Biology subjects

Rizzi, G.

Publications and source records attributed to Rizzi, G..

6 recordsLinked to original sources

bioRxiv2023

Resolving genotype-phenotype discrepancies of the Kidd blood group using nanopore sequencing

Due to substantial improvement in read accuracy, third-generation long-read sequencing holds great potential in blood group diagnostics, particularly in cases where traditional genotyping or sequencing techniques, primarily targeting exons, fail to explain serological phenotypes. In this study, we employed Oxford Nanopore sequencing to resolve all genotype-phenotype discrepancies in the Kidd blood group system (JK, encoded by SLC14A1) observed over seven years of routine high-throughput donor genotyping using a mass spectrometry based platform at Blood Transfusion Service Zurich. Discrepant results from standard serological typing and donor genotyping were confirmed by commercial PCR-SSP kits. To resolve discrepancies, we amplified the entire coding region of SLC14A1 ([~]24 kb, exons 3 to 10) in two overlapping long-range PCRs in all samples. Amplicons were barcoded and sequenced on a MinION flow cell. Sanger sequencing and bridge-PCRs were used to confirm findings. Among 11,972 donors with both serological and genotype data available for the Kidd system, we identified 10 cases with unexplained conflicting results. Five were linked to known weak and null alleles caused by variants not included in the routine donor genotyping. In two cases, we identified novel null alleles on the JK*01 (Gly40Asp; c.119G>A) and JK*02 (Gly242Glu; c.725G>A) haplotype, respectively. Remarkably, the remaining three cases were associated with a yet unknown deletion of [~]5 kb spanning exons 9-10 of the JK*01 allele, which other molecular methods had failed to detect. Overall, nanopore sequencing demonstrated reliable and accurate performance for detecting both single-nucleotide and structural variants. It possesses the potential to become a robust tool in the molecular diagnostic portfolio, particularly for addressing challenging structural variants such as hybrid genes, deletions and duplications.

genetics↗

bioRxiv2022

Goat PRP14 (gPRP14) has xeno-antigenic properties and works as a vaccine in preclinical models of cancer.

We studied the activity of recombinant goat PRP14 (gPRP14), a member of the RID protein family, as a xeno-antigen in preclinical models of cancer. Antisera from rabbits and mice immunized with gPRP14 showed strong reactivity against several tumor cell types, which was absent towards normal cells: the tumor selectivity was related to surface and intra-cellular expression in tumor cells, and to an exclusively intra-cellular localization in normal cells. In vitro, binding to tumor cells was followed by cytotoxicity which could be rescued by the addition of excess soluble antigen. In vivo, an anti-tumor activity of immunization with gPRP14 was observed in murine syngeneic models of breast cancer and melanoma: the anti-tumor response was present when gPRP14 was administered in a preventive setting, and persisted upon repeated challenges with tumor cells in long-term survivor mice. Finally, we showed that both the humoral and T-cell mediated responses are needed for the optimal anti-tumor effect in the murine melanoma model. Thus, we have performed an initial characterization of gPRP14 as a cancer vaccine, which -given the potential wide range of tumor cells positive for the antigen-appears as a promising, novel immunotherapy.

cancer biology↗

bioRxiv2021

GABAB receptor/HCN channel complexes in VTA dopamine neurons limit synaptic inhibition and prevent anxiety-like behavior

Aversive stimuli inhibiting dopamine neurons in the ventral tegmental area (DAVTA neurons) induce anxiety-like behaviors. The inhibition of DAVTA neurons is prolonged by GABAB receptor (GBR)-activated K+-currents, which exhibit a rapid desensitization of unknown physiological relevance. We now report that GBRs associate via auxiliary KCTD16 subunits with HCN channels, which facilitates activation of hyperpolarization- activated currents (Ih) by GBR-activated K+ currents. Activation of Ih underlies rapid K+ current desensitization in DAVTA neurons and limits GBR-mediated inhibition. Disruption of the GBR/HCN complex in KCTD16-/- mice or blockade of Ih prolongs optogenetically driven inhibition of DAVTA neuron firing. KCTD16-/- mice exhibit an increased anxiety-like behavior in response to stressful stimuli, which is reproduced by in vivo CRISPR/Cas9-mediated KCTD16 ablation in DAVTA neurons or intra-VTA infusion of HCN antagonist to wild-type mice. Our data reveal that GBR-induced Ih protect DAVTA neurons from prolonged GBR- mediated inhibition in response to stressors, which moderates anxiety-like behaviors.

neuroscience↗

bioRxiv2021

Zona Incerta activity dynamics underlying associative fear learning and fear generalization

Recent studies suggest that the Zona Incerta (ZI) plays a role in fear learning and recall. However, there is a clear gap in knowledge as to whether the ZI can encode fear evoking threats and cues that predict them. Here, we subject mice to a classical fear conditioning paradigm while recording the in-vivo calcium dynamics of ZI neurons. We observed that ZI neurons can encode not only a fear evoking stimulus, but can also learn to encode predictive cues, associate them with the unconditioned fear evoking stimulus, and discriminate them from neutral non-predictive cues. While the ZI across all mice learned to become mainly excited by fear predictive cues, only mice that did not generalize fear became largely inhibited by non-predictive cues and better discriminated the two. Together, we provide extensive evidence that the ZI differentially encode fear predictive and non-predictive cues in a generalization dependent manner.

neuroscience↗

bioRxiv2020

Zona Incerta subpopulations differentially encode and modulate anxiety

Despite recent clinical observations linking the Zona Incerta (ZI) to anxiety, little is known about whether and how the ZI processes anxiety. Here, we subject mice to anxious experiences and observe an increase in ZI cfos-labelled neurons and single-cell calcium activity as well as an efficient effect of ZI infusion of diazepam a classical anxiolytic drug. We further identify that somatostatin-(SOM), calretinin-(CR), and vesicular glutamate transporter-2 (Vglut2)-expressing cells display unique electrophysiological profiles; yet they similarly respond to anxiety-provoking stimuli and to diazepam. Interestingly, optogenetic manipulations reveals that each of these ZI neuronal population triggers specific anxiety-related behavioral phenotypes. Activation of SOM-expressing neurons induced anxiety while in stark contrast, photo-activation of CR-positive cells and photo-inhibition of Vglut2-expressing neurons produce anxiolysis. Furthermore, activation of CR- and Vglut2-positive cells provokes rearing and jumps; respectively. Our findings provide the first experimental evidence that ZI subpopulations encode and modulate different components of anxiety.

neuroscience↗

bioRxiv2020

Midbrain modulation of conditioned learning

The firing activity of striatal cholinergic interneurons (CINs) is a neural correlate of conditioned learning. However, its circuitry is not entirely described. Here we report that midbrain GABA and Glutamate neurons discriminate auditory cues and encode the association of a predictive stimulus with a footshock. Through their mono- and di-synaptic inputs via the thalamic-parafasicular sub-nucleus onto CINs, they contribute to processing fear learning.

neuroscience↗