Role of lysosome in healing neurological disorders by nano-bioengineering.

Lysosomes primarily recognized as center for cellular 'garbage-disposing-unit', which has recently emerged as a crucial regulator of cellular metabolism. This organelle is a well-known vital player in the pathology including neurodegenerative disorders. In pathological context, removal of intracellular damaged misfolded proteins, organelles and aggregates are ensured by 'Autophagy' pathway, which initially recognizes, engulfs and seals the toxic cargo at the cytosolic environment.

Leucine retention in lysosomes is regulated by starvation.

Cells acquire essential nutrients from the environment and utilize adaptive mechanisms to survive when nutrients are scarce. How nutrients are trafficked and compartmentalized within cells and whether they are stored in response to stress remain poorly understood. Here, we investigate amino acid trafficking and uncover evidence for the lysosomal transit of numerous essential amino acids.

Lysosomal Dysfunction in Down Syndrome Is APP-Dependent and Mediated by APP-βCTF (C99).

Lysosomal failure underlies pathogenesis of numerous congenital neurodegenerative disorders and is an early and progressive feature of Alzheimer's disease (AD) pathogenesis. Here, we report that lysosomal dysfunction in Down ayndrome (trisomy 21), a neurodevelopmental disorder and form of early onset AD, requires the extra gene copy of amyloid precursor protein (APP) and is specifically mediated by the β cleaved carboxy terminal fragment of APP (APP-βCTF, C99). In primary fibroblasts from individuals with DS, lysosomal degradation of autophagic and endocytic substrates is selectively impaired, causing them to accumulate in enlarged autolysosomes/lysosomes.

PIKfyve Regulates Vacuole Maturation and Nutrient Recovery following Engulfment.

The scavenging of extracellular macromolecules by engulfment can sustain cell growth in a nutrient-depleted environment. Engulfed macromolecules are contained within vacuoles that are targeted for lysosome fusion to initiate degradation and nutrient export. We have shown that vacuoles containing engulfed material undergo mTORC1-dependent fission that redistributes degraded cargo back into the endosomal network.

LAP: the protector against autoimmunity.

The autophagy-related process LC3-Associated Phagocytosis, or LAP, is known to control the degradation of engulfed cells and microorganisms. Now Martinez et al. discover that LAP controls immune responses to dying cells and its inhibition leads to development of Systemic Lupus Erythematosus-like disease.

Absence of lipofuscin in motor neurons of SOD1-linked ALS mice.

Lipofuscin, or aging pigment, is accreted as red autofluorescence in the lysosomes of motor neuron cell bodies in the ventral horn of WT mice by 3 mo of age. Strikingly, in two presymptomatic ALS mouse strains transgenic for mutant human Cu/Zn superoxide dismutase (SOD1), G85R SOD1YFP and G93A SOD1, little or no lipofuscin was detected in motor neuron cell bodies. Two markers of autophagy, sequestosome 1 (SQSTM1/p62) and microtubule-associated protein 1 light chain 3 (LC3), were examined in the motor neuron cell bodies of G85R SOD1YFP mice and found to be reduced relative to WT SOD1YFP transgenic mice.

Production of RNA for transcriptomic analysis from mouse spinal cord motor neuron cell bodies by laser capture microdissection.

Preparation of high-quality RNA from cells of interest is critical to precise and meaningful analysis of transcriptional differences among cell types or between the same cell type in health and disease or following pharmacologic treatments. In the spinal cord, such preparation from motor neurons, the target of interest in many neurologic and neurodegenerative diseases, is complicated by the fact that motor neurons represent <10% of the total cell population. Laser capture microdissection (LMD) has been developed to address this problem.

RNA-Seq profiling of spinal cord motor neurons from a presymptomatic SOD1 ALS mouse.

Mechanisms involved with degeneration of motor neurons in amyotrophic lateral sclerosis (ALS; Lou Gehrig's Disease) are poorly understood, but genetically inherited forms, comprising ~10% of the cases, are potentially informative. Recent observations that several inherited forms of ALS involve the RNA binding proteins TDP43 and FUS raise the question as to whether RNA metabolism is generally disturbed in ALS. Here we conduct whole transcriptome profiling of motor neurons from a mouse strain, transgenic for a mutant human SOD1 (G85R SOD1-YFP), that develops symptoms of ALS and paralyzes at 5-6 months of age.

Therapeutic effects of remediating autophagy failure in a mouse model of Alzheimer disease by enhancing lysosomal proteolysis.

