Biology subjects

Ferrena, A.

Publications and source records attributed to Ferrena, A..

8 recordsLinked to original sources

bioRxiv2024

Aging limits stemness and tumorigenesis in the lung by reprogramming iron homeostasis

Aging is associated with a decline in the number and fitness of adult stem cells1-4. Aging-associated loss of stemness is posited to suppress tumorigenesis5,6, but this hypothesis has not been tested in vivo. Here, using physiologically aged autochthonous genetically engineered mouse models and primary cells7,8, we demonstrate aging suppresses lung cancer initiation and progression by degrading stemness of the alveolar cell of origin. This phenotype is underpinned by aging-associated induction of the transcription factor NUPR1 and its downstream target lipocalin-2 in the cell of origin in mice and humans, leading to a functional iron insufficiency in the aged cells. Genetic inactivation of the NUPR1--lipocalin-2 axis or iron supplementation rescue stemness and promote tumorigenic potential of aged alveolar cells. Conversely, targeting the NUPR1-- lipocalin-2 axis is detrimental to young alveolar cells via induction of ferroptosis. We find that aging-associated DNA hypomethylation at specific enhancer sites associates with elevated NUPR1 expression, which is recapitulated in young alveolar cells by inhibition of DNA methylation. We uncover that aging drives a functional iron insufficiency, which leads to loss of stemness and tumorigenesis, but promotes resistance to ferroptosis. These findings have significant implications for the therapeutic modulation of cellular iron homeostasis in regenerative medicine and in cancer prevention. Furthermore, our findings are consistent with a model whereby most human cancers initiate in young individuals, revealing a critical window for such cancer prevention efforts.

cancer biology↗

bioRxiv2024

Comprehensive single cell transcriptomics analysis of murine osteosarcoma uncovers Skp2 function in metastasis, genomic instability and immune activation and reveals additional target pathways

Osteosarcoma (OS) is the most common primary pediatric bone malignancy. One promising new therapeutic target is SKP2, encoding a substrate recognition factor of the SCF E3 ubiquitin ligase responsible for ubiquitination and proteasome degradation of substrate p27, thus driving cellular proliferation. We have shown previously that knockout of Skp2 in an immunocompetent transgenic mouse model of OS improved survival, drove apoptosis, and induced tumor inflammation. Here, we applied single-cell RNA-sequencing (scRNA-seq) to study primary OS tumors derived from Osx-Cre driven conditional knockout of Rb1 and Trp53. We showed that murine OS models recapitulate the tumor heterogeneity and microenvironment complexity observed in patient tumors. We further compared this model with OS models with functional disruption of Skp2: one with Skp2 knockout and the other with the Skp2-p27 interaction disrupted (resulting in p27 overexpression). We found reduction of T cell exhaustion and upregulation of interferon activation, along with evidence of replicative and endoplasmic reticulum-related stress in the Skp2 disruption models, and showed that interferon induction was correlated with improved survival in OS patients. Additionally, our scRNA-seq analysis uncovered decreased activities of metastasis-related gene signatures in the Skp2-disrupted OS, which we validated by observation of a strong reduction in lung metastasis in the Skp2 knockout mice. Finally, we report several potential mechanisms of escape from targeting Skp2 in OS, including upregulation of Myc targets, DNA copy number amplification and overexpression of alternative E3 ligase genes, and potential alternative lineage activation. These mechanistic insights into OS tumor biology and Skp2 function suggest novel targets for new, synergistic therapies, while the data and our comprehensive analysis may serve as a public resource for further big data-driven OS research.

cancer biology↗

bioRxiv2024

scDAPP: a comprehensive single-cell transcriptomics analysis pipeline optimized for cross-group comparison

Single-cell transcriptomics profiling has increasingly been used to evaluate cross-group differences in cell population and cell-type gene expression. This often leads to large datasets with complex experimental designs that need advanced comparative analysis. Concurrently, bioinformatics software and analytic approaches also become more diverse and constantly undergo improvement. Thus, there is an increased need for automated and standardized data processing and analysis pipelines, which should be efficient and flexible too. To address these, we develop the single-cell Differential Analysis and Processing Pipeline (scDAPP), a R-based workflow for comparative analysis of single cell (or nucleus) transcriptomic data between two or more groups and at the levels of single cells or "pseudobulking" samples. The pipeline automates many steps of pre-processing using data-learnt parameters, uses previously benchmarked software, and generates comprehensive intermediate data and final results that are valuable for both beginners and experts of scRNA-seq analysis. Moreover, the analytic reports, augmented by extensive data visualization, increase the transparency of computational analysis and parameter choices, while facilitate users to go seamlessly from raw data to biological interpretation. Availability and ImplementationscDAPP is freely available for non-commercial usage as an R package under the MIT license. Source code, documentation and sample data are available at the GitHub (https://github.com/bioinfoDZ/scDAPP).

