Biology subjects

Dutto, I.

Publications and source records attributed to Dutto, I..

3 recordsLinked to original sources

bioRxiv2020

SAICAr-dependent and independent effects of ADSL deficiency on neurodevelopment

Adenylosuccinate Lyase (ADSL) functions in the de novo purine biosynthesis pathway. ADSL deficiency (ADSLD) causes numerous neurodevelopmental pathologies, including microcephaly and autism spectrum disorder. ADSLD patients have normal purine nucleotide levels but exhibit accumulation of the dephosphorylated ADSL substrates SAICAr and S-Ado. SAICAr was implicated in the neurotoxic effects of ADSLD, although its role remains unknown. We examined the effects of ADSL depletion in human cells and found increased DNA damage signaling, that was rescued by nucleosides, and impaired primary ciliogenesis, that was rescued by reducing SAICAr. By analyzing ADSL deficient chicken and zebrafish embryos we observed impaired neurogenesis and microcephaly, and neuroprogenitor attrition in zebrafish was rescued by reducing SAICAr. Zebrafish embryos also displayed phenotypes commonly linked to ciliopathies. Our results suggest that both reduced purine levels and SAICAr accumulation contribute to neurodevelopmental pathology in ADSLD and defective ciliogenesis may influence the ADSLD phenotypic spectrum.

developmental biology↗

bioRxiv2020

Sub-centrosomal mapping identifies augmin-γTuRC as part of a centriole-stabilizing scaffold

Centriole biogenesis and maintenance are crucial for cells to generate cilia and assemble centrosomes that function as microtubule organizing centers (MTOCs). Centriole biogenesis and MTOC function both require the microtubule nucleator {gamma}-tubulin ring complex ({gamma}TuRC). The widely accepted view is that {gamma}TuRC localizes to the pericentriolar material (PCM), where it nucleates microtubules. {gamma}TuRC has also been observed at centriolar regions that lack PCM, but the significance of these findings is unclear. Here we have used expansion microscopy to map spatially and functionally distinct sub-populations of centrosomal {gamma}TuRC including in the centriole lumen. Luminal localization is mediated by augmin and both complexes are linked to the centriole inner scaffold through POC5. Disruption of luminal localization impairs centriole stability and cilia assembly, defects that are also observed in {gamma}TuRC mutant fibroblasts derived from a patient suffering from microcephaly with chorioretinopathy. These results identify a novel, non-canonical role of augmin-{gamma}TuRC in the centriole lumen that is linked to human disease.

cell biology↗

bioRxiv2020

Dynamics of protein binding to sites of nascent unscheduled DNA repair synthesis in non-proliferating cells

The analysis of DNA repair mechanisms is of fundamental importance to understand how cells remove DNA damage and maintain their genome stability. Investigating the dynamic association of proteins at sites of active DNA synthesis has been successfully performed at DNA replication forks, providing important information on the process, and allowing the identification of new players acting at these sites. However, the applicability of these studies to DNA repair events at sites of nascent unscheduled DNA synthesis (UDS) in non-proliferating cells has been never tested. Here, we describe the analysis of dynamics association of protein participating in nucleotide excision repair (NER), and in other DNA repair processes, at sites of nascent UDS in non-proliferating cells, to avoid interference by DNA replication. Labeling with 5-ethynyl-2-deoxyuridine (EdU) after DNA damage, followed by click reaction to biotinylate these sites, permits the analysis of dynamic association of proteins, such as DNA polymerases {delta} and {kappa}, as well as PCNA, to active DNA repair synthesis sites. The suitability of this technique to identify new factors present at active UDS sites is illustrated by two examples of proteins previously unknown to participate in the UV-induced DNA repair process.

biochemistry↗