Identification of PPT1 substrates to elucidate the mechanisms of neurodegeneration in Neuronal Ceroid Lipofuscinosis

Identification: Gorenberg, Erica


Identification of PPT1 substrates to elucidate the mechanisms of neurodegeneration in Neuronal Ceroid Lipofuscinosis
Erica Gorenberg1, Vicky Chou2, Gregory Wirak2, TuKiet Lam4 and Sreeganga Chandra2,3
Interdepartmental Neuroscience Program1, Department of Neurology2, Department of Neuroscience3, Yale/NIDA Neuroproteomics Center4, Yale University, New Haven, CT 06511
Neuronal ceroid lipofuscinoses (NCLs) are a family of genetically inherited neurodegenerative diseases with lysosomal pathology. Loss of function mutations in the palmitoyl protein thioesterase 1 gene (PPT1; also known as CLN1) gene cause NCL. The severity of the CLN1 mutations is correlated with the age of disease onset as well as its progression, with a total loss of PPT1 activity leading to infantile NCL. PPT1 encodes a depalmitoylating enzyme which participates in the dynamic lipid modification of proteins. Palmitoylation is the covalent attachment of a 16-carbon fatty acid chain to cysteine residues on proteins. Palmitate groups are attached to proteins by protein acyl transferases, and removed by protein thioesterases, such as PPT1, which breaks the thioester link between the palmitate and the protein. In neurons, PPT1 is enriched at synapses, and its dysfunction leads to aberrant increases in palmitoylation and synaptic trafficking deficits. Despite a clear role for palmitoylation dynamics in neurodegenerative disease, the repertoire of PPT1 substrates are unknown, creating a knowledge deficit in our understanding of NCL and potential therapies. To identify PPT1 substrates, we purified palmitoylated proteins from wild type and PPT1 knockout (KO) synaptosomes and compared the palmitomes using Label Free Quantification-Mass Spectroscopy. We identified putative PPT1 substrates as proteins whose levels of palmitoylation are increased in the KO and validated select substrates using orthologous methods. We also mapped and characterized the synaptic pathways most affected by PPT1 KO. Our results reveal the critical roles PPT1 plays in synapse function, and protein palmitoylation deficits in PPT1-linked NCL. Our results are broadly relevant to other neurodegenerative diseases where aberrant palmitoylation has been noted such as Huntington's disease, Alzheimer's disease and amyotrophic lateral sclerosis.


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