Biology subjects

Biancon, G.

Publications and source records attributed to Biancon, G..

3 recordsLinked to original sources

bioRxiv2022

Microfluidic immuno-serology assay revealed a limited diversity of protection against COVID-19 in patients with altered immunity

The immune response to SARS-CoV-2 for patients with altered immunity such as hematologic malignancies and autoimmune disease may differ substantially from that in general population. These patients remain at high risk despite wide-spread adoption of vaccination. It is critical to examine the differences at the systems level between the general population and the patients with altered immunity in terms of immunologic and serological responses to COVID-19 infection and vaccination. Here, we developed a novel microfluidic chip for high-plex immuno-serological assay to simultaneously measure up to 50 plasma or serum samples for up to 50 soluble markers including 35 plasma proteins, 11 anti-spike/RBD IgG antibodies spanning all major variants, and controls. Our assay demonstrated the quintuplicate test in a single run with high throughput, low sample volume input, high reproducibility and high accuracy. It was applied to the measurement of 1,012 blood samples including in-depth analysis of sera from 127 patients and 21 healthy donors over multiple time points, either with acute COVID infection or vaccination. The protein association matrix analysis revealed distinct immune mediator protein modules that exhibited a reduced degree of diversity in protein-protein cooperation in patients with hematologic malignancies and patients with autoimmune disorders receiving B cell depletion therapy. Serological analysis identified that COVID infected patients with hematologic malignancies display impaired anti-RBD antibody response despite high level of anti-spike IgG, which could be associated with limited clonotype diversity and functional deficiency in B cells and was further confirmed by single-cell BCR and transcriptome sequencing. These findings underscore the importance to individualize immunization strategy for these high-risk patients and provide an informative tool to monitor their responses at the systems level.

bioengineering↗

bioRxiv2021

Pre-mRNA Splicing Factor U2AF2 Recognizes Distinct Conformations of Nucleotide Variants at the Center of the pre-mRNA Splice Site Signal

The essential pre-mRNA splicing factor U2AF2 (also called U2AF65) identifies polypyrimidine (Py) tract signals of nascent transcripts, despite length and sequence variations. Previous studies have shown that the U2AF2 RNA recognition motifs (RRM1 and RRM2) preferentially bind uridine-rich RNAs. Nonetheless, the specificity of the RRM1/RRM2 interface for the central Py tract nucleotide has yet to be investigated. We addressed this question by determining crystal structures of U2AF2 bound to a cytidine, guanosine, or adenosine at the central position of the Py tract, and compared U2AF2-bound uridine structures. Local movements of the RNA site accommodated the different nucleotides, whereas the polypeptide backbone remained similar among the structures. Accordingly, molecular dynamics simulations revealed flexible conformations of the central, U2AF2-bound nucleotide. The RNA binding affinities and splicing efficiencies of structure-guided mutants demonstrated that U2AF2 tolerates nucleotide substitutions at the central position of the Py tract. Moreover, enhanced crosslinking and immunoprecipitation of endogenous U2AF2 in human erythroleukemia cells showed uridine-sensitive binding sites with lower sequence conservation at the central nucleotide positions of otherwise uridine-rich, U2AF2-bound splice sites. Altogether, these results highlight the importance of RNA flexibility for protein recognition and take a step towards relating splice site motifs to pre-mRNA splicing efficiencies.

biophysics↗

bioRxiv2021

Multi-omics profiling of U2AF1 mutants dissects pathogenic mechanisms affecting RNA granules in myeloid malignancies

Somatic mutations in splicing factors are of significant interest in myeloid malignancies and other cancers. U2AF1, together with U2AF2, is essential for 3 splice site recognition. U2AF1 mutations result in aberrant splicing, but the molecular mechanism and the full spectrum of consequences on RNA biology have not been fully elucidated to date. We performed multi-omics profiling of in vivo RNA binding, splicing and turnover for U2AF1 S34F and Q157R mutants. We dissected specific binding signals of U2AF1 and U2AF2 and showed that U2AF1 mutations individually alter U2AF1-RNA binding, resulting in defective U2AF2 recruitment. We demonstrated a complex relationship between differential binding and splicing, expanding upon the currently accepted loss-of-binding model. Finally, we observed that U2AF1 mutations increase the formation of stress granules in both cell lines and primary acute myeloid leukemia samples. Our results uncover U2AF1 mutation-dependent pathogenic RNA mechanisms and provide the basis for developing targeted therapeutic strategies.

cancer biology↗