ERC ADV GRANT 2014 DENOVOSTEM, PI Stefano Piccolo, Grant ID 670126

In living organs, cells are in intimate contact with each other and with their microenvironment. In this context, cells display remarkable, and yet mysterious, "social" behaviors. Prominent examples are the ability of cells to stop proliferating (i.e., of making copies of themselves) in response to signals emanating from their surroundings or to change their behavior in response to tissue needs, as it occurs during wound healing. Healthy tissues can also recognize individual tumor cells, and either eliminate them, or induce them to "normalize" their behavior, as such taming the effect of oncogenic mutations.

These control systems are poorly understood because they represent forms of regulation that are proper of a cellular ensemble in its entirety. Thus, new experimental tools and conceptual paradigms are essential to understand the molecular basis of these tissue-level attributes. Addressing these unknowns has been a main focus of our ERC grant. We believe that understanding how cell communicate with each other and their environment will reveal a "treasure throve" of therapeutic opportunities in regenerative medicine and cancer treatment. This includes new ways to generate stem cells when the natural pool of these cells has been eroded by damage or ageing, and innovative strategies to prevent tumor emergence based on the body own healing capacities.

Our core objective has been to tackle the core question behind these biological events, that is, how normal cells change their fate once the environment is changing. In asking this, our primary perspective has been rooted to the notion that cells are embedded in mechanical signals emanating from their physical touching each other and the extracellular matrix to which they adhere. These signals are potent determinant of cell behavior affecting cells every second of their life. Cells respond to mechanical signals using special mechanosensory proteins, that are the YAP and TAZ transcription factors.

Through this ERC grant, our work revealed a striking overlap between stemness, tissue regeneration and cancer, all sharing features of cell fate plasticity dictated by the activation of YAP/TAZ transcriptional responses. Our insights in cancer biology and cell-cell communication have also with far-reaching implications in the biology of ageing and other human disorders.


Main achievements as Corresponding Author:

1) We discovered and reported a new cellular reprogramming paradigm (Panciera et al., Cell Stem Cell 2016): expression of a single transcription factor, YAP or TAZ, turns differentiated cells of a given tissue into the somatic stem cells (SC) of the same tissue. For this, we used a combination of lineage-tracing and functional assays in neurons, luminal mammary gland cells and pancreatic acini. YAP-reprogramming respects the epigenetic barriers defining "tissue identity" and thus differs from Yamanaka's type of reprogramming to a pluripotent, ES-like state.

2) During YAP-induced reprogramming, cells go back, transiently, to a progenitor state that is typical of the fetal stages of development of that tissue. Importantly, we showed that this reprogramming step occurs in vivo, when cell plasticity is required to build-up new SCs to compensate tissue damage and foster regeneration. YAP activation in these regenerative contexts is mechanically-induced by changes in the physical properties of the extracellular matrix (ECM) that follows tissue damage. See Yui et al. Cell Stem Cell 2018 (Collaborative work with Kim Jensen, BRIK, Copenhagen).

3) We have recently found that common oncogenic lesions induce, once introduced into otherwise normal differentiated cells, dedifferentiation into cells with SC-like attributes. Oncogene-induced endowment of SC attributes is entirely dependent on YAP/TAZ-mechanotransduction (Panciera et al., Nature Materials, 2020).

4) Although YAP/TAZ are difficult to target directly for cancer therapy, investigating their epigenetic functions and co-factors may provide important routes of intervention. We found that SWI/SNF, a protein complex exceedingly inactivated in human tumors, serves as a potent inhibitor of the pro-oncogenic transcriptional coactivators YAP/TAZ. This work offers a molecular framework based on YAP/TAZ regulation whereby mechanical and genetic lesions conspire to overrule tissue-level checkpoints that normally control cell fates and prevent cancer. See Chang et al., Nature 2018.

