Lysosome-related biomarkers in preeclampsia and cancers: Machine learning and bioinformatics analysis.

BACKGROUND: Lysosomes serve as regulatory hubs, and play a pivotal role in human diseases. However, the precise functions and mechanisms of action of lysosome-related genes remain unclear in preeclampsia and cancers. This study aimed to identify lysosome-related biomarkers in preeclampsia, and further explore the biomarkers shared between preeclampsia and cancers. MATERIALS AND METHODS: We obtained GSE60438 and GSE75010 datasets from the Gene Expression Omnibus database, pre-procesed them and merged them into a training cohort. The limma package in R was used to identify the differentially expressed mRNAs between the preeclampsia and normal control groups. Differentially expressed lysosome-related genes were identified by intersecting the differentially expressed mRNAs and lysosome-related genes obtained from Gene Ontology and GSEA databases. Gene Ontology annotations and Kyoto Encyclopedia of Genes and Genomes enrichment analysis were performed using the DAVID database. The CIBERSORT method was used to analyze immune cell infiltration. Weighted gene co-expression analyses and three machine learning algorithm were used to identify lysosome-related diagnostic biomarkers. Lysosome-related diagnostic biomarkers were further validated in the testing cohort GSE25906. Nomogram diagnostic models for preeclampsia were constructed. In addition, pan-cancer analysis of lysosome-related diagnostic biomarkers were identified by was performed using the TIMER, Sangebox and TISIDB databases. Finally, the Drug-Gene Interaction, TheMarker and DSigDB Databases were used for drug-gene interactions analysis. RESULTS: A total of 11 differentially expressed lysosome-related genes were identified between the preeclampsia and control groups. Three molecular clusters connected to lysosome were identified, and enrichment analysis demonstrated their strong relevance to the development and progression of preeclampsia. Immune infiltration analysis revealed significant immunity heterogeneity among different clusters. GBA, OCRL, TLR7 and HEXB were identified as lysosome-related diagnostic biomarkers with high AUC values, and validated in the testing cohort GSE25906. Nomogram, calibration curve, and decision curve analysis confirmed the accuracy of predicting the occurrence of preeclampsia based on OCRL and HEXB. Pan-cancer analysis showed that GBA, OCRL, TLR7 and HEXB were associated with the prognosis of patients with various tumors and tumor immune cell infiltration. Twelve drugs were identified as potential drugs for the treatment of preeclampsia and cancers. CONCLUSION: This study identified GBA, OCRL, TLR7 and HEXB as potential lysosome-related diagnostic biomarkers shared between preeclampsia and cancers.

Common genetic risk for Parkinson's disease and dysfunction of the endo-lysosomal system.

Parkinson's disease is a progressive neurological disorder, characterized by prominent movement dysfunction. The past two decades have seen a rapid expansion of our understanding of the genetic basis of Parkinson's, initially through the identification of monogenic forms and, more recently, through genome-wide association studies identifying common risk variants. Intriguingly, a number of cellular pathways have emerged from these analysis as playing central roles in the aetiopathogenesis of Parkinson's. In this review, the impact of data deriving from genome-wide analyses for Parkinson's upon our functional understanding of the disease will be examined, with a particular focus on examples of endo-lysosomal and mitochondrial dysfunction. The challenges of moving from a genetic to a functional understanding of common risk variants for Parkinson's will be discussed, with a final consideration of the current state of the genetic architecture of the disorder. This article is part of a discussion meeting issue 'Understanding the endo-lysosomal network in neurodegeneration'.

Lysosomal storage, impaired autophagy and innate immunity in Gaucher and Parkinson's diseases: insights for drug discovery.

Impairment of autophagic-lysosomal pathways is increasingly being implicated in Parkinson's disease (PD). GBA1 mutations cause the lysosomal storage disorder Gaucher disease (GD) and are the commonest known genetic risk factor for PD. GBA1 mutations have been shown to cause autophagic-lysosomal impairment. Defective autophagic degradation of unwanted cellular constituents is associated with several pathologies, including loss of normal protein homeostasis, particularly of α-synuclein, and innate immune dysfunction. The latter is observed both peripherally and centrally in PD and GD. Here, we will discuss the mechanistic links between autophagy and immune dysregulation, and the possible role of these pathologies in communication between the gut and brain in these disorders. Recent work in a fly model of neuronopathic GD (nGD) revealed intestinal autophagic defects leading to gastrointestinal dysfunction and immune activation. Rapamycin treatment partially reversed the autophagic block and reduced immune activity, in association with increased survival and improved locomotor performance. Alterations in the gut microbiome are a critical driver of neuroinflammation, and studies have revealed that eradication of the microbiome in nGD fly and mouse models of PD ameliorate brain inflammation. Following these observations, lysosomal-autophagic pathways, innate immune signalling and microbiome dysbiosis are discussed as potential therapeutic targets in PD and GD. This article is part of a discussion meeting issue 'Understanding the endo-lysosomal network in neurodegeneration'.

