Institut de Formation Doctoral

In  mammals,    the  discovery  of  melanopsin and
intrinsic  light  responsiveness in a small population  of retinal ganglion cells (ipRGCs)  paved  the  way for understanding light  regulation of nonvisual  functions, like the pupillary reflex or the entrainment of circadian rhythms.  The  scarcity of ipRGCs, however,  is a hurdle for investigating melanopsin signaling  mechanisms. In the  avian  retina, melanopsin expresses  abundantly in the outer  nuclear layer too, but its functionality is unknown. We  used  the  chicken embryo  retina  as  a model  system to investigate  possible physiological  roles of melanopsin in non-ganglion cells. A polyclonal antibody  targeting both  chicken melanopsin isoforms was validated  by Western  blot and  used  to corroborate the  distribution pattern in fixed retina  sections; strong immunoreactivity occurred in areas  that  likely include
horizontal, bipolar,  and  some  amacrine cells. Retinas were enzymatically dissociated to yield morphologically well-preserved  isolated  neurons; their  physiological viability was tested  with whole cell recording. Although voltage-gated  currents were found in the  different cell types, initial  recordings failed  to reveal direct  changes in  membrane current by photostimulation. However, light  could  reversibly modulate voltage-gated  currents. Fluorescence imaging  in cells loaded  with calcium indicators demonstrated a Ca fluorescence increase  in selected  bipolar  cells  and  small  neurons,  likely  to comprise a subtype of amacrine cells. Higher sensitivity measurements,  using  a  photomultiplier  and  pulsed light to extend the  recording period, showed  that  in a minority  of bipolar  cells, light evoked Ca responses with a long latency and a slow time course, spanning minutes. A reassessment  of  electrical  responses occurring on such a long  time  scale revealed  a small inward  current (tens    of  pA)  with  a  similar  time  course.    These observations indicate  that  intrinsic  photosensitivity is not  confined to rods,  cones,  and  some  ganglion cells, but extends to additional retinal cell types. Its physiological  role remains to be investigated.
Supported by Colciencias  grant  222852128276.

The brain's control of movement is thought to involve coordinated activity between cerebellar Purkinje cells. The results reported here demonstrate that somatic Ca 2+ imaging is a faithful reporter of Na +-dependent "simple spike" pauses and enables us to optically record changes in firing rates in populations of Purkinje cells. This simultaneous calcium imaging of populations of Purkinje cells reveals a striking spatial organization of pauses in Purkinje cell activity between neighboring cells. The source of this organization is shown to be the presynaptic GABAergic network and blocking GABA A Rs abolishes the synchrony. These data suggest that presynaptic interneurons synchronize (in)activity between neighboring Purkinje cells and thereby maximize their effect on downstream targets in the deep cerebellar nuclei.

The brain's control of movement is thought to involve coordinated activity between cerebellar Purkinje cells. The results reported here demonstrate that somatic calcium imaging is a faithful reporter of sodium-dependent "simple spike" pauses and enables us to optically record changes in firing rates in populations of Purkinje cells. This simultaneous calcium imaging of populations of Purkinje cells reveals a striking spatial organization of pauses in Purkinje cell activity between neighboring cells. The source of this organization is shown to be the presynaptic GABAergic network and blocking GABA-A receptors abolishes the synchrony. These data suggest that presynaptic interneurons synchronize (in)activity between neighboring Purkinje cells and thereby maximize their effect on downstream targets in the deep cerebellar nuclei.

NTS designed the research plan, performed all experiments and data analysis, prepared figures, and wrote the manuscript. JRB performed and analyzed electrophysiological recordings. DLP designed the research plan, performed experiments and analysis, and provided supervision. MRC designed the research plan, provided supervision, and wrote the manuscript.

This article describes the sound project "Floating Herstories" in all its levels: from its conception and development, to its trial in the field on the island of Trinidad, in the Republic of Trinidad and Tobago, West Indies. Inspired by the childhood experience of hearing a shell on Caribbean seashores, this sound work uses seashells as the medium to transmit stories narrated by women authors from Trinidad. Co-written by two authors, this article describes the multidisciplinary construction of the project, involving the participation of the three narrators of the soundtrack. Following a situated heritage of storytelling, "Floating Herstories" aims to tell stories collected poetically by the Caribbean Sea. The trial with the finished prototype conveyed the reactions of the population in situ, from which we gathered some meanings concerning the experience of both telling a story and listening to one. On this basis, the article sets out some questions about the value of silence and selected sound in a familiar or estranged context.

