulates Cellulose Synthase Velocity and Facilitates Cellulose Deposition in the Secondary Cell Wall.

Cellulose is a critical component of secondary cell walls and woody tissues of plants. Cellulose synthase (CESA) complexes (CSCs) produce cellulose as they move within the plasma membrane, extruding glucan chains into the cell wall that coalesce and crystallize into cellulose fibrils. Here we examine COBRA-LIKE4 (COBL4), a GPI-anchored protein on the outer leaflet of the plasma membrane that is required for normal cellulose deposition in secondary cell walls. Characterization of the Arabidopsis (Arabidopsis thaliana) cobl4 mutant alleles called irregular xylem6, irx6-2 and irx6-3, showed reduced ⍺-cellulose content and lower crystallinity, supporting a role for COBL4 in maintaining cellulose quantity and quality. In live-cell imaging, mNeon Green-tagged CESA7 moved in the plasma membrane at higher speeds in the irx6-2 background compared to wild type. To test conservation of COBL4 function between herbaceous and woody plants, poplar (Populus trichocarpa) COBL4 homologs PtCOBL4a and PtCOBL4b were transformed into, and rescued, the Arabidopsis irx6 mutants. Using the Arabidopsis secondary cell wall-inducible VND7-GR system to study poplar COBL4 dynamics, YFP-tagged PtCOBL4a localized to the plasma membrane in regions of high cellulose deposition in secondary cell wall bands. As predicted for a lipid-linked protein, COBL4 was more mobile in the plane of the plasma membrane than CESA7 or a control plasma membrane marker. Following programmed cell death, COBL4 anchored to the secondary cell wall bands. These data support a role for COBL4 as a modulator of cellulose organization in the secondary cell wall, influencing cellulose production and CSC velocity at the plasma membrane.

Interaction of motor behaviour, cortical oscillations and deep brain stimulation in Parkinson disease.

Recent progress in the study of Parkinson's disease (PD) has highlighted the pivotal role of beta oscillations within the basal ganglia-thalamo-cortical network in modulating motor symptoms. Predominantly manifesting as transient bursts, these beta oscillations are central to the pathophysiology of PD motor symptoms, especially bradykinesia. Our central hypothesis is that increased bursting duration in cortex, coupled with kinematics of movement, disrupts the typical flow of neural information, leading to observable changes in motor behavior in PD. To explore this hypothesis, we employed an integrative approach, analyzing the interplay between moment-to-moment brain dynamics and movement kinematics, and the modulation of these relationships by therapeutic deep brain stimulation (DBS). Local field potentials were recorded from the hand motor (M1) and premotor cortical (PM) areas, and internal Globus Pallidus (GPi) in 26 PD patients undergoing DBS implantation surgery. Participants executed rapid alternating hand movements in 30-second blocks, both with and without therapeutic pallidal stimulation. Behaviorally, the analysis revealed bradykinesia, with hand movement cycle width increasing linearly over time during DBS-OFF blocks. Crucially, there was a moment-to-moment correlation between M1 low beta burst duration and movement cycle width, a relationship that dissipated with therapeutic DBS. Further analyses suggest that high gamma activity correlates with enhanced motor performance with DBS-ON. Regardless of the nature of coupling, DBS's modulation of cortical bursting activity appeared to amplify the brain signals' informational content regarding instantaneous movement changes. Our findings underscore that DBS significantly reshapes the interaction between motor behavior and neural signals in PD, not only modulating specific bands but also expanding the system's capability to process and relay information for motor control. These insights shed light on the possible network mechanisms underlying DBS therapeutic effects, suggesting a profound impact on both neural and motor domains. Mirpour and Pouratian investigate the interplay between movement, brain oscillations and deep brain stimulation (DBS) in Parkinson's disease. They show that beta oscillations interfere with the moment-to-moment control of movement, and that DBS can enhance communication within brain networks, mitigating these effects.