Endolysosomal trafficking controls yolk granule biogenesis in vitellogenic Drosophila oocytes.

Endocytosis and endolysosomal trafficking are essential for almost all aspects of physiological functions of eukaryotic cells. As our understanding on these membrane trafficking events are mostly from studies in yeast and cultured mammalian cells, one challenge is to systematically evaluate the findings from these cell-based studies in multicellular organisms under physiological settings. One potentially valuable in vivo system to address this challenge is the vitellogenic oocyte in Drosophila, which undergoes extensive endocytosis by Yolkless (Yl), a low-density lipoprotein receptor (LDLR), to uptake extracellular lipoproteins into oocytes and package them into a specialized lysosome, the yolk granule, for storage and usage during later development. However, by now there is still a lack of sufficient understanding on the molecular and cellular processes that control yolk granule biogenesis. Here, by creating genome-tagging lines for Yl receptor and analyzing its distribution in vitellogenic oocytes, we observed a close association of different endosomal structures with distinct phosphoinositides and actin cytoskeleton dynamics. We further showed that Rab5 and Rab11, but surprisingly not Rab4 and Rab7, are essential for yolk granules biogenesis. Instead, we uncovered evidence for a potential role of Rab7 in actin regulation and observed a notable overlap of Rab4 and Rab7, two Rab GTPases that have long been proposed to have distinct spatial distribution and functional roles during endolysosomal trafficking. Through a small-scale RNA interference (RNAi) screen on a set of reported Rab5 effectors, we showed that yolk granule biogenesis largely follows the canonical endolysosomal trafficking and maturation processes. Further, the data suggest that the RAVE/V-ATPase complexes function upstream of or in parallel with Rab7, and are involved in earlier stages of endosomal trafficking events. Together, our study provides s novel insights into endolysosomal pathways and establishes vitellogenic oocyte in Drosophila as an excellent in vivo model for dissecting the highly complex membrane trafficking events in metazoan.

The GATOR2 complex maintains lysosomal-autophagic function by inhibiting the protein degradation of MiT/TFEs.

Lysosomes are central to metabolic homeostasis. The microphthalmia bHLH-LZ transcription factors (MiT/TFEs) family members MITF, TFEB, and TFE3 promote the transcription of lysosomal and autophagic genes and are often deregulated in cancer. Here, we show that the GATOR2 complex, an activator of the metabolic regulator TORC1, maintains lysosomal function by protecting MiT/TFEs from proteasomal degradation independent of TORC1, GATOR1, and the RAG GTPase. We determine that in GATOR2 knockout HeLa cells, members of the MiT/TFEs family are ubiquitylated by a trio of E3 ligases and are degraded, resulting in lysosome dysfunction. Additionally, we demonstrate that GATOR2 protects MiT/TFE proteins in pancreatic ductal adenocarcinoma and Xp11 translocation renal cell carcinoma, two cancers that are driven by MiT/TFE hyperactivation. In summary, we find that the GATOR2 complex has independent roles in TORC1 regulation and MiT/TFE protein protection and thus is central to coordinating cellular metabolism with control of the lysosomal-autophagic system.

The Parkinson's disease related mutant VPS35 (D620N) amplifies the LRRK2 response to endolysosomal stress.

The identification of multiple genes linked to Parkinson's disease (PD) invites the question as to how they may co-operate. We have generated isogenic cell lines that inducibly express either wild-type or a mutant form of the retromer component VPS35 (D620N), which has been linked to PD. This has enabled us to test proposed effects of this mutation in a setting where the relative expression reflects the physiological occurrence. We confirm that this mutation compromises VPS35 association with the WASH complex, but find no defect in WASH recruitment to endosomes, nor in the distribution of lysosomal receptors, cation-independent mannose-6-phosphate receptor and Sortilin. We show VPS35 (D620N) enhances the activity of the Parkinson's associated kinase LRRK2 towards RAB12 under basal conditions. Furthermore, VPS35 (D620N) amplifies the LRRK2 response to endolysosomal stress resulting in enhanced phosphorylation of RABs 10 and 12. By comparing different types of endolysosomal stresses such as the ionophore nigericin and the membranolytic agent l-leucyl-l-leucine methyl ester, we are able to dissociate phospho-RAB accumulation from membrane rupture.