Vibrio alginolyticus is a ubiquitous Gram-negative bacterium which is normally distributed in the coastal and estuarine environments. It has been suggested to be an opportunistic pathogen to both marine animals and humans, Here, the completed genome sequence of V. alginolyticus ZJ-T was determined by Illumina high-throughput sequencing.

The pathogenicity of methicillin-resistant S. aureus (MRSA) strains relies on their ability to produce a wide variety of tightly regulated virulence factors. Current in vivo models to analyze host-pathogen interactions are limited and difficult to manipulate.

Staphylococcus aureus is a predominant cause of chronic lung infections. While the airway environment is rich in highly sialylated mucins, the interaction of S. aureus with sialic acid is poorly characterized. Using S. aureus USA300 as well as clinical isolates, we demonstrate that quorum-sensing dysfunction, a hallmark of S. aureus adaptation, correlates with a greater ability to consume free sialic acid, providing a growth advantage in an air-liquid interface model and in vivo. Furthermore, RNA-seq experiment reveals that free sialic acid triggers transcriptional reprogramming promoting S. aureus chronic lifestyle. To support the clinical relevance of our results, we show the co-occurrence of S. aureus, sialidase-producing microbiota and free sialic acid in the airway of patients with cystic fibrosis. Our findings suggest a dual role for sialic acid in S. aureus airway infection, triggering virulence reprogramming and driving S. aureus adaptive strategies through the selection of quorum-sensing dysfunctional strains. Staphylococcus aureus is a common opportunistic pathogen infecting the lungs of patients with chronic lung diseases such as cystic fibrosis (CF) 1-4. CF is the most common life-limiting autosomal recessive disorder in the Caucasian population 5. Respiratory epithelial cells of CF patients produce thick and sticky mucus as a consequence of defective transmembrane conductance regulator CFTR gene 6 , which impedes bacterial mucociliary clearance 7. CF patients are chronically infected by lung-adapted S. aureus clones 8-10. S. aureus adaptive mechanisms include the emergence of small colony variants (SCVs) 11-13 , mucoid phenotypes 14 , and deregulation of toxins 10,15. Agr (accessory gene regulatory) system is a quorum-sensing system involved in both virulence-and metabolism-associated gene regulation 16. The role of the Agr system in the interplay between S. aureus and host-derived metabolites has received less attention 16 compared to the regulation of virulence factors 17,18. By investigating the genetic mutations acquired during the course of chronic lung infection across multiple patients with CF, we observed a convergent evolution with genetic alterations at the agr locus leading to quorum-sensing system dysfunction 10. Indeed, Agr dysfunction, which is associated with increased biofilm formation, is a common adaptive strategy identified in chronic

Bacterial pathogens adapt and replicate within host cells, while host cells develop mechanisms to eliminate them. Using a dual proteomic approach, we characterized the intra-macrophage proteome of the facultative intracellular pathogen, Francisella novicida. More than 900 Francisella proteins were identified in infected macrophages after a 10-h infection. Biotin biosynthesis-related proteins were upregulated, emphasizing the role of biotin-associated genes in Francisella replication. Conversely, proteins encoded by the Francisella pathogenicity island (FPI) were downregulated, supporting the importance of the F. tularensis Type VI Secretion System for vacuole escape, not cytosolic replication. In the host cell, over 300 proteins showed differential expression among the 6200 identified during infection. The most upregulated host protein was cis-aconitate decarboxylase IRG1, known for itaconate production with antimicrobial properties in Francisella. Surprisingly, disrupting IRG1 expression did not impact Francisella's intracellular life cycle, suggesting redundancy with other immune proteins or inclusion in larger complexes. Over-representation analysis highlighted cell-cell contact and actin polymerization in macrophage deregulated proteins. Using flow cytometry and live cell imaging, we demonstrated that merocytophagy involves diverse cell-to-cell contacts and actin polymerization-dependent processes. These findings lay the groundwork for further exploration of merocytophagy and its molecular mechanisms in future research. Data are available via ProteomeXchange with identifier PXD035145. Abbreviations WT Francisella novicida U112 BMM Bone marrow-derived mouse macrophages CFU Colony forming unit DMEM Dulbecco's modified eagle medium IRG1 Immunoresponsive gene 1 Subsp Subspecies KEGG Kyoto encyclopedia of genes and genomes LC-MS/MS Liquid chromatography coupled to tandem mass spectrometry