Biology subjects

Fernandez-Perez, D.

Publications and source records attributed to Fernandez-Perez, D..

4 recordsLinked to original sources

bioRxiv2026

Leukemia stem cell expansion cultures reveal clonal drivers of leukemogenesis and therapy response

Leukemia stem cells (LSCs) contain the highest capacity for leukemia-reinitiation and therapy-resistance across all leukemic cells, but our understanding of their molecular and cellular properties remains limited due to their relative rarity and ineffective cell culture systems to maintain their purity at scale. Here, we develop Polymer-based Leukemic STem-cell Cultures (PLSTCs) and demonstrate their capacity to derive and propagate large numbers of Npm1cA/Flt3ITD acute myeloid leukemia (AML) stem cells at high purities. Compared to traditional cultures, PLSTCs show more than 1000-fold enrichment in functional LSCs based on single-cell gene expression signatures and leukemia-initiating assays. Tracing LSC clones with genomic LARRY barcodes during ex vivo expansion, we reveal that PLSTCs can sustain a diversity of self-renewing LSC states with stable, heritable transcriptional programs. Using dynamic state-fate analysis, we characterize clonal programs that are linked with enhanced ex vivo self-renewal, in vivo leukemia initiation, and therapeutic response to induction chemotherapy. LSC clones primed to resist treatment were enriched for a rare cell state that underwent a fate-switch and produced megakaryocytic-erythroid-like leukemic cells that expanded in the spleen. Targeting LSC programs through pooled CRISPR and single-cell sequencing (CROPseq) in PLSTCs, we reveal that chondroitin-sulfate synthesis is required to maintain a primitive LSC state and leukemic recovery from chemotherapy. In sum, our studies showcase the powerful application of scalable leukemic stem-cell expansion cultures and dynamic state-fate analysis of AML LSCs. We anticipate these systems will accelerate our understanding and interception of stem cell plasticity in cancer.

cancer biology↗

bioRxiv2024

Pre-existing stem cell heterogeneity dictates clonal responses to acquisition of cancer driver mutations

Cancer cells display wide phenotypic variation even across patients with the same mutations. Differences in the cell of origin provide a potential explanation, but these assays have traditionally relied on surface markers, lacking the clonal resolution to distinguish heterogeneous subsets of stem and progenitor cells. To address this challenge, we developed STRACK, an unbiased framework to longitudinally trace clonal gene expression and expansion dynamics before and after acquisition of cancer mutations. We studied two different leukemia driver mutations, Dnmt3a-R882H and Npm1cA, and found that the response to both mutations was highly variable across different stem cell states. Specifically, a subset of differentiation-biased stem cells, which normally become outcompeted with time, can efficiently expand with both mutations. Npm1c mutations surprisingly reversed the intrinsic bias of the clone-of-origin, with stem-biased clones giving rise to more mature malignant states. We propose a clonal "reaction norm", in which pre-existing clonal states dictate different cancer phenotypic potential. Highlights- Single cell tracing of cancer initiation at the clonal level (STRACK). - Ex vivo expansion cultures sustain intrinsic and heritable HSC heterogeneity. - Premalignant mutations enhance the self-renewal of high-output stem cells, increasing their survival probability. - Transforming mutations reprogram low-output stem cell fates to more mature malignant states.

cancer biology↗

bioRxiv2020

BAP1 activity regulates PcG occupancy and global chromatin condensation counteracting diffuse PCGF3/5-dependent H2AK119ub1 deposition

BAP1 is recurrently mutated or deleted in a large number of diverse cancer types, including mesothelioma, uveal melanoma and hepatocellular cholangiocarcinoma. BAP1 is the catalytic subunit of the Polycomb Repressive De-Ubiquitination complex (PR-DUB) which removes PRC1 mediated H2AK119ub1. We and others have shown that H2AK119ub1 is essential for maintaining transcriptional repression and contributes to PRC2 chromatin recruitment. However, the precise relationship between BAP1 and PRC1 remains mechanistically elusive. Using embryonic stem cells, we show that a major function of BAP1 is to restrict H2AK119ub1 deposition to target sites. This increases the stability of PcG complexes with their targets and prevents diffuse accumulation of H2AK119ub1 and H3K27me3 modifications. Loss of BAP1 results in a broad increase in H2AK119ub1 levels that are primarily dependent on PCGF3/5-PRC1 complexes with a mechanism that is reminiscent of X-chromosome inactivation. Increased genome-wide H2AK119ub1 levels titrates away PRC2 from its targets and stimulates diffuse H3K27me3 accumulation across the genome. This decreases the activity of PcG repressive machineries at physiological targets and induces a general compaction of the entire chromatin. Our findings provide evidences for a unifying model that resolves the apparent contradiction between BAP1 catalytic activity and its role in vivo, uncovering molecular vulnerabilities that could be useful for BAP1-related pathologies.

molecular biology↗

bioRxiv2019

Histone H2AK119 Mono-Ubiquitination is Essential for Polycomb-Mediated Transcriptional Repression

The major function of Polycomb group proteins (PcG) is to maintain transcriptional repression to preserve cellular identity. This is exerted by two distinct repressive complexes, PRC1 and PRC2, that modify histones by depositing H2AK119ub1 and H3K27me3, respectively. Both complexes are essential for development and are deregulated in several types of human tumors. PRC1 and PRC2 exist in different variants and show a complex regulatory cross-talk. However, the contribution that H2AK119ub1 plays in mediating PcG repressive functions remains largely controversial. Coupling an inducible system with the expression of a fully catalytic inactive RING1B mutant, we demonstrated that H2AK119ub1 deposition is essential to maintain PcG-target genes repressed in ESC. Loss of H2AK119ub1 induced a rapid displacement of PRC2 activity and a loss of H3K27me3 deposition. This affected both PRC2.1 and PRC2.2 variants and further correlated with a strong displacement and destabilization of canonical PRC1. Finally, we find that variant PRC1 forms can sense H2AK119ub1 deposition, which contributes to their stabilization specifically at sites where this modification is highly enriched. Overall our data place H2AK119ub1 deposition as central hub that mount PcG repressive machineries to preserve cell transcriptional identity.

molecular biology↗