Characterization of Novel Human ß-glucocerebrosidase Antibodies for Parkinson's Disease Research.

BACKGROUND: Mutations in GBA1, which encodes the lysosome enzyme ß-glucocerebrosidase (also referred to as acid ß-glucosidase or GCase), are the most common genetic risk factor for Parkinson's disease (PD) and dementia with Lewy bodies (DLB). Evidence also suggests that loss of GCase activity is implicated in PD without GBA1 mutations. Consequently, therapies targeting GCase are actively being pursued as potential strategies to modify the progression of PD and related synucleinopathies. Despite this significant interest in GCase as a therapeutic target, the lack of well-characterized GCase antibodies continues to impede progress in the development of GCase-targeted therapies. OBJECTIVE: This study aims to independently evaluate human GCase (hGCase) antibodies to provide recommendations for western blot, immunofluorescence, immunoprecipitation, and AlphaLISA (Amplified Luminescent Proximity Homogeneous Assay) assays. METHODS: Two mouse monoclonal antibodies, hGCase-1/17 and hGCase-1/23, were raised against hGCase using imiglucerase, the recombinant enzyme developed to treat patients, as the antigen. These novel antibodies, alongside commonly used antibodies in the field, underwent evaluation in a variety of assays. RESULTS: The characterization of hGCase-1/17 and hGCase-1/23 using genetic models including GBA1 loss-of-function human neuroglioma H4 line and neurons differentiated from human embryonic stem cells revealed their remarkable specificity and potency in immunofluorescence and immunoprecipitation assays. Furthermore, a hGCase AlphaLISA assay with excellent sensitivity, a broad dynamic range, and suitability for high throughput applications was developed using hGCase-1/17 and hGCase-1/23, which enabled a sandwich assay configuration. CONCLUSIONS: The hGCase immunofluorescence, immunoprecipitation, and AlphaLISA assays utilizing hGCase-1/17 and hGCase-1/23 will not only facilitate improved investigations of hGCase biology, but can also serve as tools to assess the distribution and effectiveness of GCase-targeted therapies for PD and related synucleinopathies.

Development and validation of a novel lysosome-related LncRNA signature for predicting prognosis and the immune landscape features in colon cancer.

Lysosomes are essential components for managing tumor microenvironment and regulating tumor growth. Moreover, recent studies have also demonstrated that long non-coding RNAs could be used as a clinical biomarker for diagnosis and treatment of colorectal cancer. However, the influence of lysosome-related lncRNA (LRLs) on the progression of colon cancer is still unclear. This study aimed to identify a prognostic LRL signature in colon cancer and elucidated potential biological function. Herein, 10 differential expressed lysosome-related genes were obtained by the TCGA database and ultimately 4 prognostic LRLs for conducting a risk model were identified by the co-expression, univariate cox, least absolute shrinkage and selection operator analyses. Kaplan-Meier analysis, principal-component analysis, functional enrichment annotation, and nomogram were used to verify the risk model. Besides, the association between the prognostic model and immune infiltration, chemotherapeutic drugs sensitivity were also discussed in this study. This risk model based on the LRLs may be promising for potential clinical prognosis and immunotherapeutic responses related indicator in colon cancer patients.

Screening of four lysosome-related genes in sepsis based on RNA sequencing technology.