5) We explored at the genome-wide level of the mechanisms by which YAP/TAZ regulate gene expression. Following our original identification of YAP/TAZ ChIP-Seq analyses (reviewed in Totaro et al., Nature Cell Biology 2018), we discovered that YAP/TAZ physically engage the general coactivator BRD4, dictating the genome-wide association of BRD4 to chromatin. Inhibitors of BRD4 phenocopy the effects of YAP/TAZ inactivation causing an epigenetic reprogramming of cancer cells that leads to regression of pre-existing early neoplastic lesions. Zanconato et al., Nature Medicine 2018. We have more recently expanded, in collaboration with Massimiliano Pagani team (at IFOM, Milan) our investigation of the epigenomic landscape of human colorectal cancers and other cancer types (Della Chiara et al., Nature Communications 2021); the data reveal that YAP/TAZ control a core of pan-cancer enhancers that are exclusive of tumor cells and not of normal cells.

6) We advanced on the mechanisms by which YAP/TAZ mechanotransduction controls epidermal stem cell fates by integrating with Notch signaling for fine-grained orchestration of SC decisions. Published in Totaro et al., Nature Communications 2017; and Totaro et al., Trends Cell Biol 2018). This opened to the possibility to use Notch inhibitor for expanding the passaging potential of epidermal stem cells.

7) We have connected mechanotransduction and cell fate plasticity to new players and biological processes. We found that YAP/TAZ mechanotransduction controls autophagy, and that this form of renewing cellular structures is intrinsic to cell plasticity (Totaro et al., PNAS, 2019).

8) We have expanded the functional exploration of YAP/TAZ in non-epithelial tumors. By single-cell analyses we recently characterized the native, endogenous Glioblastoma SCs (GSCs) at the apex of the differentiation hierarchy of glioblastoma, the most frequent and uncurable form of brain cancer. By reconstructing the GSCs' regulatory network, we identified the YAP/TAZ coactivators as master regulators of this cell state. We found that YAP/TAZ install GSC properties in primary, otherwise normal astrocytes, and this occurs downstream of multiple oncogenic lesions. YAP/TAZ are required for GMB initiation and maintenance in vivo in different mouse and human GBM models. In this context, YAP/TAZ act as main roadblock of GSC differentiation, and their inhibition irreversibly locks differentiated GBM cells into a nontumorigenic state, preventing plasticity and regeneration of GSC-like cells. Thus, GSC identity is linked to YAP/TAZ as regulators of the balance between stemness and differentiation, as such serving as integrating hubs of cancer genetics and microenvironmental inputs controlling the multifaceted biology of GBM. See our recent work by Castellan et al. Nature Cancer, 2021.

9) We have reviewed the field in several Reviews and Perspective articles, that we think have helped the field, including Zanconato et al., Cancer Cell 2016; Panciera et al., Nature Review Mol Cell Biol., 2017; Zanconato et al., Nature Reviews Cancer, 2019; Battilana et al., Cell Stress 2021).

Collaborative work as coauthors.

10) DENOVOSTEM (Silvio Bicciato team only, collaboration with Michele De Luca, UniMoRe) contributed to work published in Nature, Hirsch et al., 2017, on the grow of artificial skin then successfully transplanted on a child suffering from a genetic skin disease.

11) We collaborated to pioneering work on the connections between YAP/TAZ and inflammation. In particular, in collaboration with Marco Foiani's team at IFOM, Milan, we found that ATR affects cell mechanics at the nuclear envelope, and this is key to preserve YAP/TAZ mechanosignaling, prevent cGAS/STING signaling, and, as such limit innate immune responses (Kidiyoor et al., Nature Communications 2020). Alterations of epithelial stem and progenitor cell function is a contributing factor in inflammation-related diseases in human. In collaboration with Freddy Radtke at EPFL, we contributed to the discovery that chronic inflammation imposes aberrant cell fate plasticity during epithelial regeneration through YAP/TAZ mechanotransduction Nowell et al., Nature Cell Biology 2016). Also in line with the links between YAP/TAZ and inflammation - this time in the context of cancer - is collaborative work with V. Bronte, on the role of YAP/TAZ as sensors of cell mechanics in Tumor Associated Macrophages (TAM), where they play an essential role in escorting tumor cell dissemination at the tumor front by remodeling the ECM (Marigo et al., Cancer Discov. 2020).

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