Biology subjects

Dotto, G.-P.

Publications and source records attributed to Dotto, G.-P..

5 recordsLinked to original sources

bioRxiv2023

Nuclear lamin A/C phosphorylation by loss of Androgen Receptor is a global determinant of cancer-associated fibroblast activation.

Alterations of nuclear structure and function, and associated impact on gene transcription, are a hallmark of cancer cells. Little is known of these alterations in Cancer-Associated Fibroblasts (CAFs), a key component of the tumor stroma. Here we show that loss of androgen receptor (AR), which triggers early steps of CAF activation in human dermal fibroblasts (HDFs), leads to nuclear membrane alterations and increased micronuclei formation, which are unlinked from induction of cellular senescence. Similar alterations occur in fully established CAFs, which are overcome by restored AR function. AR associates with nuclear lamin A/C and loss of AR results in a substantially increased lamin A/C nucleoplasmic redistribution. Mechanistically, AR functions as a bridge between lamin A/C with the protein phosphatase PPP1. In parallel with a decreased lamin-PPP1 association, AR loss results in a marked increase of lamin A/C phosphorylation at Ser 301, which is also a feature of CAFs. Phosphorylated lamin A/C at Ser 301 binds to the transcription promoter regulatory region of several CAF effector genes, which are upregulated due to the loss of AR. More directly, expression of a lamin A/C Ser301 phosphomimetic mutant alone is sufficient to convert normal fibroblasts into tumor-promoting CAFs of the myofibroblast subtype, without an impact on senescence. These findings highlight the pivotal role of the AR-lamin A/C-PPP1 axis and lamin A/C phosphorylation at Ser 301 in driving CAF activation.

cancer biology↗

bioRxiv2022

Androgen Receptor is a Determinant of Melanoma Targeted Drug Resistance

Melanoma provides a primary benchmark for targeted drug therapy. Most melanomas with BRAFV600 mutations regress in response to BRAF/MEK inhibitors (BRAFi/MEKi). However, nearly all relapse within the first two years, and there is a connection between pathways involved in BRAFi/MEKi-resistance and poor response to immune checkpoint therapy. We recently showed that androgen receptor (AR) activity is required for melanoma cell proliferation and tumorigenesis. Here we find that AR expression is markedly increased in BRAFi resistant melanoma cells as well as in sensitive cells soon after BRAFi exposure. Increased AR expression is by itself sufficient to render melanoma cells BRAFi-resistant, eliciting transcriptional changes of BRAFi resistant subpopulations and elevated EGFR and SERPINE1 expression of likely clinical significance. Inhibition of AR expression and activity blunts changes in gene expression and suppresses proliferation and tumorigenesis of BRAFi-resistant melanoma cells, enhances MHC I expression and CD8+ T cells infiltration. Our findings point to targeting AR as a possible co-adjuvant approach for the prevention and management of the disease.

cancer biology↗

bioRxiv2021

Sphingolipid Control of Fibroblast Heterogeneity Revealed by Single-Cell Lipidomics

Human cells produce thousands of lipids that impact a wide range of biological processes in ways we are only starting to characterize. The cellular composition in lipids changes during differentiation events and also varies across individual cells of the same type. Yet, the precise differences in lipid composition that directly affect cell phenotypes remain unknown. Here we have measured the lipidomes and transcriptomes of individual human dermal fibroblasts by coupling high-resolution mass spectrometry imaging to single-cell transcriptomics. We found that the cell-to-cell variation of specific lipid metabolic pathways contributes to the establishment of cell states involved in wound repair and in skin cancer growth. Sphingolipid composition defined fibroblast subpopulations while sphingolipid metabolic rewiring drove cell state transitions. These data uncover a role for cell-to-cell lipid heterogeneity in the determination of cell states and reveal a new regulatory component to the homeostasis and self-organization of multicellular systems.

biochemistry↗

bioRxiv2020

Sustained Androgen Receptor signaling is a determinant of melanoma cell growth potential and tumorigenesis

Melanoma is a benchmark of major clinical significance for cancer development with greater aggressiveness in the male than the female population. Surprisingly little is known on the role of androgen receptor (AR) signaling in the disease. Irrespectively of expression levels, genetic and pharmacological suppression of AR activity in a large panel of melanoma cells, derived from both male and female patients, suppresses proliferation and self-renewal potential while, conversely, increased AR expression or ligand stimulation enhance proliferation. AR gene silencing in multiple melanoma lines elicits a shared gene expression signature related to interferon- and inflammatory cytokines signaling with an inverse association with DNA repair-associated genes, which is significantly linked with better patients survival. AR plays an essential function in maintenance of genome integrity: in both cultured melanoma cells and tumors, loss of AR activity leads to chromosomal DNA breakage, leakage into the cytoplasm, and stimulator of interferon genes (STING) activation. In vivo, reduced tumorigenesis resulting from AR gene silencing or pharmacological inhibition is associated with intratumor macrophage infiltration and, in an immune competent mouse model, cytotoxic T cell activation. Although at different levels, androgens are produced in both male and female individuals and AR targeting provides an attractive therapy approach for improved management of melanoma irrespective of patients sex and gender. SignificanceThe study uncovers an essential role of androgen receptor (AR) signaling in melanoma cell expansion and tumorigenesis, with loss of AR activity inducing cellular senescence, genomic DNA breakage, a STING dependent inflammatory cascade and immune cells recruitment. Use of AR inhibitors as growth inhibitory and DNA damaging agents in melanoma cells can provide an attractive venue for new combination approaches for management of the disease.

cancer biology↗

bioRxiv2019

Visualizing cellular heterogeneity by quantifying the dynamics of MAPK activity in live mammalian cells with synthetic fluorescent biosensors

Mitogen-Activated Protein Kinases (MAPKs) control a wide array of cellular functions by transducing extracellular information into defined biological responses. In order to understand how these pathways are regulated, dynamic single cell measurements are highly needed. Fluorescence microscopy is well suited to perform these measurements, however, more dynamic and sensitive biosensors that allow the quantification of signaling activity in living mammalian cells are required. We have engineered a synthetic fluorescent substrate for human MAPKs that relocates from the nucleus to the cytoplasm when phosphorylated by the kinase. We demonstrate that this reporter provides a better sensitivity relative to other similar biosensors and has allowed the monitoring of ERK MAPK activity pulses upon a single physiological EGF stimulation. In addition, we display its applicability to other MAPKs and in multiple cancer cell lines or primary cells as well as its application in vivo in developing tumors. Using our newly developed biosensors, dynamic single cell measurements with high temporal resolution can be obtained. These reporters can be widely applied to the analysis of molecular mechanisms of MAPK signaling in healthy and diseased state, in cell culture assays or in vivo.

cell biology↗