Polycomb Repressive Complexes 1 and 2 (PRC1 and PRC2) control cell identity by establishing facultative heterochromatin repressive domains at common sets of target genes, from the early stages of development to adulthood. How they functionally preserve transcriptional silencing has been deeply explored in embryonic stem cells (ESC), but remains poorly described in an in vivo context. Taking advantage of Cre-dependent conditional knockout mouse models targeting distinct Polycomb activities in the intestinal epithelium, the present work proposes to characterize the interplay among the PRC1 subcomplexes and their relationship with PRC2 activity. In particular, it functionally dissects how PRC1 and PRC2 cooperate or have independent roles to establish the epigenetic signature of intestinal cells. Since catalytic inactivation of PRC1 (RING1A/B double knockout) affects intestinal homeostasis by exhausting the stem cell pool, we have ablated single PCGF activities (PRC1.1-1.6) to determine the role of variant (vPRC1) and canonical PRC1 (cPRC1) subcomplexes in the transcriptional identity of intestinal epithelial cells. By coupling in vivo studies with genome-wide analyses, we have found that none of them reproduces the effects of RING1A/B loss of function, resulting in the compensation of H2AK119ub1 deposition and gene repression. Moreover, we have evaluated the dependency of PRC2 function on the downstream cPRC1 activity, demonstrating that PRC2 acts autonomously to control transcriptional silencing and secretory lineage commitment. Unexpectedly, we have found that removal of PRC1.1 rescued the effects of PRC2 inactivation in lineage skewing while maintaining extensive transcriptional derepression, revealing an intimate connection between distinct Polycomb activities in the control of intestinal cell plasticity and identity. Finally, by validating in parallel some of these in vivo findings in the mouse ESC model, we have also observed that promiscuous interactions between vPRC1 and cPRC1 occur, identifying a novel mechanism of functional compensation among Polycomb subcomplexes.

CROSSTALK BETWEEN PRC1 AND PRC2 IN INTESTINAL LINEAGE COMMITMENT AND CELL IDENTITY / S. Amato ; internal advisor: S. Campaner ; external advisor: G. Gargiulo ; supervisor: D. Pasini. Dipartimento di Scienze della Salute, 2023 Apr 13. 34. ciclo, Anno Accademico 2022.

CROSSTALK BETWEEN PRC1 AND PRC2 IN INTESTINAL LINEAGE COMMITMENT AND CELL IDENTITY

S. Amato
2023

Abstract

Polycomb Repressive Complexes 1 and 2 (PRC1 and PRC2) control cell identity by establishing facultative heterochromatin repressive domains at common sets of target genes, from the early stages of development to adulthood. How they functionally preserve transcriptional silencing has been deeply explored in embryonic stem cells (ESC), but remains poorly described in an in vivo context. Taking advantage of Cre-dependent conditional knockout mouse models targeting distinct Polycomb activities in the intestinal epithelium, the present work proposes to characterize the interplay among the PRC1 subcomplexes and their relationship with PRC2 activity. In particular, it functionally dissects how PRC1 and PRC2 cooperate or have independent roles to establish the epigenetic signature of intestinal cells. Since catalytic inactivation of PRC1 (RING1A/B double knockout) affects intestinal homeostasis by exhausting the stem cell pool, we have ablated single PCGF activities (PRC1.1-1.6) to determine the role of variant (vPRC1) and canonical PRC1 (cPRC1) subcomplexes in the transcriptional identity of intestinal epithelial cells. By coupling in vivo studies with genome-wide analyses, we have found that none of them reproduces the effects of RING1A/B loss of function, resulting in the compensation of H2AK119ub1 deposition and gene repression. Moreover, we have evaluated the dependency of PRC2 function on the downstream cPRC1 activity, demonstrating that PRC2 acts autonomously to control transcriptional silencing and secretory lineage commitment. Unexpectedly, we have found that removal of PRC1.1 rescued the effects of PRC2 inactivation in lineage skewing while maintaining extensive transcriptional derepression, revealing an intimate connection between distinct Polycomb activities in the control of intestinal cell plasticity and identity. Finally, by validating in parallel some of these in vivo findings in the mouse ESC model, we have also observed that promiscuous interactions between vPRC1 and cPRC1 occur, identifying a novel mechanism of functional compensation among Polycomb subcomplexes.
13-apr-2023
internal advisor: S. Campaner ; external advisor: G. Gargiulo ; supervisor: D. Pasini
Dipartimento di Oncologia ed Emato-Oncologia
English
34
2022
MEDICINA DEI SISTEMI
Settore BIO/11 - Biologia Molecolare
Ricerca di base
Pubblicazione scientifica
Polycomb; epigenetics; transcription; cell commitment; cell identity
CAMPANER, STEFANO
PASINI, DIEGO
Doctoral Thesis
Prodotti della ricerca::Tesi di dottorato
-2.0
open
Università degli Studi di Milano
info:eu-repo/semantics/doctoralThesis
1
S. Amato
CROSSTALK BETWEEN PRC1 AND PRC2 IN INTESTINAL LINEAGE COMMITMENT AND CELL IDENTITY / S. Amato ; internal advisor: S. Campaner ; external advisor: G. Gargiulo ; supervisor: D. Pasini. Dipartimento di Scienze della Salute, 2023 Apr 13. 34. ciclo, Anno Accademico 2022.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/958336
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