Life in the lumen: The multivesicular endosome.

The late endosomes/endo-lysosomes of vertebrates contain an atypical phospholipid, lysobisphosphatidic acid (LBPA) (also termed bis[monoacylglycero]phosphate [BMP]), which is not detected elsewhere in the cell. LBPA is abundant in the membrane system present in the lumen of this compartment, including intralumenal vesicles (ILVs). In this review, the current knowledge on LBPA and LBPA-containing membranes will be summarized, and their role in the control of endosomal cholesterol will be outlined. Some speculations will also be made on how this system may be overwhelmed in the cholesterol storage disorder Niemann-Pick C. Then, the roles of intralumenal membranes in endo-lysosomal dynamics and functions will be discussed in broader terms. Likewise, the mechanisms that drive the biogenesis of intralumenal membranes, including ESCRTs, will also be discussed, as well as their diverse composition and fate, including degradation in lysosomes and secretion as exosomes. This review will also discuss how intralumenal membranes are hijacked by pathogenic agents during intoxication and infection, and what is the biochemical composition and function of the intra-endosomal lumenal milieu. Finally, this review will allude to the size limitations imposed on intralumenal vesicle functions and speculate on the possible role of LBPA as calcium chelator in the acidic calcium stores of endo-lysosomes.

Development of Organelle Replacement Therapy Using a Stearyl-Polyhistidine Peptide against Lysosomal Storage Disease Cells.

We previously reported on a polyhistidine peptide, His16 peptide, as a new cell-penetrating peptide. This peptide is anticipated to be a new carrier for drug delivery systems (DDSs) for targeting intracellular lysosomes because it can transport macromolecules (e.g., liposomes) into these organelles. In the present study, we examined the application of His16 peptide as a DDS carrier against lysosomal storage disease (LSD) cells. LSDs are metabolic disorders caused by loss of specific lysosomal enzymes. For the treatment of LSD cells, we devised a system designated organelle replacement therapy (ORT). ORT is a strategy for transporting exogenous lysosomes containing all kinds of lysosomal enzymes from normal cells into endogenous lysosomes in LSD cells using His16 peptide. To develop the ORT system, we prepared His16 peptide-modified healthy lysosomes (His16-Lyso) by insertion of a stearyl-His16 peptide into a hydrophobic region in the lysosomal membrane. His16-Lyso showed cellular uptake and localization to endogenous lysosomes in LSD cells. His16-Lyso also restored the proliferation of LSD cells, which otherwise showed slower proliferation than normal cells. These results suggested that His16-Lyso replenished deficient lysosomal enzymes in LSD cells. The results further suggest that His16-Lyso are promising candidates as a treatment tool for LSD cells and to establish a foundation for ORT.

Patient with Fabry disease undergoing cardiac surgery: a word of caution.

Fabry disease (FD) is a rare genetic disorder that affects various organs and systems in the body. The disease is caused by a deficiency in the lysosomal enzyme α-galactosidase A (AGAL), which leads to the accumulation of globotriaosylceramide (Gb3) within lysosomes. This accumulation can cause damage to cells and organ systems, leading to a wide range of symptoms and complications. FD is a heterogeneous disorder, with a wide range of clinical phenotypes, ranging from the classic form, which is severe and associated with early onset, to milder non-classical forms, which are often limited to one organ and manifest later in life. We describe the case of a 23-year-old FD patient who was admitted as an emergency transfer due to newly discovered severe aortic regurgitation and suspected aortic valve endocarditis with vegetations of high embolic potential. Three years ago, the patient underwent a living donor kidney transplantation-the kidney graft lost its function 1 year after transplantation, and a chronic hemodialysis program was reinstituted. Supplementary Information: The online version contains supplementary material available at 10.1007/s12055-024-01717-6.

Presynaptic Endosomal Cathepsin D Regulates the Biogenesis of GABAergic Synaptic Vesicles.

