Mutazioni del gene CLN8 sono causa di varianti di lipofuscinosi ceroidee neuronali (neuronal ceroid lipofuscinoses, NCLs), un gruppo di patologie neurodegenerative autosomiche-recessive principalmente dell'infanzia, associate a geni e variabili cliniche differenti. Generalmente i sintomi comprendono un progressivo deterioramento mentale e motorio, epilessia, atassia, in alcuni casi anche perdita della visione. Si conoscono a tutt'oggi almeno 13 geni differenti implicati nelle varie forme NCLs: CLN1/PPT1, CLN2/TPP1, CLN3, CLN4/DNAJC5, CLN5, CLN6, CLN7/MFSD8, CLN8, CLN10/CTSD, CLN13/CTSF, CLN11/GRN, CLN12/ATP13A2 e CLN14/KCTD7, dei quali solo alcuni codificano proteine a funzione conosciuta (http://www.ucl.ac.uk/ncl/), mentre altri hanno funzione ignota. Ciò rappresenta ancora un grosso limite alla conoscenza della patologia e dei possibili bersagli terapeutici. Da qualche tempo, il gruppo di ricerca si occupa dello studio della funzione della proteina CLN8, la cui struttura molecolare possiede un dominio TLC (TRAM-LAG1-CLN8) di 200 amminoacidi che consente di suggerire un possibile ruolo nella regolazione della sintesi e del trasporto degli sfingolipidi (Winter and Ponting, TIBS, 2002). Gli sfingolipidi sono lipidi polari di membrana particolarmente abbondanti nelle cellule nervose, implicati nello sviluppo e plasticità del sistema nervoso e con una distribuzione spazio-temporale essenziale al mantenimento delle funzioni neuronali. Nostri studi hanno identificato una potenziale capacità della proteina CLN8 di interagire con diverse componenti proteiche cellulari che regolano meccanismi quali il trasporto di vescicole/membrane, la formazione di autofago-lisosomi di smaltimento, la degenerazione neuronale apoptotica e la sintesi e il trasporto di lipidi (Passantino et al., BBA, 2013). In particolare, l'identificazione della proteina VAPA (vesicle-associated membrane protein-associated-protein-A) tra gli interattori della CLN8 ha indirizzato la nostra proposta di studio nell'ambito del progetto finanziato dalla Fondazione Telethon (Telethon Grants 2016). La proteina VAPA ha un ruolo di "docking site" per le proteine che legano i lipidi, svolgendo una funzione determinate nel metabolismo degli sfingolipidi, e pertanto può rappresentare un partner molecolare ad hoc del CLN8 (Peretti et al., MBC, 2008). Il nostro studio intende definire le basi strutturali e funzionali dell'interazione CLN8/VAPA in associazione ad analisi analitiche di sfingolipidomica con l'obiettivo ultimo di definire la funzione della proteina CLN8 e identificare meccanismi patogenetici e bersagli specifici della patologia NCL. Lo studio verrà esteso anche su cellule di fibroblasti di pazienti con difetti in CLN8, ottenute dalla collaborazione con istituzioni in Italia, Finlandia e Germania, considerando le variabili genotipo-fenotipo.
