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Microtubule-lipid crosstalk during neuronal cytokinesis

Projectomschrijving

Erfelijke ziekten waarbij de hersenen van een kind onvoldoende groeien veroorzaken geestelijke en vaak ook lichamelijke beperkingen. Een van de oorzaken van deze zogenaamde congenitale microcephalieën is dat de deling van zenuwcelvoorlopers niet goed verloopt. Hierdoor vormen zich te weinig zenuwcellen en blijven de hersenen dus klein. Het is bekend dat het celskelet een belangrijke rol speelt bij de celdeling. Wij hebben een nieuw eiwit gevonden, dat gemuteerd is in een klein aantal kinderen met ernstige microcephalie. Dit enzym is belangrijk voor de celdeling en regelt de omzetting van bepaalde vetten in de celmembraan. Wij willen de functie van dit enzym in de hersenontwikkeling bestuderen en hoe het celskelet en membraanvetten van de cel met elkaar communiceren zodat de zenuwceldeling goed verloopt. Door ons onderzoek wordt een nieuw aspect van de celdeling in kaart gebracht, waardoor mogelijk ook andere microcephalie-gerelateerde genen kunnen worden geïdentificeerd.

Producten

Titel: Loss of ER and nuclear envelope-associated neutral sphingomyelinase SMPD4 causes a severe neurodevelopmental disorder with microcephaly and congenital arthrogryposis
Auteur: Maarten Fornerod
Titel: Proximity Ligation Mapping of Microcephaly Associated SMPD4 Shows Association with Components of the Nuclear Pore Membrane
Auteur: Alexandra C. A. Piët, Marco Post, Dick Dekkers, Jeroen A. A. Demmers, Maarten Fornerod
Magazine: Cells
Titel: Loss of SMPD4 Causes a Developmental Disorder Characterized by Microcephaly and Congenital Arthrogryposis
Auteur: Magini, P. Smits, D. J. Vandervore, L. Schot, R. Columbaro, M. Kasteleijn, E. van der Ent, M. Palombo, F. Lequin, M. H. Dremmen, M. de Wit, M. C. Y. Severino, M. Divizia, M. T. Striano, P. Ordonez-Herrera, N. Alhashem, A. Al Fares, A. Al Ghamdi, M. Rolfs, A. Bauer, P. Demmers, J. Verheijen, F. W. Wilke, M. van Slegtenhorst, M. van der Spek, P. J. Seri, M. Jansen, A. C. Stottmann, R. W. Hufnagel, R. B. Hopkin, R. J. Aljeaid, D. Wiszniewski, W. Gawlinski, P. Laure-Kamionowska, M. Alkuraya, F. S. A
Magazine: American Journal of Human Genetics
Titel: Generating Membrane Curvature at the Nuclear Pore: A Lipid Point of View
Auteur: Bas W A Peeters, Alexandra C A Piët, Maarten Fornerod
Magazine: Cells

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Samenvatting van de aanvraag

Mendelian malformations of cortical development (MCD) encompass a wide spectrum of disorders with various underlying genetic aetiologies and clinical manifestations, including microcephaly, or smaller brain. A significant number of mutations that give rise to microcephaly are in genes (e.g. KIF2A) encoding microtubule (MT) associated proteins (MAPs) involved in spindle organization during mitosis and other MT-related cell cycle processes. These results suggest that defects in (a)symmetric neuronal mitosis might underlie the formation of a smaller brain. Cytokinesis is the last stage of mitosis during which the cytoplasm of a single cell is partitioned into two daughter cells by cleavage furrow ingression and membrane abscission. Cytokinesis is governed by the midbody, a dense network of MTs, MAPs, and other components. Interestingly, many MAPs mutated in microcephaly localize to the midbody suggesting that these proteins are involved in MT-based events during cytokinesis. We study MAPs called CLASPs, which are major MT growth promoters. Our interest in cytokinesis was sparked by the observations that CLASPs interact with KIF2A and co-localize with KIF2A at the midbody (1), that CLASP1 was shown to regulate cytokinesis (2), and that knock down of KIF2A results in strong cytokinesis defects with overt MT stabilization (3). Thus, proper regulation of midbody MTs is required for succesful completion of cytokinesis. Recently, we identified a mutation in the SMPD4 gene, which appears to encode a neutral sphingomyelinase (see below), in four individuals from a large consanguineous pedigree presenting with severe primary microcephaly and simplified gyral pattern, small cerebellum and brain stem and global hypomyelination. Within our collaborative international network two additional independent families were subsequently identified with microcephaly and SMPD4 mutations. These results show that SMPD4 regulates neuronal proliferation and possibly differentiation. All SMPD4 patients are characterized by severe microcephaly, lack of any psychomotor development, congenital arthrogryposis and distal limb underdevelopment. Patients often die in the first few months, but some survive into childhood, suggesting the presence of some kind of stochastic determinant of threshold enzyme activity. Strikingly, SMPD4 was identified in a genetic screen for lipid metabolizing genes required for cytokinesis. Knock-down of SMPD4 caused significant delay during metaphase in mitosis, followed by failure in cytokenesis. In this study, significant increases in total and midbody ceramide and hexosyl-ceramide levels were detected, which is difficult to reconcile with a sphingomyelinase activity of SMPD4 (of which ceramides are expected to be the reaction products). Nevertheless, the unexpected finding of a lipid metabolizing enzyme being required for cytokinesis and normal neurocortical development provides us with a unique opportunity to obtain insight into MT-lipid crosstalk during neuronal cytokinesis. As both lipid composition and MT behaviour at the midbody need to be carefully orchestrated for succesful completion of cytokinesis, some form of crosstalk between these systems must exist. Our working hypothesis is that a cytokinesis checkpoint monitors whether lipids and MTs evolve in a timely fashion, and that cytokinesis is delayed when one of the two systems lags behind. The first objective of this proposal is to examine MT-lipid crosstalk and the existence of a checkpoint during neuronal cytokinesis. An unsolved question is the molecular function of the SMPD4 protein. Studies leading to the identification of the SMPD4 gene were based on purification of a protein fraction with sphingomyelinase activity. However, both SMPD4 knockdown experiments and the Smpd4 knock-out mice show accumulation of sphingolipids which are supposed to be the product of sphingomyelinase activity (i.e. ceramides) or substrates of an enzyme with different catalytic activity (i.e. glucosylceramidase). The second aim of this proposal is to investigate the function of SMPD4 in lipid metabolism, by using a combination of metabolomics, proteomics and biochemistry. An Smpd4 knock-out mouse model mimics several of the phenotypes of SMPD4 patients. Interestingly, both patients and knockout mice show substantial perinatal mortality, however, survival is also observed. The third aim of this proposal is to examine stochastic events that lead to cell-to-cell variation of SMPD4 activity. We will mainly use single cell transcriptomics to tackle this issue. Finally, we aim to investigate the function of SMPD4 in neuronal differentiation.

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Kenmerken

Projectnummer:
91217045
Looptijd: 100%
Looptijd: 100 %
2018
2023
Onderdeel van programma:
Gerelateerde subsidieronde:
Projectleider en penvoerder:
prof. dr. ir. N.J. Galjart
Verantwoordelijke organisatie:
Erasmus Medisch Centrum