Three-dimensional organization of HP1C-complex target genes
- Autores: Marta Puerto Plasencia
- Directores de la Tesis: Ferran Azorín Marín (dir. tes.), Pedro Martínez Serra (tut. tes.)
- Lectura: En la Universitat de Barcelona ( España ) en 2022
- Idioma: inglés
- Tribunal Calificador de la Tesis: Montserrat (Corominas Guiu) Corominas (presid.), Jordi Bernués Martínez (secret.), Eulàlia de Nadal Clanchet (voc.)
- Programa de doctorado: Programa de Doctorado en Genética por la Universidad de Barcelona
- Materias:
- Enlaces
- Tesis en acceso abierto en: TESEO
- Resumen
It has been previously shown that the Drosophila heterochromatin protein 1c (HP1c) forms a complex with the transcription factors (TF) WOC and ROW, and the extraproteasomal ubiquitin receptor dDsk2. The HP1c-complex localizes at the transcription start site (TSS) of active genes and regulates RNApol II pausing. Previous work also showed that the HP1c-complex interacts with the architectural proteins Z4, Chromator and BEAF-32, which are involved in the regulation of the three-dimensional (3D) organization of the genome inside the nucleus. In this work, we have analyzed the 3D organization of HP1c-complex target genes and studied the contribution of the architectural protein Z4. HiChIP experiments performed with αROW antibodies show that HP1c-complex target genes are extensively involved in 3D interactions with non-target genes, forming clusters of high connectivity (HCRROW). Genes within these clusters are expressed and, although the average expression of genes in different HCRROW clusters is similar, dispersion of the expression levels of genes within each cluster is lower than respect to the genes in the rest of clusters, suggesting that gene expression within HCRROW clusters is coordinated. Our results also show that the architectural protein Z4 stabilizes 3D organization of HP1c-complex target genes since its depletion reduces αROW HiChIP interactions. Unexpectedly, we observed that Z4 has an opposite effect when 3D interactions are studied by whole-genome HiC. We show that 3D interactions of HP1c-complex target regions, which decreased intensity in αROW HiChIP experiments upon Z4 depletion, show increased intensity in HiC experiments. These results suggest that, in contrast to HiC experiments that interrogate 3D interactions in the whole population of cells, αROW HiChIP experiments capture the interactions in only a minor subpopulation, in which ROW is actually bound. Therefore, while Z4 stabilizes the interactions when ROW is bound, it has the opposite effect when ROW is not bound. Further analysis of HiC data showed that Z4 depletion results in a general increase of short-range 3D interactions, including interactions between active (A) compartments, gene loop interactions and Polycomb (Pc) loop interactions. Similar effects where observed in Z4-depleted cells subjected to hyperosmotic stress. We observed that, while in control cells hyperosmotic stress causes a general loss of 3D interactions that swiftly recover after stress release, in Z4-depleted cells 3D interactions were more resistant to hyperosmotic stress and recovered to a higher intensity upon stress release. Interestingly, we also observed that Z4 antagonizes pairing between homologous chromosomes, as cells lacking Z4 show increased pairing. A similar behavior was reported earlier for the condensin subunit CapH2. As a matter of fact, co-IP experiments provide support for the interaction between Z4 and CapH2. Moreover, ChIP-seq experiments show that Z4 extensively co-localizes with CapH2 and that CapH2 chromatin binding depends on Z4. These results suggest that Z4 and CapH2 co-operate to prevent chromosome pairing. Notably, we observed that hyperosmotic stress abolishes chromosome pairing. Altogether these results suggest a link between chromosome pairing and 3D interactions since conditions that favor pairing (ie, Z4 depletion) reinforce 3D interactions, while conditions that inhibit pairing (ie, hyperosmotic stress) weaken 3D interactions.