The clinical and genetic spectrum of inherited glycosylphosphatidylinositol deficiency disorders.

Inherited glycosylphosphatidylinositol deficiency disorders (IGDs) are a group of rare multisystem disorders arising from pathogenic variants in glycosylphosphatidylinositol anchor pathway (GPI-AP) genes. Despite associating 24 of at least 31 GPI-AP genes with human neurogenetic disease, prior reports are limited to single genes without consideration of the GPI-AP as a whole and with limited natural history data. In this multinational retrospective observational study, we systematically analyse the molecular spectrum, phenotypic characteristics and natural history of 83 individuals from 75 unique families with IGDs, including 70 newly reported individuals; the largest single cohort to date. Core clinical features were developmental delay or intellectual disability (DD/ID, 90%), seizures (83%), hypotonia (72%) and motor symptoms (64%). Prognostic and biologically significant neuroimaging features included cerebral atrophy (75%), cerebellar atrophy (60%), callosal anomalies (57%) and symmetric restricted diffusion of the central tegmental tracts (60%). Sixty-one individuals had multisystem involvement including gastrointestinal (66%), cardiac (19%) and renal (14%) anomalies. Though dysmorphic features were appreciated in 82%, no single dysmorphic feature had a prevalence >30%, indicating substantial phenotypic heterogeneity. Follow-up data were available for all individuals, 15 of whom were deceased at the time of writing. Median age at seizure onset was 6 months. Individuals with variants in synthesis stage genes of the GPI-AP exhibited a significantly shorter time to seizure onset than individuals with variants in transamidase and remodelling stage genes of the GPI-AP (P = 0.046). Forty individuals had intractable epilepsy. The majority of individuals experienced delayed or absent speech (95%), motor delay with non-ambulance (64%), and severe-to-profound DD/ID (59%). Individuals with a developmental epileptic encephalopathy (51%) were at greater risk of intractable epilepsy (P = 0.003), non-ambulance (P = 0.035), ongoing enteral feeds (P < 0.001) and cortical visual impairment (P = 0.007). Serial neuroimaging showed progressive cerebral volume loss in 87.5% and progressive cerebellar atrophy in 70.8%, indicating a neurodegenerative process. Genetic analyses identified 93 unique variants (106 total), including 22 novel variants. Exploratory analyses of genotype-phenotype correlations using unsupervised hierarchical clustering identified novel genotypic predictors of clinical phenotype and long-term outcome with meaningful implications for management. In summary, we expand both the mild and severe phenotypic extremities of the IGDs, provide insights into their neurological basis, and vitally, enable meaningful genetic counselling for affected individuals and their families.

Functional partnerships between GPI-anchored proteins and adhesion GPCRs.

Specific extracellular interaction between glycophosphatidylinositol (GPI)-anchored proteins and adhesion G protein-coupled receptors (aGPCRs) plays an important role in unique biological functions. GPI-anchored proteins are derived from a novel post-translational modification of single-span membrane molecules, while aGPCRs are bona fide seven-span transmembrane proteins with a long extracellular domain. Although various members of the two structurally-distinct protein families are known to be involved in a wide range of biological processes, many remain as orphans. Interestingly, accumulating evidence has pointed to a complex interaction and functional synergy between these two protein families. I discuss herein current understanding of specific functional partnerships between GPI-anchored proteins and aGPCRs.

Structure and Function of the Glycosylphosphatidylinositol Transamidase, a Transmembrane Complex Catalyzing GPI Anchoring of Proteins.