bioinformatics↗

bioRxiv2024

Facilitate integrated analysis of single cell multiomic data by binarizing gene expression values

The identity of a cell type can be revealed by its transcriptome and epigenome profiles, both of which can be in flux temporally and spatially, leading to distinct cell states or subtypes. The popular and standard workflow for single cell RNA-seq (scRNA-seq) data analysis applies feature selection, dimensional reduction, and clustering on the gene expression values quantified by read counts, but alternative approaches using a simple classification of a gene to "on" and "off" (i.e., binarization of the gene expression) has been proposed for classifying cells and other downstream analyses. Here, we demonstrate that a direct concatenation of the binarized scRNA-seq data and the standard single cell ATAC-seq data is sufficient and effective for integrated clustering analysis, after applying term-frequency-inverse document frequency (TF-IDF) and single value decomposition (also called latent semantic indexing, LSI) algorithms to the combined data, when the two modalities of omic data are collected using paired multiomic technology. This proposed approach avoids the need for converting scATAC-seq data to gene activity scores for combined analysis and furthermore enables a direct investigation into the contribution of each data type to resolving cell type identity.

bioinformatics↗

bioRxiv2023

Targeted inhibition of SCF-SKP2 confers anti-tumor activities resulting in a survival benefit in osteosarcoma

Osteosarcoma(OS) is a highly aggressive bone cancer for which treatment has remained essentially unchanged for decades. Although OS is characterized by extensive genomic heterogeneity and instability, RB1 and TP53 have been shown to be the most commonly inactivated tumor suppressors in OS. We previously generated a mouse model with a double knockout (DKO) of Rb1 and Trp53 within cells of the osteoblastic lineage, which largely recapitulates human OS with nearly complete penetrance. SKP2 is a repression target of pRb and serves as a substrate recruiting subunit of the SCFSKP2 complex. In addition, SKP2 plays a central role in regulating the cell cycle by ubiquitinating and promoting the degradation of p27. We previously reported the DKOAA transgenic model, which harbored a knock-in mutation in p27 that impaired its binding to SKP2. Here, we generated a novel p53-Rb1-SKP2 triple-knockout model (TKO) to examine SKP2 function and its potential as a therapeutic target in OS. First, we observed that OS tumorigenesis was significantly delayed in TKO mice and their overall survival was markedly improved. In addition, the loss of SKP2 also promoted an apoptotic microenvironment and reduced the stemness of DKO tumors. Furthermore, we found that small-molecule inhibitors of SKP2 exhibited anti-tumor activities in vivo and in OS organoids as well as synergistic effects when combined with a standard chemotherapeutic agent. Taken together, our results suggest that SKP2 inhibitors may reduce the stemness plasticity of OS and should be leveraged as next-generation adjuvants in this cancer.

cancer biology↗

bioRxiv2023

SKP2 knockout in Rb1/p53 deficient mouse models of osteosarcoma induces immune infiltration and drives a transcriptional program with a favorable prognosis

