Biology subjects

Chemes, L. B.

Publications and source records attributed to Chemes, L. B..

3 recordsLinked to original sources

bioRxiv2022

Genome-wide CRISPR screens identify novel regulators of wild-type and mutant p53 stability

Tumour suppressor p53 (TP53) is the most frequently mutated gene in cancer. Several hotspot p53 mutants not only lose tumour suppressive capabilities, but also function in a dominant-negative manner, suppressing canonical wild-type p53 function. Furthermore, some hotspot p53 mutants promote oncogenesis by gain-of-function mechanisms. Levels of p53 are regulated predominantly through regulation of protein stability and while wild-type p53 is normally kept at very low levels at steady-state, p53 mutants are often stabilized in tumours, which may be vital for their oncogenic properties. Here, we systematically profiled the factors that regulate protein stability of wild-type and mutant p53 using marker-based genome-wide CRISPR screens. We found that most proteins that regulate wild-type p53 also regulate a subset of p53 mutants with the exception of p53 R337H regulators, which are largely private to this mutant. Mechanistically, we identified FBXO42 as a novel positive regulator of a subset of p53 mutants comprising R273H, R248Q and R248W. We show that FBXO42 acts together with CCDC6 to regulate USP28-mediated p53 stabilization. Our work also identifies C16orf72 as a negative regulator of the stability of wild-type p53 and of all p53 mutants tested. C16orf72 is amplified in breast cancer, and we show that C16orf72 regulates p53 levels in mammary epithelium of mice and its overexpression results in accelerated breast cancer with reduced p53 levels. Together, this work provides a network view of the processes that regulate p53 stability, which might provide clues for reinforcing wild-type p53 or targeting mutant p53 in cancer.

cancer biology↗

bioRxiv2021

A novel bacterial protease inhibitor adjuvant in RBD-based COVID-19 vaccine formulations increases neutralizing antibodies, specific germinal center B cells and confers protection against SARS-CoV-2 infection.

In this work we evaluated recombinant receptor binding domain (RBD) based vaccine formulation prototypes with potential for further clinical development. We assessed different formulations containing RBD plus Alum, AddaS03, AddaVax or the combination of Alum and U-Omp19: a novel Brucella spp. protease inhibitor vaccine adjuvant. Results show that the vaccine formulation composed of U-Omp19 and Alum as adjuvants have a better performance: it significantly increased mucosal and systemic neutralizing antibodies in comparison to antigen plus Alum, AddaVax or AddaS03. Antibodies induced with the formulation containing U-Omp19 not only increased their neutralization capacity against the wild-type virus but also cross neutralized alpha, lambda and gamma variants with similar potency. Also, addition of U-Omp19 to vaccine formulation increased the frequency of RBD-specific geminal center B cells and plasmablasts. Additionally, U-Omp19+Alum formulation induced RBD-specific Th1 and CD8+ T cell responses in spleens and lungs. Finally, this vaccine formulation conferred protection against an intranasal SARS-CoV-2 challenge of K18-hACE2 mice.

immunology↗

bioRxiv2021

Conformational buffering underlies functional selection in intrinsically disordered protein regions

Many disordered proteins conserve essential functions in the face of extensive sequence variation. This makes it challenging to identify the forces responsible for functional selection. Viruses are robust model systems to investigate functional selection and they take advantage of protein disorder to acquire novel traits. Here, we combine structural and computational biophysics with evolutionary analysis to determine the molecular basis for functional selection in the intrinsically disordered adenovirus early gene 1A (E1A) protein. E1A competes with host factors to bind the retinoblastoma (Rb) protein, triggering early S-phase entry and disrupting normal cellular proliferation. We show that the ability to outcompete host factors depends on the picomolar binding affinity of E1A for Rb, which is driven by two binding motifs tethered by a hypervariable disordered linker. Binding affinity is determined by the spatial dimensions of the linker, which constrain the relative position of the two binding motifs. Despite substantial sequence variation across evolution, the linker dimensions are finely optimized through compensatory changes in amino acid sequence and sequence length, leading to conserved linker dimensions and maximal affinity. We refer to the mechanism that conserves spatial dimensions despite large-scale variations in sequence as conformational buffering. Conformational buffering explains how variable disordered proteins encode functions and could be a general mechanism for functional selection within disordered protein regions.

biophysics↗