Glycosylphosphatidylinositol (GPI) anchoring of proteins is a ubiquitous posttranslational modification in eukaryotic cells. GPI-anchored proteins (GPI-APs) play critical roles in enzymatic, signaling, regulatory, and adhesion processes. Over 20 enzymes are involved in GPI synthesis, attachment to client proteins, and remodeling after attachment. The GPI transamidase (GPI-T), a large complex located in the endoplasmic reticulum membrane, catalyzes the attachment step by replacing a C-terminal signal peptide of proproteins with GPI. In the last three decades, extensive research has been conducted on the mechanism of the transamidation reaction, the components of the GPI-T complex, the role of each subunit, and the substrate specificity. Two recent studies have reported the three-dimensional architecture of GPI-T, which represent the first structures of the pathway. The structures provide detailed mechanisms for assembly that rationalizes previous biochemical results and subunit-dependent stability data. While the structural data confirm the catalytic role of PIGK, which likely uses a caspase-like mechanism to cleave the proproteins, they suggest that unlike previously proposed, GPAA1 is not a catalytic subunit. The structures also reveal a shared cavity for GPI binding. Somewhat unexpectedly, PIGT, a single-pass membrane protein, plays a crucial role in GPI recognition. Consistent with the assembly mechanisms and the active site architecture, most of the disease mutations occur near the active site or the subunit interfaces. Finally, the catalytic dyad is located ~22 Å away from the membrane interface of the GPI-binding site, and this architecture may confer substrate specificity through topological matching between the substrates and the elongated active site. The research conducted thus far sheds light on the intricate processes involved in GPI anchoring and paves the way for further mechanistic studies of GPI-T.

Identification of the glycosylphosphatidylinositol-specific phospholipase A2 (GPI-PLA2) that mediates GPI fatty acid remodeling in Trypanosoma brucei.

The biosynthesis of glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) in the parasitic protozoan Trypanosoma brucei involves fatty acid remodeling of the GPI precursor molecules before they are transferred to protein in the endoplasmic reticulum. The genes encoding the requisite phospholipase A2 and A1 activities for this remodeling have thus far been elusive. Here, we identify a gene, Tb927.7.6110, that encodes a protein that is both necessary and sufficient for GPI-phospholipase A2 (GPI-PLA2) activity in the procyclic form of the parasite. The predicted protein product belongs to the alkaline ceramidase, PAQR receptor, Per1, SID-1, and TMEM8 (CREST) superfamily of transmembrane hydrolase proteins and shows sequence similarity to Post-GPI-Attachment to Protein 6 (PGAP6), a GPI-PLA2 that acts after transfer of GPI precursors to protein in mammalian cells. We show the trypanosome Tb927.7.6110 GPI-PLA2 gene resides in a locus with two closely related genes Tb927.7.6150 and Tb927.7.6170, one of which (Tb927.7.6150) most likely encodes a catalytically inactive protein. The absence of GPI-PLA2 in the null mutant procyclic cells not only affected fatty acid remodeling but also reduced GPI anchor sidechain size on mature GPI-anchored procyclin glycoproteins. This reduction in GPI anchor sidechain size was reversed upon the re-addition of Tb927.7.6110 and of Tb927.7.6170, despite the latter not encoding GPI precursor GPI-PLA2 activity. Taken together, we conclude that Tb927.7.6110 encodes the GPI-PLA2 of GPI precursor fatty acid remodeling and that more work is required to assess the roles and essentiality of Tb927.7.6170 and the presumably enzymatically inactive Tb927.7.6150.

Persistent ER stress causes GPI anchor deficit to convert a GPI-anchored prion protein into pro-PrP via the ATF6-miR449c-5p-PIGV axis.

