Biology subjects

Clerici, M.

Publications and source records attributed to Clerici, M..

6 recordsLinked to original sources

bioRxiv2022

Fip1 is a multivalent interaction scaffold for processing factors in human mRNA 3' end biogenesis

3 end formation of most eukaryotic mRNAs is dependent on the assembly of a [~]1.5 megadalton multiprotein complex, that catalyzes the coupled reaction of pre-mRNA cleavage and polyadenylation. In mammals, the cleavage and polyadenylation specificity factor (CPSF) constitutes the core of the 3 end processing machinery onto which the remaining factors, including cleavage stimulation factor (CstF) and poly(A) polymerase (PAP), assemble. These interactions are mediated by Fip1, a CPSF subunit characterized by high degree of intrinsic disorder. Here, we report two crystal structures revealing the interactions of human Fip1 (hFip1) with CPSF30 and CstF77. We demonstrate that CPSF contains two copies of hFip1, each binding to the zinc finger (ZF) domains 4 and 5 of CPSF30. Using polyadenylation assays we show that the two hFip1 copies are functionally redundant in recruiting one copy of PAP, thereby increasing the processivity of RNA polyadenylation. We further show that the interaction between hFip1 and CstF77 is mediated via a short motif in the N-terminal "acidic" region of hFip1. In turn, CstF77 competitively inhibits CPSF-dependent PAP recruitment and 3 polyadenylation. Taken together, these results provide a structural basis for the multivalent scaffolding and regulatory functions of hFip1 in 3 end processing.

biochemistry↗

bioRxiv2020

Simplexviruses successfully adapt to their host by fine-tuning immune responses

Primate herpes simplex viruses are relatively harmless to their natural hosts, whereas cross-species transmission can result in severe disease. We performed a genome-wide scan for signals of adaptation of simplexviruses to hominins. We found evidence of positive selection in three glycoproteins, with selected sites located in antigenic determinants. Positively selected non-core proteins were involved in different immune-escape mechanisms. By expressing mutants of one of these proteins (ICP47), we show that the amino acid status at the positively selected sites is sufficient to induce HLA-G. HSV-1/HSV-2 ICP47 induced HLA-G when mutated to recapitulate residues in B virus, whereas the mutated version of B virus ICP47 failed to determine HLA-G expression. Thus, the evolution of ICP47 in HSV-1/HSV-2 determined the loss of an immunosuppressive effect, suggesting that simplexviruses tune immune responses to promote successful co-existence with their hosts. These results also help explain the high pathogenicity of B virus in humans.

microbiology↗

bioRxiv2020

Antigenic variation of SARS-CoV-2 in response to immune pressure

The ongoing evolution of SARS-CoV-2 is expected to be at least partially driven by the selective pressure imposed by the human immune system. We exploited the availability of a large number of high-quality SARS-CoV-2 genomes, as well as of validated epitope predictions, to show that B cell epitopes in the spike glycoprotein (S) and in the nucleocapsid protein (N) have higher diversity than non-epitope positions. Similar results were obtained for other human coronaviruses. Conversely, in the SARS-CoV-2 population, epitopes for CD4+ and CD8+ T cells were not more variable than non-epitope positions. A significant reduction in epitope variability was instead observed for some of the most immunogenic proteins (S, N, ORF8, and ORF3a). Analysis over longer evolutionary time-frames indicated that this effect is not due to differential constraints. These data indicate that SARS-CoV-2 is evolving to elude the host humoral immune response, whereas recognition by T cells might benefit the virus.

microbiology↗

bioRxiv2019

Early life adversity and borderline intellectual functioning negatively impact limbic system connectivity in childhood: a connectomics-based study

Epigenetic factors related to early life adversity (ELA) in childhood are major risk factors for borderline intellectual functioning (BIF). BIF affects both adaptive and intellectual abilities, commonly leading to school failure and to an increased risk to develop mental and social problems in the adulthood. This study aimed to investigate the neurobiological underpinnings of ELA associated with BIF in terms of global topological organization and structural connectivity and their relation with intellectual functioning. BIF (N=32) and age-matched typical development (TD, N=14) children were evaluated for intelligence quotient (IQ), behavioral competencies, and ELA. Children underwent an anatomical and diffusion-weighted MR imaging (DWI) protocol. Global brain topological organization was assessed measuring segregation and integration indexes. Moreover, structural matrices, measuring normalized number of fibers (NFn), were compared between the 2 groups using network-based statistics. Finally, a linear regression model was used to explore the relationship between network parameters and clinical measures. Results showed increased behavioral difficulties and ELA, together with decreased network integration in BIF children. Moreover, significantly lower NFn was observed in the BIF group (p=.039) in a sub-network comprising anterior and posterior cingulate, the pericallosal sulcus, the orbital frontal areas, amygdala, basal ganglia, the accumbens nucleus, and the hippocampus. Linear regression showed that NFn significantly predicted IQ (p<.0001). This study demonstrated that ELA in children with BIF is associated with a decreased information integration at the global level, and with an altered structural connectivity within the limbic system strictly related to the intellectual functioning.

neuroscience↗

bioRxiv2018

Stavudine Reduces NLRP3-Inflammasome Activation and Upregulates Aβ-Autophagy

Alzheimers disease (AD) is associated with amyloid-beta (A{beta}) deposition and neuroinflammation, possibly driven by activation of the NLRP3 inflammasome. Nucleoside reverse transcriptase inhibitors (NRTI) hamper the assembly of the NLRP3 inflammasome; we analyzed whether stavudine (D4T), a prototypical NRTI, modulates A{beta}-mediated inflammasome activation; because neuroinflammation impairs A{beta} clearance by phagocytes, phagocytosis and autophagy were examined as well. THP-1-derived macrophages were stimulated in vitro with A{beta}42 alone or after LPS priming with/without D4T. NLRP3 and TREM2 expression was analyzed by RT-PCR, phagocytosis and ASC-Speck by AmnisFlowSight, NLRP3-produced cytokines by ELISA, authophagy by P-ELISA evaluation of P-ERK and P-AKT. Results showed that IL1{beta}, IL18 and caspase-1 were increased whereas A{beta}-phagocytosis and TREM2 were reduced in LPS+A{beta}42-stimulated cells. D4T reduced NLRP3 assembly as well as IL18 and caspase-1 production, but not IL1{beta}, phagocytosis, and TREM2. P-AKT expression was augmented and P-ERK was reduced by D4T, suggesting a stimulatory effect on autophagy. D4T reduces NLRP3 inflammasome-associated inflammation, possibly restoring autophagy, in an in vitro model of AD; it will be interesting to verify its possibly beneficial effects in the clinical scenario.

molecular biology↗

bioRxiv2017

Active site alanine substitutions can convert deubiquitinating enzymes into avid ubiquitin-binding domains

A common strategy for studying the biological role of deubiquitinating enzymes (DUBs) in different pathways is to study the effects of replacing the wild type DUB with a catalytically inactive mutant in cells. We report here that a commonly studied DUB mutation, in which the catalytic cysteine is replaced with alanine, can dramatically increase the affinity of some DUBs for ubiquitin. Overexpression of these tight-binding mutants thus has the potential to sequester cellular pools of monoubiquitin and ubiquitin chains. As a result, cells expressing these mutants may display unpredictable dominant negative physiological effects that are not related to loss of DUB activity. The structure of the SAGA DUB module bound to free ubiquitin reveals the structural basis for the 30-fold higher affinity of Ubp8C146A for ubiquitin. We show that an alternative option, substituting the active site cysteine with arginine, can inactivate DUBs while also decreasing the affinity for ubiquitin.

biochemistry↗