Biology subjects

Carbognin, E.

Publications and source records attributed to Carbognin, E..

4 recordsLinked to original sources

bioRxiv2023

TGF-beta dynamically controls epithelial identity in a 3D model of human epiblast

Pluripotency is the ability to give rise to all cell types of the body and is first observed in a mass of disorganised cells of the embryo. Upon implantation, pluripotent cells form a columnar epithelium and undergo lumenogenesis. At gastrulation, a portion of the pluripotent epiblast will undergo epithelial to mesenchymal transition (EMT), forming the primitive streak (PS). It still remains unclear what molecular mechanism supports the epithelial identity of the pluripotent epiblast before gastrulation. Here we developed an optimised, chemically defined 3D model of human pluripotent epiblast formation in which conventional pluripotent stem cells (PSCs) self-organise into a columnar epithelium with a lumen in 48 hours. From 72 hours we observed spontaneous symmetry breaking and specification of PS-like cells, as confirmed by single-cell RNA sequencing. We found that Insulin and FGF signalling are both required for the proliferation and survival of the pluripotent epiblast model. Conversely, TGF-beta signalling maintains epithelial identity. Epithelial identity appears uncoupled from the expression of canonical pluripotency markers OCT4, NANOG and PRDM14, but under the control of ZNF398. Once the pluripotent epithelium is established, TGF-beta inhibition is inconsequential, and stimulation with Activin A leads to highly efficient PS induction. We conclude that TGF-beta dynamically orchestrates epithelial identity of human pluripotent cells.

developmental biology↗

bioRxiv2023

KLF7 is a general inducer of human pluripotency

Pluripotency is the capacity to give rise to all differentiated cells of the body and the germ line and is governed by a self-reinforcing network of transcription factors. The forced expression of only some of these factors enables the reprogramming of somatic cells to pluripotency. In murine cells, several kruppel-like factors (KLFs) have been identified as stabilisers and inducers of pluripotency. Human somatic cells are routinely reprogrammed by expression of KLF4 in combination with OCT4, SOX2 and cMYC (OSKM). An extensive transcriptome analysis revealed, however, that KLF4 is barely expressed in conventional human pluripotent stem cells (PSCs). Here we show that KLF7 is robustly expressed in conventional human PSCs and it allows transcription factor-mediated somatic reprogramming. Moreover, we find that KLF7 is highly expressed in naive PSCs and its forced expression in conventional hPSCs induces upregulation of naive markers and boosts efficiency of chemical resetting to naive PSCs, overall suggesting that KLF7 is a general human pluripotency factor and an inducer of pluripotency.

developmental biology↗

bioRxiv2023

Identification of druggable host dependency factors shared by multiple SARS-CoV-2 variants of concern

The high mutation rate of SARS-CoV-2 leads to emergence of several variants, some of which are resistant to vaccines and drugs targeting viral elements. Targeting host dependency factors - cell proteins required for viral replication - would help avoid resistance. However, whether different SARS-CoV-2 variants induce conserved cell responses and exploit the same core host factors is still unclear. We compared three variants of concern and observed that the host transcriptional response was conserved, differing only in kinetics and magnitude. By CRISPR screening we identified the host genes required for infection by each variant: most of the identified genes were shared by multiple variants, both in lung and colon cells. We validated our hits with small molecules and repurposed FDA-approved drugs. All drugs were highly effective against all tested variants, including delta and omicron, new variants that emerged during the study. Mechanistically, we identified ROS production as a pivotal step in early virus propagation. Antioxidant drugs, such as N-acetyl cysteine (NAC), were effective against all variants both in human lung cells, and in a humanised mouse model. Our study supports the use of available antioxidant drugs, such as NAC, as a general and effective anti-COVID-19 approach.

microbiology↗

bioRxiv2017

A common molecular logic determines embryonic stem cell self-renewal and reprogramming

During differentiation and reprogramming new cell identities are generated by reconfiguration of gene regulatory networks. Here we combined automated formal reasoning with experimentation to expose the logic of network activation during induction of naive pluripotency. We find that a Boolean network architecture defined for maintenance of naive state embryonic stem cells (ESC) also explains transcription factor behaviour and potency during resetting from primed pluripotency. Computationally identified gene activation trajectories were experimentally substantiated at single cell resolution. Contingency of factor availability explains the counterintuitive observation that Klf2, which is dispensable for ESC maintenance, is required during resetting. We tested 136 predictions formulated by the dynamic network, finding a predictive accuracy of 78.7%. Finally, we show that this network explains and predicts experimental observations of somatic cell reprogramming. We conclude that a common deterministic program of gene regulation is sufficient to govern maintenance and induction of naive pluripotency. The tools exemplified here could be broadly applied to delineate dynamic networks underlying cell fate transitions.

developmental biology↗