PurposeOsteosarcoma (OS) is an aggressive bone malignancy with a poor prognosis. One putative proto-oncogene in OS is SKP2, encoding a substrate recognition factor of the SCF E3 ubiquitin ligase. We previously demonstrated that SKP2 knockout in murine OS improved survival and delayed tumorigenesis. Here we aim to define the SKP2 drives transcriptional program and its clinical implication in OS. Experimental DesignWe performed RNA-sequencing (RNA-seq) on tumors from a transgenic OS mouse model with conditional Trp53 and Rb1 knockouts in the osteoblast lineage ("DKO": Osx1-Cre;Rb1lox/lox;p53lox/lox) and a triple-knockout model with additional Skp2 germline knockout ("TKO": Osx1-Cre;Rb1lox/lox;p53lox/lox;SKP2-/-). We validated our RNA-seq findings using qPCR and immunohistochemistry. To investigate the clinical implications of our results, we analyzed a human OS patient cohort ("NCI-TARGET OS") with RNA-seq and clinical data. ResultsWe found large differences in gene expression after SKP2 knockout. Strikingly, we observed increased expression of genes related to immune microenvironment infiltration in TKO tumors. We observed significant increases in signature genes for macrophages and to a lesser extent, T cells, B cells and vascular cells. We also uncovered a set of relevant transcription factors that may mediate the changes. In OS patient cohorts, high expression of genes upregulated in TKO was correlated with favorable overall survival, which was largely explained by the macrophage gene signatures. This relationship was further supported by our finding that SKP2 expression was negatively correlated with macrophage infiltration in the NCI-TARGET OS and the TCGA Sarcoma cohort. ConclusionOur findings indicate that SKP2 may mediate immune exclusion from the OS tumor microenvironment, suggesting that SKP2 modulation in OS may induce anti-tumor immune activation. Translational RelevanceOsteosarcoma (OS) is an aggressive bone malignancy. Standard care treatment involving chemotherapy and surgery remains unchanged for decades. OS prognosis remains poor and targeted therapies are critically needed. Immunotherapy failed in clinical trials despite extensive genomic instability, suggesting OS tumors employ active immune exclusion. One putative oncogene in OS is SKP2, which activates cell proliferation. SKP2 is correlated with worse prognosis in patients and knockout of SKP2 improved survival in murine OS. Here, we performed comparative transcriptomic analysis between transgenic OS murine models with SKP2 knockout and controls. We showed that SKP2 knockout dramatic increased immune gene expression largely due to macrophage, and to a lesser extent lymphocytes, infiltration. Interestingly, we found that the increased gene program uncovered from our SKP2 knockout model was correlated with improved survival in OS patients. Our findings indicate a completely new function for SKP2 in mediating immune exclusion in sarcoma and suggest that SKP2 inhibition may lead to improved immune activation and potential treatment.

cancer biology↗

bioRxiv2023

Six3 and Six6 jointly regulate the identities and developmental trajectories of multipotent retinal progenitor cells in the mouse retina

Formation, maintenance, and differentiation of tissue-specific progenitor cells are fundamental tasks during organogenesis. Retinal development is an excellent model for dissecting these processes; mechanisms of retinal differentiation can be harnessed for retinal regeneration toward curing blindness. Using single-cell RNA sequencing of embryonic mouse eye cups in which transcription factor Six3 was conditionally inactivated in peripheral retinas on top of germline deletion of its close paralog Six6 ("DKO"), we identified cell clusters and then inferred developmental trajectories in the integrated dataset. In control retinas, naive retinal progenitor cells had two major trajectories leading to ciliary margin cells and retinal neurons, respectively. The ciliary margin trajectory was directly from naive retinal progenitor cells at G1 phase, and the retinal neuron trajectory was through a neurogenic state marked by Atoh7 expression. Upon Six3 and Six6 dual deficiency, both naive and neurogenic retinal progenitor cells were defective. Ciliary margin differentiation was enhanced, and multi-lineage retinal differentiation was disrupted. An ectopic neuronal trajectory lacking the Atoh7+ state led to ectopic neurons. Differential expression analysis not only confirmed previous phenotype studies but also identified novel candidate genes regulated by Six3/Six6. Six3 and Six6 were jointly required for balancing the opposing gradients of the Fgf and Wnt signaling in the central-peripheral patterning of the eye cups. Taken together, we identify transcriptomes and developmental trajectories jointly regulated by Six3 and Six6, providing deeper insight into molecular mechanisms underlying early retinal differentiation.

developmental biology↗

bioRxiv2022

Single cell transcriptomics uncovers a non-autonomous Tbx1-dependent genetic program controlling cardiac neural crest cell deployment and progression

Disruption of cardiac neural crest cells (CNCCs) results in congenital heart disease, yet we do not understand the cell fate dynamics as these cells differentiate to vascular smooth muscle cells. Here we utilized single-cell RNA-sequencing of NCCs from the pharyngeal apparatus with heart in control mouse embryos and when Tbx1, the gene for 22q11.2 deletion syndrome, is inactivated. We uncovered three dynamic transitions of pharyngeal NCCs expressing Tbx2 and Tbx3 through differentiated CNCCs expressing cardiac transcription factors with smooth muscle genes, and that these transitions are altered non-autonomously by loss of Tbx1. Further, inactivation of Tbx2 and Tbx3 in early CNCCs resulted in aortic arch branching defects due to failed smooth muscle differentiation. Loss of Tbx1 interrupted mesoderm to CNCC cell-cell communication with upregulation of BMP signaling with reduced MAPK signaling and failed dynamic transitions of CNCCs leading to disruption of aortic arch artery formation and cardiac outflow tract septation.

developmental biology↗