Endoplasmic reticulum (ER) stress and unfolded protein response are cells' survival strategies to thwart disruption of proteostasis. Tumor cells are continuously being challenged by ER stress. The prion protein, PrP, normally a glycosylphosphatidylinositol (GPI)-anchored protein exists as a pro-PrP retaining its GPI-peptide signal sequence in human pancreatic ductal cell adenocarcinoma (PDAC). Higher abundance of pro-PrP indicates poorer prognosis in PDAC patients. The reason why PDAC cells express pro-PrP is unknown. Here, we report that persistent ER stress causes conversion of GPI-anchored PrP to pro-PrP via a conserved ATF6-miRNA449c-5p-PIGV axis. Mouse neurons and AsPC-1, a PDAC cell line, express GPI-anchored PrP. However, continuous culture of these cells with the ER stress inducers thapsigargin or brefeldin A results in the conversion of a GPI-anchored PrP to pro-PrP. Such a conversion is reversible; removal of the inducers allows the cells to re-express a GPI-anchored PrP. Mechanistically, persistent ER stress increases the abundance of an active ATF6, which increases the level of miRNA449c-5p (miR449c-5p). By binding the mRNA of PIGV at its 3'-UTRs, miR449c-5p suppresses the level of PIGV, a mannosyltransferase pivotal in the synthesis of the GPI anchor. Reduction of PIGV leads to disruption of the GPI anchor assembly, causing pro-PrP accumulation and enhancing cancer cell migration and invasion. The importance of ATF6-miR449c-5p-PIGV axis is recapitulated in PDAC biopsies as the higher levels of ATF6 and miR449c-5p and lower levels of PIGV are markers of poorer outcome for patients with PDAC. Drugs targeting this axis may prevent PDAC progression.

Identification of GPI-anchored protein LYPD1 as an essential factor for odontoblast differentiation in tooth development.

Lipid rafts are membrane microdomains rich in cholesterol, sphingolipids, glycosylphosphatidylinositol-anchored proteins (GPI-APs), and receptors. These lipid raft components are localized at the plasma membrane and are essential for signal transmission and organogenesis. However, few reports have been published on the specific effects of lipid rafts on tooth development. Using microarray and single-cell RNA sequencing methods, we found that a GPI-AP, lymphocyte antigen-6/Plaur domain-containing 1 (Lypd1), was specifically expressed in preodontoblasts. Depletion of Lypd1 in tooth germ using an ex vivo organ culture system and in mouse dental pulp (mDP) cells resulted in the inhibition of odontoblast differentiation. Activation of bone morphogenetic protein (BMP) signaling by BMP2 treatment in mDP cells promoted odontoblast differentiation via phosphorylation of Smad1/5/8, while this BMP2-mediated odontoblast differentiation was inhibited by depletion of Lypd1. Furthermore, we created a deletion construct of the C terminus containing the omega site in LYPD1; this site is necessary for localizing GPI-APs to the plasma membrane and lipid rafts. We identified that this site is essential for odontoblast differentiation and morphological change of mDP cells. These findings demonstrated that LYPD1 is a novel marker of preodontoblasts in the developing tooth; in addition, they suggest that LYPD1 is important for tooth development and that it plays a pivotal role in odontoblast differentiation by regulating Smad1/5/8 phosphorylation through its effect as a GPI-AP in lipid rafts.

Towards a thorough understanding of mammalian glycosylphosphatidylinositol-anchored protein biosynthesis.

Glycosylphosphatidylinositols (GPIs) are glycolipids found ubiquitously in eukaryotes. They consist of a glycan and an inositol phospholipid, and act as membrane anchors of many cell-surface proteins by covalently linking to their C-termini. GPIs also exist as unlinked, free glycolipids on the cell surface. In human cells, at least 160 proteins with various functions are GPI-anchored proteins. Because the attachment of GPI is required for the cell-surface expression of GPI-anchored proteins, a thorough knowledge of the molecular basis of mammalian GPI-anchored protein biosynthesis is important for understanding the basic biochemistry and biology of GPI-anchored proteins and their medical significance. In this paper, I review our previous knowledge of the biosynthesis of mammalian GPI-anchored proteins and then examine new findings made since 2020.

Antibody profiles in the mosaic of "seronegative" APS syndrome.

