Biology subjects

Piazza, S.

Publications and source records attributed to Piazza, S..

14 recordsLinked to original sources

bioRxiv2026

Haplotype specific analyses in the phased genomes era: the case of apple cv "Golden Delicious"

Apple is one of the major cultivated fruit crops in temperate regions. To better support breeding programs and facilitate the development of improved cultivars, we generated a new haplotype-resolved version of the Golden Delicious genome, one of the major founders of many modern apple lineages. The assembly features the separation of the two haplotypes, with a total size of 647.3 Mb and 649.2 Mb, respectively. The phasing was accurately validated with 10,321 curated SNPs. Telomere-to-telomere continuity was verified by the analysis of telomeric sequence composition at the end of each chromosome. Gene prediction identified a total of 45,116 genes in haplotype 1 and 45,063 genes in haplotype 2. A pangenome analysis employing 6 haplotype resolved genomes identified both common and unique gene families. The availability of a phased genome enabled the assessment of genome-wide allelic specific expression. Our case study, focusing on Md-PG1 (a key regulator of fruit softening), revealed that the allelic form on haplotype 2 (GDH2-10g24673) was the dominant contributor to total gene expression. In addition, the phased genome also showed specific miRNA chromosomal distribution patterns, as well as a distinct methylation profile. Altogether, these genomic resources provide new insights into the allelic regulation of key agronomic traits and represent a valuable tool to accelerate apple breeding.

genomics↗

bioRxiv2025

KODAMA enables self-guided weakly supervised learning in spatial transcriptomics

Spatial transcriptomics provides researchers with a powerful tool to investigate gene expression patterns within their native positional context in tissues, revealing intricate cellular relationships. However, analyzing spatial transcriptomics data presents unique challenges due to their high dimensionality and complexity. Several competitive tools have emerged, each aiming to integrate spatial dependencies into their workflow. Yet, the overall performance of these approaches remains constrained by limitations in prediction accuracy and compatibility with specific data types. Here, we introduce the third version of the KODAMA algorithm, specifically tailored for spatial transcriptomics analysis. This upgraded version incorporates a novel approach that effectively reduces data dimensionality while preserving spatial information. At its core, the KODAMA algorithm employs parallel iterations that enforce spatial constraints throughout an embedded clustering process. Our method provides the flexibility to simultaneously analyze multiple samples, streamlining analysis workflows. Additionally, it extends beyond traditional 2D analysis to accommodate multidimensional datasets, including 3D spatial information. KODAMA seamlessly integrates into various pipelines, such as Seurat and Giotto. Extensive evaluations of KODAMA on 10x Visium, Visium HD, and image-based 3D datasets demonstrate its high accuracy in spatial domain prediction. Comparative analyses against alternative dimensionality reduction techniques and spatial analysis tools consistently validate and highlight KODAMAs superior performance in unraveling the spatial organization of cellular components across both single and integrated tissue samples.

bioinformatics↗

bioRxiv2024

The DNA-repair protein APE1 participates with hnRNPA2B1 to motif-enriched and prognostic miRNA secretion

The base excision repair (BER) Apurinic/apyrimidinic endonuclease 1 (APE1) enzyme is endowed with several non-repair activities including miRNAs processing. APE1 is overexpressed in many cancers but its causal role in the tumorigenic processes is largely unknown. We recently described that APE1 can be actively secreted by mammalian cells through exosomes. However, APE1 role in EVs or exosomes is still unknown, especially regarding a putative regulatory function on small non-coding RNAs vesicular secretion. Through dedicated transcriptomic analysis on cellular and vesicular small RNAs of different APE1-depleted cancer cell lines, we found that miRNAs loading into EVs is a regulated process, dependent on APE1, distinctly conveying RNA subsets into vesicles. We identified APE1-dependent secreted miRNAs characterized by enriched sequence motifs and possible binding sites for APE1. In 33 out of 34 APE1-dependent-miRNA precursors, we surprisingly found EXO-motifs and proved that APE1 cooperates with hnRNPA2B1 for the EV-sorting of a subset of miRNAs, including miR-1246, through direct binding to GGAG stretches. Using TCGA-datasets, we showed that these miRNAs identify a signature with high prognostic significance in cancer. In summary, we provided evidence that APE1 is part of the protein cargo of secreted EVs, suggesting a novel post-transcriptional role for this ubiquitous DNA-repair enzyme that could explain its role in cancer progression.

