Equipe d’Accueil : Molecular Genetics and Epigenetics
Intitulé de l’Unité : Department of Mycology, Institut Pasteur
Nom du Responsable de l’Unité : Guilhem JANBON
Nom du Responsable de l’Équipe : Eugene GLADYSHEV
Adresse : 28 Rue du Dr Roux, 75015 Paris
Responsable de l’encadrement : Eugene GLADYSHEV
Tél : 0680663280 E-mail: eugene.gladyshev@pasteur.fr

Résumé du projet
Our team is interested in discovering new mechanisms responsible for the initiation of epigenetic silencing. This work is done in the fungus Neurospora crassa, for the following reasons: (1) N. crassa features all principal epigenetic pathways, including RNAi, DNA methylation, constitutive (“H3K9me3”), and facultative (“H3K27me3”) heterochromatin; (2) in these pathways, the key effectors remain strongly conserved; yet (3) they can be inactivated, individually or altogether, without causing substantial fitness defects under standard conditions; (4) at the same time, N. crassa evolved several extremely efficient genome-defense mechanisms that employ these pathways to effectuate epigenetic silencing; in addition, (5) N. crassa is a haploid microbial organism, with a relatively small genome devoid of naturally occurring repetitive DNA and gene paralogs, thus greatly simplifying the analysis; (6) lastly, N. crassa represents an established model system, which is easy to grow and manipulate in the lab. These reasons make N. crassa simply the best model organism to pursue fundamental mechanisms of epigenetic silencing that escaped their proper identification in other organisms.
One of our projects is focused on the mechanism by which a chromatin remodeler ATRX [alphathalassemia mental retardation X-linked] is able to trigger both transcriptional and post-transcriptional silencing. In humans, ATRX malfunction is linked to cancerogenesis, neuropathologies, and other disease, and a great deal of attention has been paid to ATRX in the literature; yet the results have been confusing, in part, due to the complexity of the mammalian system itself. In N. crassa, deletion of the gene encoding ATRX does not produce any apparent growth defects, yet it does inactivate the following genome-defense processes: meiotic silencing by unpaired DNA (MSUD), heterochromatinrelated pathway of repeat-induced point mutation (RIP), somatic silencing of perturbed chromatin (Carlier et al, 2024) and repeat-induced gene silencing (RIGS). How ATRX directs the initiation of both heterochromatin-mediated and RNAi-mediated silencing in such diverse processes represents the question that the proposed M2 internship is aimed to address.
This M2 project may pursue one or several themes in combination, as follows:
– dissect sequence requirements of a natural locus subject to strong ATRX-dependent transcriptiona silencing: this locus corresponds to an integrated chromovirus, present as a single copy and defined by its LTRs (which appear dispensable for silencing, thus pointing to the existence of an unknown mechanism);
– characterize new factors that work together with ATRX: this effort will (1) use mass-spec to identify proteins that co-IP with tagged ATRX and (2) establish their relevance by functional analysis;
– characterize the thresholding process that defines chromatin for ATRX-mediated silencing: here we will purify a model locus by a high-affinity method and analyze it by mass-spec.

Dernières Publications en lien avec le projet :
[Research papers]
[1] Carlier F, Castro Ramirez S, Kilani J, Chehboub S, Loïodice I, Taddei A, Gladyshev E. 2024. Remodeling of perturbed chromatin can initiate de novo transcriptional and post-transcriptional silencing. PNAS 121: e2402944121. doi: 10.1073/pnas.2402944121.
[2] Rhoades N, Nguyen TS, Witz G, Cecere G, Hammond T, Mazur AK, Gladyshev E. 2021. Recombination-independent recognition of DNA homology for meiotic silencing in Neurospora crassa. PNAS 118: e2108664118. doi: 10.1073/pnas.2108664118.
[3] Carlier F, Nguyen TS, Mazur AK, Gladyshev E. 2021. Modulation of C-to-T mutation by recombination-independent pairing of closely positioned DNA repeats. Biophys J. 120: 4325-4336. doi: 10.1016/j.bpj.2021.09.014.

[Reviews]
[4] Mazur AK, Gladyshev E. 2023. C-DNA may facilitate homologous DNA pairing. Trends Genet. 39: 575-585. doi: 10.1016/j.tig.2023.01.008.

[Preprints]
[5] Mazur AK, Maaloum M, Gladyshev E. (2024). Properties of DNA in concentrated aqueous solutions of LiCl suggest transition to C-DNA. doi: 10.1101/2024.09.20.613475. bioRxiv.

Ce projet s’inscrit dans la perspective d’une thèse
Type de financement prévu : –
Ecole Doctorale de rattachement : BioSPC