Clinical manifestations, as distinct from thrombotic and obstetric morbidity, were recently included in the update of classification criteria of the antiphospholipid syndrome (APS). However, the existence of several patients with clinical manifestations suggestive of APS, but negative for criteria antiphospholipid antibodies (aPLs) [anti-cardiolipin antibodies (aCL), anti-ß2-glycoprotein I antibodies (aß2-GPI) and lupus anticoagulant] may suggest an update of diagnostic criteria. In this study, we analyzed the prevalence of six non-criteria aPLs in a large monocentric cohort of patients with seronegative APS (SN-APS), to investigate their possible diagnostic role. aCL IgA, aß2-GPI IgA and aß2-GPI Domain 1 antibodies were detected by chemiluminescence, anti-phosphatidylserine/prothrombin (aPS/PT) IgG, anti-vimentin/cardiolipin (aVim/CL) IgG and anti-carbamylated-ß2-glycoprotein I (aCarb-ß2-GPI) IgG by ELISA in sera from 144 SN-APS patients. In SN-APS patients, aCL IgA were detected in 4/144 (2.77%), aß2-GPI IgA in 2/144 (1.39%), aß2-GPI-Domain 1 in 1/144 (0.69%), aPS/PT in 16/144 (11.11%), aVim/CL in 37/144 (25.69%) and aCarb-ß2-GPI in 43/144 patients (29.86%). Patients negative for all non-criteria aPL assays were 77/144 (53.47%). Notably, the Venn diagram showed that aCarb-ß2-GPI together with aVim/CL represented the prevalent combination of positive antibodies. In SN-APS patients, aCL IgA were associated with recurrent thrombosis (OR11.48; p=0.03); in obstetric SN-APS patients, aPS/PT were significantly associated with foetal deaths (OR4.84; p=0.01), aVim/CL with spontaneous abortions (OR2.71; p=0.016). This study indicates that aPS/PT, aVim/CL and aCarb-ß2-GPI antibodies may represent useful tools to identify "seronegative" APS patients, who are negative for criteria aPLs, supporting the need to make testing for non-criteria aPLs more accessible in patients with SN-APS.

Missense variant in PIGM associated with severe cystic encephalomalacia and portal vein thrombosis: Phenotypic and genotypic expansion of the glycosylphosphatidylinositol biosynthesis defect-1 (GPIBD1).

Glycosylphosphatidylinositols (GPIs) are a type of glycolipid responsible for anchoring many important proteins to the cell membrane surface. Defects in the synthesis of GPIs can lead to a group of multisystem disorders known as the inherited GPI deficiencies (IGDs). Homozygosity for the c.-270C > G variant in the promoter of PIGM has been associated with a IGD subtype known as glycosylphosphatidylinositol biosynthesis defect-1 (GPIBD1). The several cases reported in the literature have been described to have a milder neurologic phenotype in comparison to the other IGDs and have been treated with sodium phenylbutyrate with some degree of success. These patients typically present with portal and hepatic vein thrombosis and mostly develop absence seizures. Here we describe a patient homozygous for a nonsynonymous variant in PIGM who deceased at 9 weeks of life and had multiple physical dysmorphisms (rocker bottom feet, midline cleft palate, thickened and lichenified skin), portal vein thrombosis, CNS structural anomalies (progressive multicystic encephalomalacia and ventriculomegaly), and a neurological phenotype of a diffuse encephalopathy. This is the first known case report of a PIGM-related IGD/CDG due to a coding variant.

Ethanolamine-phosphate on the second mannose is a preferential bridge for some GPI-anchored proteins.

Glycosylphosphatidylinositols (GPIs) are glycolipids that anchor many proteins (GPI-APs) on the cell surface. The core glycan of GPI precursor has three mannoses, which in mammals, are all modified by ethanolamine-phosphate (EthN-P). It is postulated that EthN-P on the third mannose (EthN-P-Man3) is the bridge between GPI and the protein and the second (EthN-P-Man2) is removed after GPI-protein attachment. However, EthN-P-Man2 may not be always transient, as mutations of PIGG, the enzyme that transfers EthN-P to Man2, result in inherited GPI deficiencies (IGDs), characterized by neuronal dysfunctions. Here, we show that EthN-P on Man2 is the preferential bridge in some GPI-APs, among them, the Ect-5'-nucleotidase and Netrin G2. We find that CD59, a GPI-AP, is attached via EthN-P-Man2 both in PIGB-knockout cells, in which GPI lacks Man3, and with a small fraction in wild-type cells. Our findings modify the current view of GPI anchoring and provide a mechanistic basis for IGDs caused by PIGG mutations.