Presynaptic endosomes reportedly participate in synaptic vesicle (SV) recycling. However, it remains unclear whether they differentially regulate SV biogenesis and synaptic transmission in different types of synapses and how they are implicated in diseases. Using cryo-electron tomography and endocytic tracing, we uncover different endocytic modes and dynamics associated with distinct SV morphology between glutamatergic and GABAergic synapses. We further find that cathepsin D (CatD), a lysosomal storage disease (LSD) protein, is selectively located in GABAergic presynaptic endosomes. Inactivation of CatD results in enlarged presynaptic endosomes, reduces the readily releasable pool, and impairs synaptic transmission in GABAergic, but not glutamatergic, synapses. Moreover, CatD-deficient mice exhibit hyperactivity and increased sensitivity to seizure, mimicking epileptic behavior in CatD-related LSD patients. These data reveal an important role for presynaptic endosomal CatD in regulating GABAergic SV biogenesis and provide mechanistic insights for understanding the synaptic pathology and behavioral defects in CatD-associated LSD.

Progranulin associates with hexosaminidase A and ameliorates GM2 ganglioside accumulation and lysosomal storage in Tay-Sachs disease.

Tay-Sachs disease (TSD) is a lethal lysosomal storage disease (LSD) caused by mutations in the HexA gene, which can lead to deficiency of ß-hexosaminidase A (HexA) activity and consequent accumulation of its substrate, GM2 ganglioside. Recent reports that progranulin (PGRN) functions as a chaperone of lysosomal enzymes and its deficiency is associated with LSDs, including Gaucher disease and neuronal ceroid lipofuscinosis, prompted us to screen the effects of recombinant PGRN on lysosomal storage in fibroblasts from 11 patients affected by various LSDs, which led to the isolation of TSD in which PGRN demonstrated the best effects in reducing lysosomal storage. Subsequent in vivo studies revealed significant GM2 accumulation and the existence of typical TSD cells containing zebra bodies in both aged and ovalbumin-challenged adult PGRN-deficient mice. In addition, HexA, but not HexB, was aggregated in PGRN-deficient cells. Furthermore, recombinant PGRN significantly reduced GM2 accumulation and lysosomal storage in these animal models. Mechanistic studies indicated that PGRN bound to HexA through granulins G and E domain and increased the enzymatic activity and lysosomal delivery of HexA. More importantly, Pcgin, an engineered PGRN derivative bearing the granulin E domain, also effectively bound to HexA and reduced the GM2 accumulation. Collectively, these studies not only provide new insights into the pathogenesis of TSD but may also have implications for developing PGRN-based therapy for this life-threatening disorder. KEY MESSAGES: GM2 accumulation and the existence of typical TSD cells containing zebra bodies are detected in both aged and ovalbumin-challenged adult PGRN deficient mice. Recombinant PGRN significantly reduces GM2 accumulation and lysosomal storage both in vivo and in vitro, which works through increasing the expression and lysosomal delivery of HexA. Pcgin, an engineered PGRN derivative bearing the granulin E domain, also effectively binds to to HexA and reduces GM2 accumulation.

Defective autophagy degradation and abnormal tight junction-associated signaling drive epithelial dysfunction in cystinosis.

Cystinosis is a lysosomal storage disease due to inactivating mutations in CTNS, the cystinosin transporter that exports cystine out of lysosomes. The lysosomal accumulation of cystine leads to severe dysfunction of the epithelial cells lining the proximal tubule of the kidney, causing defective endocytosis and massive losses of solutes in the urine. The mechanisms linking lysosomal defect and epithelial dysfunction were unknown, preventing the development of disease-modifying therapies. We recently reported that lysosomal alterations in cystinosis lead to defective autophagic clearance of damaged mitochondria, generating oxidative stress. The latter destabilizes tight junctions and activates an abnormal YBX3 (Y box binding protein 3) transcriptional program driving a loss of differentiation and defective apical endocytosis in cystinosis cells. Correction of the primary lysosomal defect, neutralization of mitochondrial oxidative stress, or blockage of tight junction-associated YBX3 signaling rescue epithelial function and endocytic uptake. Our findings suggest a cascade that links lysosomal disease, defective autophagy and epithelial dysfunction, providing new perspectives for cystinosis and lysosomal storage disorders.