PURPOSE: Screening of lysosome-related genes in sepsis patients to provide direction for lysosome-targeted therapy. METHODS: Peripheral blood samples were obtained from 22 patients diagnosed with sepsis and 10 normal controls for the purpose of RNA sequencing and subsequent analysis of differential gene expression. Concurrently, lysosome-related genes were acquired from the Gene Ontology database. The intersecting genes between the differential genes and lysosome-related genes were then subjected to PPI, GO and KEGG analyses. Core genes were identified through survival analysis, and their expression trends in different groups were determined using meta-analysis. Single-cell RNA sequencing was used to clarify the cellular localization of core genes. RESULTS: The intersection of 1328 sepsis-differential genes with 878 lysosome-related genes yielded 76 genes. PPI analysis showed that intersecting genes were mainly involved in Cellular process, Response to stimulus, Immune system process, Signal transduction, Lysosome. GO and KEGG analysis showed that intersecting genes were mainly involved in leukocyte mediated immunity, cell activation involved in immune response, lytic vacuole, lysosome. Survival analysis screened four genes positively correlated with sepsis prognosis, namely GNLY, GZMB, PRF1 and RASGRP1. The meta-analysis revealed that the expression levels of these four genes were significantly higher in the normal control group compared to the sepsis group, which aligns with the findings from RNA sequencing data. Furthermore, single-cell RNA sequencing demonstrated that T cells and NK cells exhibited high expression levels of GNLY, GZMB, PRF1, and RASGRP1. CONCLUSION: GNLY, GZMB, PRF1, and RASGRP1, which are lysosome-related genes, are closely linked to the prognosis of sepsis and could potentially serve as novel research targets for sepsis, offering valuable insights for the development of lysosome-targeted therapy. The clinical trial registration number is ChiCTR1900021261, and the registration date is February 4, 2019.

Erbin accelerates TFEB-mediated lysosome biogenesis and autophagy and alleviates sepsis-induced inflammatory responses and organ injuries.

Mounting attention has been focused on defects of the autophagy-lysosomal pathway in sepsis, however, the precise mechanisms governing the autophagy-lysosomal process in sepsis are poorly known. We have previously reported that Erbin deficiency aggravated the inflammatory response and organ injuries caused by sepsis. In the present study, we found that Erbin knockout impaired the autophagy process in both muramyl dipeptide (MDP)-induced bone marrow-derived macrophages (BMDMs) and sepsis mouse liver and lung, as detected by the accumulation of LC3-II and SQSTM1/p62, and autophagosomes. Pretreatment with autophagy inhibitor chloroquine (CQ) further aggravated inflammatory response and organ injuries in vivo and in vitro sepsis model. We also observed that the impaired lysosomal function mediated autophagic blockade, as detected by the decreased expression of ATP6V, cathepsin B (CTSB) and LAMP2 protein. Immunoprecipitation revealed that the C-terminal of Erbin (aa 391-964) interacts with the N-terminal of transcription factor EB (TFEB) (aa 1-247), and affects the stability of TFEB-14-3-3 and TFEB-PPP3CB complexes and the phosphorylation status of TFEB, thereby promote the nucleus translocation of TFEB and the TFEB target genes transcription. Thus, our study suggested that Erbin alleviated sepsis-induced inflammatory responses and organ injuries by rescuing dysfunction of the autophagy-lysosomal pathway through TFEB-14-3-3 and TFEB-PPP3CB pathway.

Construction and validation of a novel lysosomal signature for hepatocellular carcinoma prognosis, diagnosis, and therapeutic decision-making.

Lysosomes is a well-recognized oncogenic driver and chemoresistance across variable cancer types, and has been associated with tumor invasiveness, metastasis, and poor prognosis. However, the significance of lysosomes in hepatocellular carcinoma (HCC) is not well understood. Lysosomes-related genes (LRGs) were downloaded from Genome Enrichment Analysis (GSEA) databases. Lysosome-related risk score (LRRS), including eight LRGs, was constructed via expression difference analysis (DEGs), univariate and LASSO-penalized Cox regression algorithm based on the TCGA cohort, while the ICGC cohort was obtained for signature validation. Based on GSE149614 Single-cell RNA sequencing data, model gene expression and liver tumor niche were further analyzed. Moreover, the functional enrichments, tumor microenvironment (TME), and genomic variation landscape between LRRSlow/LRRShigh subgroup were systematically investigated. A total of 15 Lysosomes-related differentially expressed genes (DELRGs) in HCC were detected, and then 10 prognosis DELRGs were screened out. Finally, the 8 optimal DELRGs (CLN3, GBA, CTSA, BSG, APLN, SORT1, ANXA2, and LAPTM4B) were selected to construct the LRRS prognosis signature of HCC. LRRS was considered as an independent prognostic factor and was associated with advanced clinicopathological features. LRRS also proved to be a potential marker for HCC diagnosis, especially for early-stage HCC. Then, a nomogram integrating the LRRS and clinical parameters was set up displaying great prognostic predictive performance. Moreover, patients with high LRRS showed higher tumor stemness, higher heterogeneity, and higher genomic alteration status than those in the low LRRS group and enriched in metabolism-related pathways, suggesting its underlying role in the progression and development of liver cancer. Meanwhile, the LRRS can affect the proportion of immunosuppressive cell infiltration, making it a vital immunosuppressive factor in the tumor microenvironment. Additionally, HCC patients with low LRRS were more sensitive to immunotherapy, while patients in the high LRRS group responded better to chemotherapy. Upon single-cell RNA sequencing, CLN3, GBA, and LAPTM4B were found to be specially expressed in hepatocytes, where they promoted cell progression. Finally, RT-qPCR and external datasets confirmed the mRNA expression levels of model genes. This study provided a direct links between LRRS signature and clinical characteristics, tumor microenvironment, and clinical drug-response, highlighting the critical role of lysosome in the development and treatment resistance of liver cancer, providing valuable insights into the prognosis prediction and treatment response of HCC, thereby providing valuable insights into prognostic prediction, early diagnosis, and therapeutic response of HCC.