Candida auris from the Egyptian cobra: Role of snakes as potential reservoirs.

Candida auris represents one of the most urgent threats to public health, although its ecology remains largely unknown. Because amphibians and reptiles may present favorable conditions for C. auris colonization, cloacal and blood samples (n = 68), from several snake species, were cultured and molecularly screened for C. auris using molecular amplification of glycosylphosphatidylinositol protein-encoding genes and ribosomal internal transcribed spacer sequencing. Candida auris was isolated from the cloacal swab of one Egyptian cobra (Naja haje legionis) and molecularly identified in its cloaca and blood. The isolation of C. auris from wild animals is herein reported for the first time, thus suggesting the role that these animals could play as reservoirs of this emerging pathogen. The occurrence of C. auris in blood requires further investigation, although the presence of cationic antimicrobial peptides in the plasma of reptiles could play a role in reducing the vitality of the fungus.

Candida auris represents one of the most urgent threats to public health. In this study, we reported for the first time the isolation of C. auris from snake thus suggesting the role of these animals as reservoirs of this emerging pathogen.

Establishment of ELISA-comparable moderate and high thresholds for anticardiolipin and anti-ß2 glycoprotein I chemiluminescent immunoassays according to the 2023 ACR/EULAR APS classification criteria and evaluation of their diagnostic performance.

OBJECTIVES: Recently published 2023 ACR/EULAR APS classification criteria emphasize the importance of quantifying single-, double-, and triple-antiphospholipid antibody positivity, distinguishing between IgG and IgM isotypes, and delineating moderate/high levels of anticardiolipin (aCL) and anti-ß2 glycoprotein I (anti-ß2GPI) antibodies. We aimed to establish clinically important moderate/high thresholds for aCL and anti-ß2GPI IgG/IgM chemiluminescent immunoassays (CLIA), in particular QUANTA Flash, comparable to our in-house ELISAs used for over two decades, and to evaluate their diagnostic performance. METHODS: QUANTA Flash CLIA and in-house ELISAs were used to measure aCL and anti-ß2GPI IgG/IgM. Moderate thresholds for QUANTA Flash CLIA were determined using a non-parametric approach, calculating a 99th percentile on serum samples from 139 blood donors, and by mirroring the diagnostic performance of in-house ELISA on 159 patient samples. RESULTS: Thresholds for QUANTA Flash CLIA achieving diagnostic performance equivalent to in-house ELISAs were 40 CU for moderate and 80 CU for high levels for aCL and anti-ß2GPI IgG and IgM. The assays showed good qualitative agreement, ranging from 76.10 to 91.19 %. When considering in-house ELISA results, 14 out of 80 (17.5 %) patients did not fulfill the new ACR/EULAR laboratory classification criteria, while 27 out of 80 (33.8 %) did not when considering QUANTA Flash CLIA results. CONCLUSIONS: We determined moderate and high thresholds for aCL and anti-ß2GPI IgG and IgM detected with QUANTA Flash CLIA, aligning with long-established in-house ELISA thresholds. These thresholds are crucial for seamlessly integrating of the new 2023 ACR/EULAR classification criteria into future observational clinical studies and trials.

Deep brain stimulation of globus pallidus internus and subthalamic nucleus in Parkinson's disease: a multicenter, retrospective study of efficacy and safety.