molecular biology↗

bioRxiv2023

A common druggable signature of oncogenic CMYC, mutant KRAS and mutant p53 reveals functional redundancy and competition of the oncogenes in cancer

Major driver oncogenes CMYC, mutant KRAS and mutant TP53 often co-exist and cooperate in promoting human neoplasia. By CRISPR-Cas9-mediated downregulation we determined their proteomics and transcriptomics downstream programs in a panel of cell lines with activated either single or three oncogenes - in cancers of lung, colon and pancreas. This allowed to define and screen the oncogenes common functional program for anti-cancer target candidates, and find protocols which efficiently kill cancer cells and organoids by targeting pathways represented by a signature of three genes: RUVBL1, HSPA9 and XPO1. We found that these genes were controlled by the driver oncoproteins in a redundant or competitive manner, rather than by cooperation. Each oncoprotein individually was able to upregulate the three target genes, while upon oncogene co-expression each target was controlled preferably by a specific oncoprotein which reduced the influence of the others. Mechanistically this redundancy was mediated by parallel routes of the target gene activation - as in the case of mutant KRAS signaling to C-JUN and GLI-2 transcription factors bypassing CMYC, and by competition - as in the case of mutant p53 and CMYC competing for biding to the target promoters. The transcriptomics data from the cell lines and patient samples indicate that the redundancy of the oncogenic programs is a broad phenomenon which may comprise even a majority of the genes dependent on the oncoprotein, as shown for mutant p53 in colon and lung cancer cell lines. Nevertheless, we demonstrate that the redundant oncogene programs harbor targets of efficient anti-cancer drug combinations, bypassing limitations of a direct oncoprotein inhibition.

cancer biology↗

bioRxiv2023

A Compact and Effective Photon-Resolved Image Scanning Microscope

Fluorescence confocal laser-scanning microscopy (LSM) is one of the most popular tools for life science research. This popularity is expected to grow thanks to single-photon array detectors tailored for LSM. These detectors offer unique single-photon spatiotemporal information, opening new perspectives for gentle and quantitative super-resolution imaging. However, a flawless recording of this information poses significant challenges for the microscope data-acquisition system. Here, we present a data-acquisition module based on the digital frequency domain principle, able to record photons essential spatial and temporal features. We use this module to extend the capabilities of established imaging techniques based on single-photon avalanche diode (SPAD) array detectors, such as fluorescence lifetime image scanning microscopy. Furthermore, we use the module to introduce a robust multi-species approach encoding the fluorophores excitation spectra in the time domain. Lastly, we combine time-resolved stimulated emission depletion microscopy with image scanning microscopy, boosting spatial resolution. Our results demonstrate how a conventional fluorescence laser scanning microscope can transform into a simple, information-rich, super-resolved imaging system with the simple addition of a SPAD array detector with a tailored data acquisition system. We expected a blooming of advanced single-photon imaging techniques which effectively harness all the sample information encoded in each photon.