CLN8 mutations cause clinical variant forms of neuronal ceroid lipofuscinoses (NCLs), which are a genetic heterogeneous group of autosomal recessive neurodegenerative diseases manifest mainly in childhood. Symptoms generally include progressive mental and motor deterioration, epilepsy, ataxia, in some cases also visual loss. To date, at least 13 different genes have been found characterizing the different NCLs: CLN1/PPT1, CLN2/TPP1, CLN3, CLN4/DNAJC5, CLN5, CLN6, CLN7/MFSD8, CLN8, CLN10/CTSD, CLN13/CTSF, CLN11/GRN, CLN12/ATP13A2 and CLN14/KCTD7 genes; of these, only few genes encode proteins with known function, whilst the others have undefined function (http://www.ucl.ac.uk/ncl/). This represents an obstacle to the knowledge of pathogenic features of these diseases and to the identification of precise targets for the development of successful therapies. In these last years, the research team has been involved in studying CLN8 protein, whose molecular structure possesses a TLC (TRAM-LAG1-CLN8) domain of 200 a.a. residues which may indicate a role in the synthesis and trafficking of sphigolipids (Winter and Ponting, TIBS, 2002). Sphingolipids are polar membrane lipids particularly abundant in nervous cells, involved in neural differentiation and development, and with a peculiar spatial-temporal distribution for the maintenance of neuronal functions. Our previous studies identified potential interactions of CLN8 with proteins involved in vesicular/membrane trafficking, autophagosome-lysosome formation, neuronal apoptotic death and in the synthesis and transport of lipids (Passantino et al., BBA, 2013). Among CLN8 interactors, the identification of VAPA protein (vesicle-associated membrane protein-associated-protein-A) is the starting point of our proposal in the project approved by Telethon Foundation (Telethon Grants 2016). VAPA protein plays a key role as a "docking site" for lipid-binding proteins at the endoplasmic reticulum, and so contributes to sphingolipid metabolism, being then a potential molecular partner of CLN8 (Peretti et al., MBC, 2008). Our project will develop studies to structurally and functionally define CLN8/VAPA interaction along with a sphingolipidomic profile. The final objective of these studies is to provide a comprehensive understanding of CLN8 function, bringing to a closer definition of pathomechanisms and therapeutic targets of CLN8 diseases. In the project, investigations will be extended to fibroblast cells from patients with CLN8-associated diseases provided by collaborations with institutions in Italy, Finland and Germany, in order to sort out genotype-phenotype differences.
Targeting lipids in CLN8-associated NCL diseases: structural and functional interaction of CLN8 with Vesicle-Associated Membrane Protein-associated Protein A (VAPA), and genotype-phenotype correlations
Patrizia Guarneri
2017
Abstract
CLN8 mutations cause clinical variant forms of neuronal ceroid lipofuscinoses (NCLs), which are a genetic heterogeneous group of autosomal recessive neurodegenerative diseases manifest mainly in childhood. Symptoms generally include progressive mental and motor deterioration, epilepsy, ataxia, in some cases also visual loss. To date, at least 13 different genes have been found characterizing the different NCLs: CLN1/PPT1, CLN2/TPP1, CLN3, CLN4/DNAJC5, CLN5, CLN6, CLN7/MFSD8, CLN8, CLN10/CTSD, CLN13/CTSF, CLN11/GRN, CLN12/ATP13A2 and CLN14/KCTD7 genes; of these, only few genes encode proteins with known function, whilst the others have undefined function (http://www.ucl.ac.uk/ncl/). This represents an obstacle to the knowledge of pathogenic features of these diseases and to the identification of precise targets for the development of successful therapies. In these last years, the research team has been involved in studying CLN8 protein, whose molecular structure possesses a TLC (TRAM-LAG1-CLN8) domain of 200 a.a. residues which may indicate a role in the synthesis and trafficking of sphigolipids (Winter and Ponting, TIBS, 2002). Sphingolipids are polar membrane lipids particularly abundant in nervous cells, involved in neural differentiation and development, and with a peculiar spatial-temporal distribution for the maintenance of neuronal functions. Our previous studies identified potential interactions of CLN8 with proteins involved in vesicular/membrane trafficking, autophagosome-lysosome formation, neuronal apoptotic death and in the synthesis and transport of lipids (Passantino et al., BBA, 2013). Among CLN8 interactors, the identification of VAPA protein (vesicle-associated membrane protein-associated-protein-A) is the starting point of our proposal in the project approved by Telethon Foundation (Telethon Grants 2016). VAPA protein plays a key role as a "docking site" for lipid-binding proteins at the endoplasmic reticulum, and so contributes to sphingolipid metabolism, being then a potential molecular partner of CLN8 (Peretti et al., MBC, 2008). Our project will develop studies to structurally and functionally define CLN8/VAPA interaction along with a sphingolipidomic profile. The final objective of these studies is to provide a comprehensive understanding of CLN8 function, bringing to a closer definition of pathomechanisms and therapeutic targets of CLN8 diseases. In the project, investigations will be extended to fibroblast cells from patients with CLN8-associated diseases provided by collaborations with institutions in Italy, Finland and Germany, in order to sort out genotype-phenotype differences.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