BACKGROUND: Deep brain stimulation (DBS) is an established therapeutic option in advanced Parkinson's disease (PD). Literature data and recent guidelines remain inconclusive about the best choice as a target between the subthalamic nucleus (STN) and the globus pallidus internus (GPi). MATERIALS AND METHODS: We retrospectively reviewed the clinical efficacy outcomes of 48 DBS-implanted patients (33 STN-DBS and 15 GPi-DBS) at a short- (<1 year from the surgery) and long-term (2-5 years) follow-up. Also, clinical safety outcomes, including postoperative surgical complications and severe side effects, were collected. RESULTS: We found no difference between STN-DBS and GPi-DBS in improving motor symptoms at short-term evaluation. However, STN-DBS achieved a more prominent reduction in oral therapy (L-DOPA equivalent daily dose, P = .02). By contrast, GPi-DBS was superior in ameliorating motor fluctuations and dyskinesia (MDS-UPDRS IV, P < .001) as well as motor experiences of daily living (MDS-UPDRS II, P = .03). The greater efficacy of GPi-DBS on motor fluctuations and experiences of daily living was also present at the long-term follow-up. We observed five serious adverse events, including two suicides, all among STN-DBS patients. CONCLUSION: Both STN-DBS and GPi-DBS are effective in improving motor symptoms severity and complications, but GPi-DBS has a greater impact on motor fluctuations and motor experiences of daily living. These results suggest that the two targets should be considered equivalent in motor efficacy, with GPi-DBS as a valuable option in patients with prominent motor complications. The occurrence of suicides in STN-treated patients claims further attention in target selection.

Inherited glycosylphosphatidylinositol deficiency: a review from molecular and clinical perspectives.

Glycosylphosphatidylinositol (GPI) is a highly conserved post-translational modification in eukaryotes, which is essential for anchoring various proteins to the cell surface. Dysfunction of GPI biogenesis leads to human diseases, such as inherited GPI deficiency (IGD) caused by germline mutations in GPI-related genes. With accumulating reports on individuals with IGD, there has been increasing interest and studies on disease mechanism, diagnosis, and therapy. This review outlines the biosynthetic pathway of GPI-anchored proteins (GPI-APs) and summarizes clinical IGD cases from a molecular perspective. We also review current diagnostic and therapeutic approaches for IGD. Finally, we discuss future research directions to facilitate the understanding and treatment of GPI-related disorders.

A CRISPR-Cas9-engineered mouse model for GPI-anchor deficiency mirrors human phenotypes and exhibits hippocampal synaptic dysfunctions.

Pathogenic germline mutations in PIGV lead to glycosylphosphatidylinositol biosynthesis deficiency (GPIBD). Individuals with pathogenic biallelic mutations in genes of the glycosylphosphatidylinositol (GPI)-anchor pathway exhibit cognitive impairments, motor delay, and often epilepsy. Thus far, the pathophysiology underlying the disease remains unclear, and suitable rodent models that mirror all symptoms observed in human patients have not been available. Therefore, we used CRISPR-Cas9 to introduce the most prevalent hypomorphic missense mutation in European patients, Pigv:c.1022C > A (p.A341E), at a site that is conserved in mice. Mirroring the human pathology, mutant Pigv341E mice exhibited deficits in motor coordination, cognitive impairments, and alterations in sociability and sleep patterns, as well as increased seizure susceptibility. Furthermore, immunohistochemistry revealed reduced synaptophysin immunoreactivity in Pigv341E mice, and electrophysiology recordings showed decreased hippocampal synaptic transmission that could underlie impaired memory formation. In single-cell RNA sequencing, Pigv341E-hippocampal cells exhibited changes in gene expression, most prominently in a subtype of microglia and subicular neurons. A significant reduction in Abl1 transcript levels in several cell clusters suggested a link to the signaling pathway of GPI-anchored ephrins. We also observed elevated levels of Hdc transcripts, which might affect histamine metabolism with consequences for circadian rhythm. This mouse model will not only open the doors to further investigation into the pathophysiology of GPIBD, but will also deepen our understanding of the role of GPI-anchor-related pathways in brain development.