biophysics↗

bioRxiv2023

HuR modulation with tanshinone mimics impairs LPS response in murine macrophages

Lipopolysaccharide exposure to macrophages induces an inflammatory response that is heavily regulated at the transcriptional and post-transcriptional levels. HuR (ELAVL1) is an RNA binding protein that binds and regulates the maturation and half-life of AU/U rich elements (ARE) containing cytokines and chemokines transcripts, mediating the LPS-induced response. Here we investigated how and to what extent small molecule tanshinone mimics (TMs) inhibiting HuR-RNA interaction counteract LPS stimulus in macrophages. We show TMs exist in solution in keto-enolic tautomerism and that, by molecular dynamic calculations, the orto quinone form is the bioactive species interacting with HuR and inhibiting its binding mode vs mRNA targets. A chemical blockage of the diphenolic, reduced form as a diacetate caused the loss of activity of TMs in vitro but resulted to prodrug-like activity in vivo. The murine macrophage cell line RAW264.7 was treated with LPS and TMs, and the modulation of cellular LPS-induced response was monitored by RNA and Ribonucleoprotein immunoprecipitation sequencing. Correlation analyses indicated that LPS induced a strong coupling between differentially expressed genes and HuR-bound genes, and that TMs reduced such interactions. Functional annotation addressed a specific set of genes involved in chemotaxis and immune response, such as Cxcl10, Il1b, Cd40, and Fas, with a decreased association with HuR, a reduction of their expression and protein secretion. The same effect was observed in primary murine bone marrow-derived macrophages, and in vivo in an LPS induced peritonitis model, in which the serum level of Cxcl10 and Il1b was strongly reduced, endowing TMs such as TM7nox with remarkable anti-inflammatory properties in vivo.

molecular biology↗

bioRxiv2022

Optical-quality assessment of a miniaturized intraocular telescope

PurposeEvaluating the optical transmission and geometrical aberrations of an intraocular device, namely, the Small-Incision New Generation Implantable Miniature Telescope (SING IMT, Samsara Vision), designed to correct age-related macular degeneration. MethodsOptical transmission in the spectral range 350-750 nm of the implantable optics was recorded with a fiber-optic spectrometer. Geometrical aberrations were studied by measuring the wavefront of a laser beam after passing through the implantable optics and performing an expansion of the measured wavefront into a Zernike polynomial basis. The study was conducted under in-vitro experimental conditions. A second monofocal intraocular lens (SY60WF, Alcon) was tested and used as reference for assessing the optical quality of the SING IMT device. ResultsSpectroscopy measurements revealed that the SING IMT and monofocal IOL element feature UV-rejection and blue-rejection capabilities, respectively. Wavefront concavity indicated that the SING IMT behaves as a diverging lens with a focal length of approximately -100 mm; Zernike analysis showed that SING IMT has negligible coma, trefoil, astigmatism, and spherical aberrations of any order and along any direction. ConclusionsThe SING IMT exhibited even optical transmission in the whole visible spectrum and curvature capable of magnifying the retinal images without introducing geometrical aberrations, which proves the feasibility of this device as high-quality optical element for imaging. The rigidity of the compound lens of the SING IMT prevents mechanically-induced distortions, an issue encountered with polymeric lenses. Translational RelevanceSpectrometry and in vitro wavefront analysis provide evidence supporting the new generation miniaturized telescopic intraocular lens as a favorable option to intraocular implant in age-related macular degeneration.

biophysics↗

bioRxiv2022

Focus-ISM for Sharp and Gentle Super-Resolved Microscopy

Super-resolution microscopy is routinely used for fixed and thin samples, while its feasibility for imaging live and thick samples is still limited. In the case of stimulated emission depletion (STED) microscopy, the high-intensity illumination required to achieve effective sub-diffraction resolution can introduce photo-damage, thus reducing the compatibility of the technique with live-cell imaging. Moreover, the out-of-focus fluorescence background may overcome the often faint signal stemming from the focal point, thus constraining imaging to thin samples. Here, we combined STED microscopy with image-scanning microscopy (ISM) to mitigate these limitations without any practical disadvantages. We first enhanced a laser scanning microscope (LSM) by introducing a detector array, hence providing access to a set of additional spatial information that is not available with a typical single-element detector. Then, we exploited this extended dataset to implement focus-ISM, a novel method that relaxes the high-intensity requirement of STED microscopy and removes the out-of-focus background. Additionally, we generalized the focus-ISM method to conventional LSM, namely without a STED beam. The proposed approach requires minimal architectural changes compared with conventional STED microscopes but provides substantial advantages for live and thick sample imaging while maintaining all compatibility with all recent advances in STED and confocal microscopy. As such, focus-ISM represents an essential step towards a universal super-resolved LSM technique for subcellular imaging.