New tools can propel research in lysosomal storage diseases.

Historically, the clinical manifestations of lysosomal storage diseases offered an early glimpse into the essential digestive functions of the lysosome. However, it was only recently that the more subtle role of this organelle in the dynamic regulation of multiple cellular processes was appreciated. With the need for precise interrogation of lysosomal interplay in health and disease comes the demand for more sophisticated functional tools. This demand has recently been met with 1) induced pluripotent stem cell-derived models that recapitulate the disease phenotype in vitro, 2) methods for lysosome affinity purification coupled with downstream omics analysis that provide a high-resolution snapshot of lysosomal alterations, and 3) gene editing and CRISPR/Cas9-based functional genomic strategies that enable screening for genetic modifiers of the disease phenotype. These emerging methods have garnered much interest in the field of neurodegeneration, and their use in the field of metabolic disorders is now also steadily gaining momentum. Looking forward, these robust tools should accelerate basic science efforts to understand lysosomal dysfunction distal to substrate accumulation and provide translational opportunities to identify disease-modifying therapies.

Lysosome damage triggers direct ATG8 conjugation and ATG2 engagement via non-canonical autophagy.

Cells harness multiple pathways to maintain lysosome integrity, a central homeostatic process. Damaged lysosomes can be repaired or targeted for degradation by lysophagy, a selective autophagy process involving ATG8/LC3. Here, we describe a parallel ATG8/LC3 response to lysosome damage, mechanistically distinct from lysophagy. Using a comprehensive series of biochemical, pharmacological, and genetic approaches, we show that lysosome damage induces non-canonical autophagy and Conjugation of ATG8s to Single Membranes (CASM). Following damage, ATG8s are rapidly and directly conjugated onto lysosome membranes, independently of ATG13/WIPI2, lipidating to PS (and PE), a molecular hallmark of CASM. Lysosome damage drives V-ATPase V0-V1 association, direct recruitment of ATG16L1 via its WD40-domain/K490A, and is sensitive to Salmonella SopF. Lysosome damage-induced CASM is associated with formation of dynamic, LC3A-positive tubules, and promotes robust LC3A engagement with ATG2, a lipid transfer protein central to lysosome repair. Together, our data identify direct ATG8 conjugation as a rapid response to lysosome damage, with important links to lipid transfer and dynamics.

Distinct sets of lysosomal genes define synucleinopathy and tauopathy.

Neurodegenerative diseases are characterized by distinct protein aggregates, such as those of α-synuclein and tau. Lysosomal defect is a key contributor to the accumulation and propagation of aberrant protein aggregates in these diseases. The discoveries of common proteinopathies in multiple forms of lysosomal storage diseases (LSDs) and the identification of some LSD genes as susceptible genes for those proteinopathies suggest causative links between LSDs and the proteinopathies. The present study hypothesized that defects in lysosomal genes will differentially affect the propagation of α-synuclein and tau proteins, thereby determining the progression of a specific proteinopathy. We established an imaging-based high-contents screening (HCS) system in Caenorhabditis elegans (C. elegans) model, by which the propagation of α-synuclein or tau is measured by fluorescence intensity. Using this system, we performed RNA interference (RNAi) screening to induce a wide range of lysosomal malfunction through knock down of 79 LSD genes, and to obtain the candidate genes with significant change in protein propagation. While some LSD genes commonly affected both α-synuclein and tau propagation, our study identified the distinct sets of LSD genes that differentially regulate the propagation of either α-synuclein or tau. The specificity and efficacy of these LSD genes were retained in the disease-related phenotypes, such as pharyngeal pumping behavior and life span. This study suggests that distinct lysosomal genes differentially regulate the propagation of α-synuclein and tau, and offer a steppingstone to understanding disease specificity. [BMB Reports 2023; 56(12): 657-662].