Trehalose upregulates progranulin expression in human and mouse models of GRN haploinsufficiency: a novel therapeutic lead to treat frontotemporal dementia.

BACKGROUND: Progranulin (PGRN) is a secreted growth factor important for neuronal survival and may do so, in part, by regulating lysosome homeostasis. Mutations in the PGRN gene (GRN) are a common cause of frontotemporal lobar degeneration (FTLD) and lead to disease through PGRN haploinsufficiency. Additionally, complete loss of PGRN in humans leads to neuronal ceroid lipofuscinosis (NCL), a lysosomal storage disease. Importantly, Grn-/- mouse models recapitulate pathogenic lysosomal features of NCL. Further, GRN variants that decrease PGRN expression increase the risk of developing Alzheimer's disease (AD) and Parkinson's disease (PD). Together these findings demonstrate that insufficient PGRN predisposes neurons to degeneration. Therefore, compounds that increase PGRN levels are potential therapeutics for multiple neurodegenerative diseases. RESULTS: Here, we performed a cell-based screen of a library of known autophagy-lysosome modulators and identified multiple novel activators of a human GRN promoter reporter including several common mTOR inhibitors and an mTOR-independent activator of autophagy, trehalose. Secondary cellular screens identified trehalose, a natural disaccharide, as the most promising lead compound because it increased endogenous PGRN in all cell lines tested and has multiple reported neuroprotective properties. Trehalose dose-dependently increased GRN mRNA as well as intracellular and secreted PGRN in both mouse and human cell lines and this effect was independent of the transcription factor EB (TFEB). Moreover, trehalose rescued PGRN deficiency in human fibroblasts and neurons derived from induced pluripotent stem cells (iPSCs) generated from GRN mutation carriers. Finally, oral administration of trehalose to Grn haploinsufficient mice significantly increased PGRN expression in the brain. CONCLUSIONS: This work reports several novel autophagy-lysosome modulators that enhance PGRN expression and identifies trehalose as a promising therapeutic for raising PGRN levels to treat multiple neurodegenerative diseases.

Differential mechanisms of action of the mucolipin synthetic agonist, ML-SA1, on insect TRPML and mammalian TRPML1.

Mucolipin synthetic agonist 1 (ML-SA1) was recently identified to activate mammalian TRPML channels and shown to alleviate lipid accumulation in lysosomes of cellular models of lysosome storage diseases, mucolipidosis type IV (MLIV) and Niemann-Pick's disease type C (NPC). Owning to its potential use in complimenting genetic studies in Drosophila melanogaster to elucidate the cellular and physiological functions of TRPML channels, we examined the effect of ML-SA1 on Drosophila TRPML expressed in HEK293 cells using whole-cell, inside-out, and whole-lysosome electrophysiological recordings. We previously showed that when expressed in HEK293 cells, Drosophila TRPML was localized and functional on both plasma membrane and endolysosome. We show here that in both inside-out patches excised from the plasma membrane and whole-lysosome recordings from enlarged endolysosome vacuoles, ML-SA1 failed to activate TRPML unless exogenous phosphatidylinositol 3,5-bisphosphate [PI(3,5)P2] was applied. At 1 µM ML-SA1, the sensitivity of TRPML to PI(3,5)P2 increased approximately by 10-fold and at 10 µM ML-SA1, the deactivation of PI(3,5)P2-evoked TRPML currents was markedly slowed. On the other hand, constitutive activation of TRPML by a mutation that mimics the varitint-waddler (Va) mutation of mouse TRPML3 rendered the insect channel sensitive to activation by ML-SA1 alone. Moreover, different from the insect TRPML, mouse TRPML1 was readily activated by ML-SA1 independent of PI(3,5)P2. Thus, our data reveal that while ML-SA1 acts as a true agonist at mouse TRPML1, it behaves as an allosteric activator of the Drosophila TRPML, showing dependence on and the ability to stabilize open conformation of the insect channels.