biophysics↗

bioRxiv2021

Defective linear and circular RNAs biogenesis in Huntington's disease: CAG repeat expansion hijacks neuronal splicing

Alternative splicing (AS) appears to be altered in Huntingtons disease (HD), but its significance for early, pre-symptomatic disease stages has not been inspected. Here, taking advantage of Htt CAG knock-in mouse in vitro and in vivo models, we demonstrate a strong correlation between Htt CAG repeat length and increased aberrant linear AS, specifically affecting neural progenitors and, in vivo, the striatum prior to overt behavioral phenotypes stages. Remarkably, expanded Htt CAG repeats reflect on a previously neglected, global impairment of back-splicing, leading to decreased circular RNAs production in neural progenitors. Though the mechanisms of this dysregulation remain uncertain, our study unveils network of transcriptionally altered micro-RNAs and RNA-binding proteins (CELF, hnRNPS, PTBP, SRSF) which, in turn, might influence the AS machinery, primarily in neural cells. We suggest that this unbalanced expression of linear and circular RNAs might result in altered neural fitness, contributing to HD striatal vulnerability.

genetics↗

bioRxiv2021

Plasticity of the gastrocnemius elastic system in response to decreased work and power demand during growth

Elastic energy storage and release can enhance performance that would otherwise be limited by the force-velocity constraints of muscle. While functional influence of a biological spring depends on tuning between components of an elastic system (the muscle, spring, driven mass, and lever system), we do not know whether elastic systems systematically adapt to functional demand. To test whether altering work and power generation during maturation alters the morphology of an elastic system, we prevented growing guinea fowl (Numida Meleagris) from jumping. At maturity, we compared the jump performance of our treatment group to that of controls and measured the morphology of the gastrocnemius elastic system. We found that restricted birds jumped with lower jump power and work, yet there were no significant between-group differences in the components of the elastic system. Further, subject-specific models revealed no difference in energy storage capacity between groups, though energy storage was most sensitive to variations in muscle properties (most significantly operating length and least dependent on tendon stiffness). We conclude that the gastrocnemius elastic system in the guinea fowl displays little to no plastic response to decreased demand during growth and hypothesize that neural plasticity may explain performance variation.

physiology↗

bioRxiv2021

The Arabidopsis pattern recognition receptor EFR enhances fire blight resistance in apple

Fire blight disease, caused by the bacterium Erwinia amylovora (E. amylovora), is responsible for substantial losses in cultivated apple worldwide. An important mechanism of plant immunity is based on the recognition of conserved microbial molecules, named pathogen- or microbe-associated molecular patterns (PAMPs or MAMPs), through pattern recognition receptors (PRRs), leading to pattern-triggered immunity (PTI). The interspecies transfer of PRRs represents a promising strategy to engineer broad spectrum and durable disease resistance in crops. EFR, the Arabidopsis thaliana PRR for the PAMP elf18 derived from the elongation factor thermal unstable (EF-Tu) proved to be effective in improving bacterial resistance when expressed into Solanaceae and other plant species. In this study, we tested whether EFR can affect the interaction of apple with E. amylovora by its ectopic expression in the susceptible apple rootstock M.26. Stable EFR expression led to the activation of PAMP-triggered immune response in apple leaves upon treatment with supernatant of E. amylovora, as measured by production of reactive oxygen species and the induction of known defense genes. The amount of tissue necrosis associated with E. amylovora infection was significantly reduced in the EFR transgenic rootstock compared to the wild-type. Our results show that the expression of EFR in apple rootstock may be a valuable biotechnology strategy to improve the resistance of apple to fire blight.

