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FICD activity and AMPylation remodelling modulate human neurogenesis

ABSTRACT: The AMPylation is novel protein post-translation modification. In consist of attachment of the adenosine monophosphate onto the serine, threonine or tyrosine amino acid residues of the protein. This process is catalysed by bacterial effectors or in human by protein FICD (also called HYPE) which contain conserved fic domain. In our study we have focused on identification of the unknown AMPylated proteins and their function in human cells (HeLa, A549, SH-SY5Y, iPSCs, NPCs and neurons) and cerebral organoids (COs). For this purpose we designed and synthesized the small molecule N6-propargyl phosphoramidate adenosine (pro-N6pA) which was used for in situ labelling of the AMPylation in cells and COs. Subsequnetly the the propargyl tag was employed for preparation of chemical-proteomic samples which were measured by LC-MS/MS. This approach confirmed the known AMPylation of heat shock protein HSPA5 and found diverse group of novel AMPylation targets. Several cathepsins found as AMPylation targets were further characterized by identification of AMP binding site. Based on our chemical proteomic data we found out that neurons and neuronal like cells contain distinct AMPylatino pattern from other studied cell types. This also suggested the importance of the AMPylation in these cell types which were then studied in cerebral organoids by overexpressing the FICD wt and E234G highly active mutant. This led to the accelerated differentiation of progenitors into the neurons. Overall our approach is suitable for identification of the AMPylated proteins in living cells and led to characterization of FICD function.

INSTRUMENT(S): Orbitrap Fusion, Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture, Fibroblast

SUBMITTER: Pavel Kielkowski

LAB HEAD: Stephan A. Sieber

PROVIDER: PXD015062 | Pride | 2019-12-04

REPOSITORIES: Pride

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FICD activity and AMPylation remodelling modulate human neurogenesis.

Kielkowski Pavel P Buchsbaum Isabel Y IY Kirsch Volker C VC Bach Nina C NC Drukker Micha M Cappello Silvia S Sieber Stephan A SA

Nature communications 20200124 1

Posttranslational modification (PTM) of proteins represents an important cellular mechanism for controlling diverse functions such as signalling, localisation or protein-protein interactions. AMPylation (also termed adenylylation) has recently been discovered as a prevalent PTM for regulating protein activity. In human cells AMPylation has been exclusively studied with the FICD protein. Here we investigate the role of AMPylation in human neurogenesis by introducing a cell-permeable propargyl ade ...[more]

PMID: 31980631

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