The extensive autophagic-lysosomal pathology in Alzheimer disease (AD) brain has revealed a major defect: in the proteolytic clearance of autophagy substrates. Autophagy failure contributes on several levels to AD pathogenesis and has become an important therapeutic target for AD and other neurodegenerative diseases. We recently observed broad therapeutic effects of stimulating autophagic-lysosomal proteolysis in the TgCRND8 mouse model of AD that exhibits defective proteolytic clearance of autophagic substrates, robust intralysosomal amyloid-β peptide (Aβ) accumulation, extracellular β-amyloid deposition and cognitive deficits.

Reversal of autophagy dysfunction in the TgCRND8 mouse model of Alzheimer's disease ameliorates amyloid pathologies and memory deficits.

Autophagy, a major degradative pathway for proteins and organelles, is essential for survival of mature neurons. Extensive autophagic-lysosomal pathology in Alzheimer's disease brain contributes to Alzheimer's disease pathogenesis, although the underlying mechanisms are not well understood. Here, we identified and characterized marked intraneuronal amyloid-β peptide/amyloid and lysosomal system pathology in the Alzheimer's disease mouse model TgCRND8 similar to that previously described in Alzheimer's disease brains.

Identification of regulators of chaperone-mediated autophagy.

Chaperone-mediated autophagy (CMA) is a selective mechanism for the degradation of cytosolic proteins in lysosomes that contributes to cellular quality control and becomes an additional source of amino acids when nutrients are scarce. A chaperone complex delivers CMA substrates to a receptor protein at the lysosomal membrane that assembles into multimeric translocation complexes. However, the mechanisms regulating this process remain, for the most part, unknown.

Induction of autophagy by cystatin C: a mechanism that protects murine primary cortical neurons and neuronal cell lines.

Cystatin C (CysC) expression in the brain is elevated in human patients with epilepsy, in animal models of neurodegenerative conditions, and in response to injury, but whether up-regulated CysC expression is a manifestation of neurodegeneration or a cellular repair response is not understood. This study demonstrates that human CysC is neuroprotective in cultures exposed to cytotoxic challenges, including nutritional-deprivation, colchicine, staurosporine, and oxidative stress. While CysC is a cysteine protease inhibitor, cathepsin B inhibition was not required for the neuroprotective action of CysC.

Entering the lysosome through a transient gate by chaperone-mediated autophagy.

A subset of cytosolic proteins can be selectively degraded in lysosomes through chaperone-mediated autophagy. The lysosomal-membrane protein type 2A (LAMP-2A) acts as the receptor for the substrates of chaperone-mediated autophagy (CMA), which should undergo unfolding before crossing the lysosomal membrane and reaching the lumen for degradation. Translocation of substrates is assisted by chaperones on both sides of the membrane, but the actual steps involved in this process and the characteristics of the translocation complex were, for the most part, unknown.

The chaperone-mediated autophagy receptor organizes in dynamic protein complexes at the lysosomal membrane.

Chaperone-mediated autophagy (CMA) is a selective type of autophagy by which specific cytosolic proteins are sent to lysosomes for degradation. Substrate proteins bind to the lysosomal membrane through the lysosome-associated membrane protein type 2A (LAMP-2A), one of the three splice variants of the lamp2 gene, and this binding is limiting for their degradation via CMA. However, the mechanisms of substrate binding and uptake remain unknown.

Altered dynamics of the lysosomal receptor for chaperone-mediated autophagy with age.

Rates of autophagy, the mechanism responsible for lysosomal clearance of cellular components, decrease with age. We have previously described an age-related decline in chaperone-mediated autophagy (CMA), a selective form of autophagy, by which particular cytosolic proteins are delivered to lysosomes after binding to the lysosome-associated membrane protein type 2A (LAMP-2A), a receptor for this pathway. Rates of CMA decrease with age because of a decrease in the levels of LAMP-2A.

Chaperone-mediated autophagy in aging and neurodegeneration: lessons from alpha-synuclein.

Different conditions, ranging from genetic mutation to post-translational modification, result in the intracellular presence of misfolded or conformationally altered proteins. These abnormal proteins tend to organize in toxic oligomeric structures often resulting in cellular death. Alterations in the function of the surveillance systems that normally repair or remove abnormal proteins are the basis of many neurodegenerative disorders.

Oxidative stress and autophagy.

Organisms respond to oxidative injury by orchestrating a stress response to prevent further damage. An increase in the intracellular levels of antioxidant agents, and at the same time the removal of already damaged components, are both part of the oxidative stress response. Lysosomes have been classically considered one of the main targets of the reactive oxygen species.