Biology subjects

Santoni, M.

Publications and source records attributed to Santoni, M..

4 recordsLinked to original sources

bioRxiv2026

Cyclin B phosphorylation by PKA safeguards the G2-arrest in vertebrate oocytes

Entry into meiotic M-phase in vertebrate oocytes requires activation of Cdk1-Cyclin B, which is restrained by the cAMP-PKA signalling during prophase arrest. Although Cdc25C and ARPP19 are established PKA substrates in this context, they do not fully explain how PKA represses Cdk1 activation. Here we used Turbo-ID approach using catalytic PKA (PKAc) as the bait to identify substrates in Xenopus laevis oocytes. Unexpectedly, this screen identified Cyclin B2, the regulatory subunit of Cdk1, as a PKA-proximal protein. Although Cyclin B2 does not stably associate with PKAc, it harbors a conserved PKA motif around Serine 271 within Cyclin Box 2. We show that PKAc phosphorylates Cyclin B2 at S271 both in vivo and in vitro. Functionally, a phosphomimic Cyclin B2 mutant at this site fails to induce meiotic maturation and Cdk1 activation. S271 phosphorylation neither alters Cyclin B2 stability nor its binding to Cdk1 in vivo. S271 phosphorylation does not impair Cdk1 activity toward a single-site substrate, such as PP1, but slows Cyclin-B-dependent multisite phosphorylation of Cdc25C, a key regulator of the Cdk1 activation network. These findings identify Cyclin B as a direct and conserved PKA target, revealing the mechanistic link between PKA activity and Cdk1-Cyclin B repression that maintains oocyte prophase arrest.

cell biology↗

bioRxiv2026

Sleep deprivation impairs information processing via dysregulation of chloride homeostasis in the prefrontal cortex

Sleep deprivation (SD) impairs information processing through alterations of prefrontal cortex (PFC) function, yet the molecular underpinnings of this process remain poorly understood. We previously showed that SD disrupts sensorimotor gating by elevating prefrontal levels of the neurosteroid allopregnanolone (AP), a positive allosteric modulator of GABA-A receptors. Here we identify a complementary, mechanistically independent process whereby SD alters GABA-A currents in the PFC of mice and rats. SD reduced membrane expression of the chloride exporter KCC2, leading to intracellular chloride accumulation and a depolarizing shift in GABA-A receptor reversal potential that weakened GABAergic inhibition. Pharmacological normalization of chloride homeostasis with bumetanide fully rescued SD-induced deficits in sensorimotor gating and information encoding. SD also upregulated BDNF, and intra-PFC antagonism of its receptor TrkB restored KCC2 expression and normalized information processing, identifying BDNF-TrkB signaling as an upstream driver of chloride dysregulation. Notably, blocking AP synthesis rescued behavioral deficits without correcting chloride imbalance, confirming mechanistic independence. Finally, combined administration of AP and a KCC2 blocker produced information-processing deficits akin to those induced by SD. These findings identify TrkB-dependent disruption of prefrontal chloride homeostasis as a druggable mechanism underlying sleep loss-induced cognitive dysfunction.

animal behavior and cognition↗

bioRxiv2024

Enhanced Intestinal Epithelial Co-Culture Model with Orbital Mechanical Stimulation: A Proof-of-Concept Application in Food Nanotoxicology

IntroductionCurrent in vitro intestinal models lack the mechanical forces present in the physiological environment, limiting their reliability for nanotoxicology studies. Here, we developed an enhanced Caco-2/HT29-MTX-E12 co-culture model incorporating orbital mechanical stimulation to better replicate intestinal conditions and investigate nanoparticle interactions. MethodsWe established co-cultures under static and dynamic conditions, validating their development through multiple approaches including barrier integrity measurements, gene expression analysis, and confocal microscopy. We introduced novel quantitative analysis of dome formation as a differentiation marker and demonstrated the model application by investigating cellular responses to titanium dioxide (TiO) nanoparticles in a digested food matrix. ResultsDynamic conditions accelerated epithelial differentiation, achieving functional barrier properties by day 14 rather than day 21, with enhanced mucin production and more organized three-dimensional structure. Mechanical stimulation selectively promoted goblet cell differentiation without affecting general epithelial markers. The optimized model successfully detected concentration-dependent oxidative stress responses to TiO exposure, revealing cellular dysfunction preceding membrane damage. DiscussionThis improved co-culture system provides a better physiological platform for nanotoxicology studies. By incorporating mechanical forces, each cell type exhibits more representative behavior, creating a more realistic experimental setup. The model bridges the gap between simple monocultures and complex 3D systems, offering a practical approach for investigating nanoparticle-epithelium interactions in a food-relevant context.

cell biology↗

bioRxiv2022

Structure of the photosynthetic Calvin-Benson-Bassham sedoheptulose-1,7-bisphosphatase SBPase from the model microalga Chlamydomonas reinhardtii

The Calvin-Benson-Bassham cycle (CBBC) performs carbon fixation in photosynthetic organisms. Among the eleven enzymes that participate in the pathway, sedoheptulose-1,7-bisphosphatase (SBPase) is expressed in photo-autotrophs and catalyzes the hydrolysis of sedoheptulose-1,7- bisphosphate (SBP) to sedoheptulose-7-phosphate (S7P). SBPase, along with nine other enzymes in the CBBC, contributes to the regeneration of ribulose-1,5-bisphosphate, the carbon-fixing co- substrate used by ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco). The metabolic role of SBPase is restricted to the CBBC, and a recent study revealed that the three-dimensional structure of SBPase from the moss Physcomitrium patens was found to be similar to that of fructose-1,6- bisphosphatase (FBPase), an enzyme involved in both CBBC and neoglucogenesis. In this study we report the first structure of an SBPase from a chlorophyte, the model unicellular green microalga Chlamydomonas reinhardtii. By combining experimental and computational structural analyses, we describe the topology, conformations and quaternary structure of Chlamydomonas reinhardtii SBPase (CrSBPase). We identify active site residues and locate sites of redox- and phospho-post- translational modifications that contribute to enzymatic functions. Finally, we observe that CrSBPase adopts distinct oligomeric states that may dynamically contribute to the control of its activity.

biochemistry↗