[Electroacupuncture at "Baihui"（GV20） and "Yongquan"（KI1） improves learning-memory ability of APP/PS1 double transgenic mice by regulating endosomal-lysosomal system].

OBJECTIVE: To explore the mechanism of electroacupuncture（EA） in improving learning-memory ability in Alzheimer's disease （AD） mice from the perspective of endosomal-lysosomal system. METHODS: Male APP/PS1 transgenic mice were randomly divided into model group and EA group （n=10 in each group） and 10 male C57BL/6 wild mice were taken as the normal group. EA （1 Hz/50 Hz, 1 mA） was applied at bilateral "Yongquan"（KI1） and acupuncture was applied at "Baihui" （GV20） for 15 min. The mice of the model and normal groups were subjected to restriction with the same method as those of the EA group for 15 min. The treatment was conducted once every other day for 6 weeks. The spatial learning-memory ability （shown by escape latency of place navigation test and the time of crossing the target platform and total swimming distance in the target quadrant in 1 min of spatial probe test ） was detected by Morris water maze test. The immunoactivity of senile plaques （SP） in the hippocampus tissue was detected by immunohistochemistry. The ultrastructural characters of hippocampal neurons were observed by transmission electron microscope, and the expression levels of Ras-related protein 5 （Rab5）, Ras-related protein 7 （Rab7） and cathepsin D （CTSD） in the hippocampus were detected by Western blot, separately. RESULTS: Compared with the normal group, the escape latency, SP immunoactivity, and protein expression levels of Rab5, Rab7 and CTSD were significantly increased （P<0.05, P<0.01）, while the number of crossing the original platform and the total swimming distance in the platform quadrant were considerably reduced （P<0.05） in the model group. In contrast to the model group, the EA group had a marked decrease in the escape latency, SP immunoactivity, and protein expression levels of Rab5, Rab7 and CTSD （P<0.05, P<0.01）, and a striking increase in the number of crossing the original platform and the swimming distance in the platform quadrant （P<0.05）. Results of transmission electron microscope showed an accumulation of endosome, lysosome, and endolysosomes in the hippocampal neurons in the model group, which was evidently milder in the EA group. CONCLUSION: EA of GV20 and KI1 can improve the learning-memory ability of AD mice, which may be related to its function in reducing hippocampal Aß deposition and down-regulating endosomal-lysosomal system activity.

Altered endosomal-lysosomal biogenesis in melanoma.

Cutaneous melanoma is the deadliest form of skin neoplasm and its high mortality rates could be averted by early accurate detection. While the detection of melanoma is currently reliant upon melanin visualisation, research into melanosome biogenesis, as a key driver of pathogenesis, has not yielded technology that can reliably distinguish between atypical benign, amelanotic and melanotic lesions. The endosomal-lysosomal system has important regulatory roles in cancer cell biology, including a specific functional role in melanosome biogenesis. Herein, the involvement of the endosomal-lysosomal system in melanoma was examined by pooled secondary analysis of existing gene expression datasets. A set of differentially expressed endosomal-lysosomal genes was identified in melanoma, which were interconnected by biological function. To illustrate the protein expression of the dysregulated genes, immunohistochemistry was performed on samples from patients with cutaneous melanoma to reveal candidate markers. This study demonstrated the dysregulation of Syntenin-1, Sortilin and Rab25 may provide a differentiating feature between cutaneous melanoma and squamous cell carcinoma, while IGF2R may indicate malignant propensity in these skin cancers.

Genetic Evidence for Endolysosomal Dysfunction in Parkinson's Disease: A Critical Overview.

