Biology subjects

Marchiori, E.

Publications and source records attributed to Marchiori, E..

4 recordsLinked to original sources

bioRxiv2025

Caffeine-regulated molecular switches for functional control of CAR T cells in vivo

The limited controllability of CAR T cells in patients represents a key challenge of this highly potent immunotherapy. A molecular ON-switch, which can be regulated with a non-toxic and readily available small molecule drug, would represent a major advance towards controllable CAR T therapeutics. For that purpose, we engineered caffeine-responsive heterodimeric ON-switches (CaffSwitches) and demonstrate their high caffeine-dependency and virtually absent leakiness. When incorporating these CaffSwitches into CARs, the resulting CaffCARs were efficiently activated by caffeine concentrations achieved in human plasma after drinking one cup of coffee. Moreover, CaffCAR T cells also showed efficient tumor clearance in an in vivo mouse model, which was completely abolished in the absence of caffeine. This tight control was even observed with c-Jun overexpressing CaffCAR T cells, despite their massive expansion. Together, we anticipate that these novel CaffSwitches will be valuable tools for the development of safe and efficient next generation CAR T cells.

synthetic biology↗

bioRxiv2024

Mapping Cryptic Phosphorylation Sites in the Human Proteome

Advances in computational and experimental methods have revealed the existence of transient, non-native protein folding intermediates that could play roles in disparate biological processes, from regulation of protein expression to disease-relevant misfolding mechanisms. Here, we tested the possibility that specific post-translational modifications may involve residues exposed during the folding process by assessing the solvent accessibility of 87,138 post-translationally modified amino acids in the human proteome. Unexpectedly, we found that one-third of phosphorylated proteins present at least one phosphosite completely buried within the proteins inner core. Computational and experimental analyses suggest that these cryptic phosphosites may become exposed during the folding process, where their modification could destabilize native structures and trigger protein degradation. Phylogenetic investigation also reveals that cryptic phosphosites are more conserved than surface-exposed phosphorylated residues. Finally, cross-referencing with cancer mutation databases suggests that phosphomimetic mutations in cryptic phosphosites can increase tumor fitness by inactivating specific onco-suppressors. These findings define a novel role for co-translational phosphorylation in shaping protein folding and expression, laying the groundwork for exploring the implications of cryptic phosphosites in health and disease.

biochemistry↗

bioRxiv2022

Long range allostery mediates the regulation of plasminogen activator inhibitor 1 by vitronectin

The serpin plasminogen activator inhibitor 1 (PAI-1) spontaneously undergoes a massive structural change from a metastable, active conformation, with a solvent accessible reactive center loop (RCL), to a stable, inactive or latent conformation in which the RCL has inserted into the central {beta} sheet. Physiologically, conversion to the latent state is regulated by the binding of vitronectin which retards the rate of this latency transition approximately 2-fold. We investigated the effects of vitronectin on the PAI-1 latency transition using all-atom path sampling simulations in explicit solvent. In simulated latency transitions of free PAI-1, the RCL is quite mobile as is the gate, the region that impedes RCL access to the central {beta} sheet. This mobility allows the formation of a transient salt bridge that facilitates the transition, and this finding rationalizes existing mutagenesis results. Vitronectin binding reduces RCL and gate mobility by allosterically rigidifying structural elements over 40 [A] away from the binding site thus blocking the transition to the latent conformation. The effects of vitronectin are propagated by a network of dynamically correlated residues including a number of conserved sites that have previously been identified as important for PAI-1 stability. Simulations also revealed a transient pocket populated only in the vitronectin bound state which corresponds to a cryptic drug binding site identified by crystallography. Overall, these results shed new light on regulation of the PAI-1 latency transition by vitronectin and illustrate the potential of path sampling simulations for understanding functional conformational changes in proteins and for facilitating drug discovery.

biophysics↗

bioRxiv2017

The importance of Chargaff’s second parity rule for genomic signatures in metagenomics

An important problem in metagenomic data analysis is to identify the source organism, or at least taxon, of each sequence. Most methods tackle this problem in two steps by using an alignment-free approach: first the DNA sequences are represented as points of a real n-dimensional space via a mapping function then either clustering or classification algorithms are applied. Those mapping functions require to be genomic signatures: the dissimilarity between the mapped points must reflect the degree of phylogenetic similarity of the source species. Designing good signatures for metagenomics can be challenging due to the special characteristics of metagenomic sequences; most of the existing signatures were not designed accordingly and they were tested only on error-free sequences sampled from a few dozens of species.\n\nIn this work we analyze comparatively the goodness of existing and novel signatures based on tetranu-cleotide frequencies via statistical models and computational experiments; we also study how they are affected by the generalized Chargaffs second parity rule (GCSPR), which states that in a given sequence longer than 50kbp, inverse oligonucleotides are approximately equally frequent. We analyze 38 million sequences of 150 bp-1,000 bp with 1% base-calling error, sampled from 1,284 microbes. Our models indicate that GCSPR reduces strand-dependence of signatures, that is, their values are less affected by the source strand; GCSPR is further exploited by some signatures to reduce the intra-species dispersion. Two novel signatures stand out both in the models and in the experiments: the combination signature and the operation signature. The former achieves strand-independence without grouping oligonucleotides; this could be valuable for alignment-free sequence comparison methods when distinguishing inverse oligonucleotides matters. Operation signature sums the frequencies of reverse, complement, and inverse tetranucleotides; having 72 features it reduces the computational intensity of the analysis.

bioinformatics↗