plant biology↗

bioRxiv2020

Stop codon readthrough of a POU transcription factor regulates steroidogenesis and developmental transitions

Translational stop codon readthrough generates C-terminally extended protein isoforms. While evidence mounts of readthrough as a global phenomenon, proofs of its functional consequences are scarce. We show that readthrough of the mRNA for the Drosophila POU/Oct transcription factor Drifter occurs at a high rate and in a spatiotemporal manner in vivo, reaching above 50% in the prothoracic gland. Phylogenetic analyses suggested that readthrough of drifter is conserved among Dipterans, with C-terminal extensions harboring intrinsically disordered regions, and amino acids streches implied in transcriptional activation. The C-terminally extended Drifter isoform is required for maintaining normal levels of the growth hormone ecdysone through regulation of its biosynthetic genes, acting in synergy with the transcription factor Molting defective. A 14-bp deletion that abolished readthrough, caused prolonged larval development and delayed metamorphosis. This study provides a striking example of alternative genetic decoding that feeds into the progression from one life cycle stage to another.

molecular biology↗

bioRxiv2020

CHD8 Suppression Impacts on Histone H3 Lysine 36 Trimethylation and Alters RNA Alternative Splicing.

Disruptive mutations in the chromodomain helicase DNA binding protein 8 (CHD8) have been recurrently associated with Autism Spectrum Disorders (ASD). In normal cellular physiology, CHD8 co-purifies with MLL1 and MOF transcriptional activation complex, with elongating RNAPII and directly binds to DNA promoters and enhancers regions, thus a regulatory role in transcriptional initiation and elongation could be postulated. Here we investigated how chromatin landscape reacts to CHD8 suppression by analyzing a panel of histone modifications in induced pluripotent stem cell-derived neural progenitors. We interrogated transcriptionally active and repressed regions, as well as active and poised enhancers. CHD8 suppression led to significant reduction (47.82%) in histone H3K36me3 peaks at gene bodies, particularly impacting on transcriptional elongation chromatin states. H3K36me3 reduction specifically affects highly expressed, CHD8-bound genes. Histone H3K36me3 reduction associated to CHD8-suppression does not functionally impact on global transcriptional levels, but correlated with altered alternative splicing patterns of [~] 2000 protein coding genes implicated in "RNA splicing", "mitotic cell cycle phase transition" and "mRNA processing", especially affecting alternative first exon and exon skipping events. In summary, our results point toward broad molecular consequences of CHD8 suppression, implicating altered histone deposition/maintenance and RNA processing regulation as important regulatory processes in ASD.

genetics↗

bioRxiv2019

Expression of telomerase prevents ALT and maintains telomeric heterochromatin in juvenile brain tumors.

The activation of a telomere maintenance mechanism (TMM) is an essential step in cancer progression to escape replicative senescence and apoptosis. Paediatric brain tumors frequently exhibit Alternative Lengthening of Telomere (ALT) as active TMM, but the mechanisms involved in the induction of ALT in brain tumor cells are not clear.\n\nHere, we report a model of juvenile zebrafish brain tumor that progressively develops ALT. We discovered that reduced expression of tert and increase in Terra expression precedes ALT development. Additionally, tumors show persistent telomeric DNA damage and loss of heterochromatin marks at chromosome ends. Surprisingly, expression of telomerase reverts ALT features. Comparative analysis of gene expression after the rescue of ALT with telomerase and analysis of telomerase positive paediatric brain cancers showed increase of telomeric heterochromatin and maintenance of telomere length compared to ALT tumors, with reduced expression of genes of the pre-replicative complex as hallmark. Thus our study identifies telomere maintenance mechanisms as major drivers of telomeric DNA replication and chromatin status in brain cancers.

cancer biology↗