Parkinson's disease (PD) is the second most common neurodegenerative disorder in the aging population, and no disease-modifying therapy has been approved to date. The pathogenesis of PD has been related to many dysfunctional cellular mechanisms, however, most of its monogenic forms are caused by pathogenic variants in genes involved in endolysosomal function (LRRK2, VPS35, VPS13C, and ATP13A2) and synaptic vesicle trafficking (SNCA, RAB39B, SYNJ1, and DNAJC6). Moreover, an extensive search for PD risk variants revealed strong risk variants in several lysosomal genes (e.g., GBA1, SMPD1, TMEM175, and SCARB2) highlighting the key role of lysosomal dysfunction in PD pathogenesis. Furthermore, large genetic studies revealed that PD status is associated with the overall "lysosomal genetic burden", namely the cumulative effect of strong and weak risk variants affecting lysosomal genes. In this context, understanding the complex mechanisms of impaired vesicular trafficking and dysfunctional endolysosomes in dopaminergic neurons of PD patients is a fundamental step to identifying precise therapeutic targets and developing effective drugs to modify the neurodegenerative process in PD.

Updates on the study of lysosomal protein dynamics: possibilities for the clinic.

INTRODUCTION: The lysosome is the main degradative organelle of almost all mammalian cells, fulfilling important functions in macromolecule recycling, metabolism, and signaling. Lysosomal dysfunction is connected to a continuously growing number of pathologic conditions, and lysosomal proteins present potential biomarkers for a variety of diseases. Therefore, there is an increasing interest in their analysis in patient samples. AREAS COVERED: We provide an overview of OMICs studies which identified lysosomal proteins as potential biomarkers for pathological conditions, covering proteomics, genomics, and transcriptomics approaches, identified through PubMed searches. With respect to discovery proteomics analyses, mainly lysosomal luminal and associated proteins were detected, while membrane proteins were found less frequently. Comprehensive coverage of the lysosomal proteome was only achieved by ultra-deep-coverage studies, but targeted approaches allowed for the reproducible quantification of lysosomal proteins in diverse sample types. EXPERT OPINION: The low abundance of lysosomal proteins complicates their reproducible analysis in patient samples. Whole proteome shotgun analyses fail in many instances to cover the lysosomal proteome, which is due to under-sampling and/or a lack of sensitivity. With the current state of the art, targeted proteomics assays provide the best performance for the characterization of lysosomal proteins in patient samples.

A central role for regulated protein stability in the control of TFE3 and MITF by nutrients.

The TFE3 and MITF master transcription factors maintain metabolic homeostasis by regulating lysosomal, melanocytic, and autophagy genes. Previous studies posited that their cytosolic retention by 14-3-3, mediated by the Rag GTPases-mTORC1, was key for suppressing transcriptional activity in the presence of nutrients. Here, we demonstrate using mammalian cells that regulated protein stability plays a fundamental role in their control. Amino acids promote the recruitment of TFE3 and MITF to the lysosomal surface via the Rag GTPases, activating an evolutionarily conserved phospho-degron and leading to ubiquitination by CUL1ß-TrCP and degradation. Elucidation of the minimal functional degron revealed a conserved alpha-helix required for interaction with RagA, illuminating the molecular basis for a severe neurodevelopmental syndrome caused by missense mutations in TFE3 within the RagA-TFE3 interface. Additionally, the phospho-degron is recurrently lost in TFE3 genomic translocations that cause kidney cancer. Therefore, two divergent pathologies converge on the loss of protein stability regulation by nutrients.

TPC Functions in the Immune System.

Two-pore channels (TPCs) are novel intracellular cation channels, which play a key role in numerous (patho-)physiological and immunological processes. In this chapter, we focus on their function in immune cells and immune reactions. Therefore, we first give an overview of the cellular immune response and the partaking immune cells. Second, we concentrate on ion channels which in the past have been shown to play an important role in the regulation of immune cells. The main focus is then directed to TPCs, which are primarily located in the membranes of acidic organelles, such as lysosomes or endolysosomes but also certain other vesicles. They regulate Ca2+ homeostasis and thus Ca2+ signaling in immune cells. Due to this important functional role, TPCs are enjoying increasing attention within the field of immunology in the last few decades but are also becoming more pertinent as pharmacological targets for the treatment of pro-inflammatory diseases such as allergic hypersensitivity. However, to uncover the precise molecular mechanism of TPCs in immune cell responses, further molecular, genetic, and ultrastructural investigations on TPCs are necessary, which then may pave the way to develop novel therapeutic strategies to treat diseases such as anaphylaxis more specifically.