Porta de la Riva, Montserrat et al. (2021) International Worm Meeting "Functional photon-based neurotransmission in a nociceptive avoidance circuit"

Sanfeliu Cerdan, Neus, Porta de la Riva, Montserrat, Krieg, Michael, Catala Castro, Frederic, Gonzalez, Adriana

[International Worm Meeting, 2021]

Communication between cells is essential during development, homeostasis and nervous system function. The distribution of information is carried out by secreted chemicals, which are released and perceived by different cells. Defects in neurotransmission can lead to neurological disorders and lack of adaptation of the surrounding environment. Here, we have developed a synthetic, photon-based signaling system that we used to overcome synaptic transmission defects in C. elegans, restoring the communication between nervous cells. In particular, we have replaced a conditional defect for glutamate release in the nociceptor neuron ASH with a calcium sensitive light emitting enzyme (enhanced Nanolantern). To prove functionality, we built an optimized microscope capable of detecting how cellular calcium increase triggers light production in vivo. Subsequently, light emission is coupled to downstream interneurons (AVA, AIB) expressing an ultrasensitive light-gated ion channel (Channelrhodopsin-XXL). Finally, we developed a new microfluidic device, which we call Trap'N'Slap, with the aim to deliver mechanical stimuli directly to ASH, while recording calcium transients (by means of Genetically Encoded Calcium Indicators) directly from AVA and AIB. This, together with behavioral nose touch assays, proved that quantum release from ASH, is able to recover functional communication between neurons in an endogenous circuit.

Sanfeliu-Cerdan, Neus et al. (2021) International Worm Meeting "Transducing touch by a titin-related protein in the worm"

Sanfeliu-Cerdan, Neus, Catala-Castro, Frederic, Krieg, Michael, Porta-de-la-Riva, Montserrat

[International Worm Meeting, 2021]

Of the five traditional senses, touch is the first one available to us when we are born. Despite being a relatively little investigated sensory modality, touch governs our daily lives, from walking and feeding to kissing and cuddling. How mechanical forces reach the mechanoelectrical transduction channels remains an open question. To address this knowledge gap, we study the mechanotransduction pathway of C. elegans' gentle touch, which relies on the six touch receptor neurons that tile the body into two receptive fields. We showed that a protein-protein interaction motif at the MEC-2 C-terminus is critical for mechanosensation, without interfering with its localization. To understand the function of MEC-2 C-terminus in touch, we conditionally coupled it to different components of the cytoskeleton. This artificial coupling was able to recover nearly wildtype functions of MEC-2, implying a role in force transfer. We then combined genetic engineering, microfluidics, and FRET-based molecular tension microscopy and found that MEC-2 is under tension when a force is applied to the body wall of the worm. To uncover an endogenous binding partner of MEC-2, we performed a neuronal RNAi screening. Surprisingly, interference of unc-89/obscn, which encodes for a protein related to the giant human titin, led to decreased touch response and colocalizes with wt but not mutant MEC-2 in TRNs. In summary, our study sheds light into how mechanical forces propagate to the mechanosensitive channels and show the first example for titin in neuronal mechanotransduction.

Aristizabal, David et al. (2010) C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany "The abnormal embryonic partitioning gene par-5 regulates replication stress-induced checkpoint response and cell proliferation in C. elegans germline"

Aristizabal, David, Schwartz, Simo, FONTRODONA, Laura, Porta-de-la-Riva, Montserrat, CERON, Julian

[C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany, 2010]

After DNA damage, cells respond through surveillance mechanisms in order to maintain the genomic stability by activating signal transduction cascades, so-called checkpoints. Such mechanisms promote mainly the inhibition of the cell cycle progression, repair of the damaged DNA and induction of apoptotic cell death if DNA is not repaired. In C. elegans germline, DNA damage and replicative stress (e.g. Hydroxyurea (HU) treatment) trigger a checkpoint response through conserved pathways. In our lab, by screening for human genes whose expression was induced after HU treatment (Guerra et al, unpublished results), we identified two genes coding for 14-3-3 proteins (Z and E) that are homologues of the C. elegans gene par-5. The partitioning defective PAR proteins regulate the asymmetric cell division that takes place at the first embryonic division. Interestingly, PAR-5 is also present in the adult germline, but its function there remains unknown. By using a HU-induced cell cycle arrest model in the germline we found that par-5 is required for a complete DNA damage checkpoint response. par-5 inactivation (by RNAi or mutation) severally impairs the cell cycle arrest induced by the DNA replication stress. Moreover, such inactivation also affects cell proliferation in the germline precursors as was evidenced by a significant decrease in the number of mitotic cells after RNAi knockdown. Supporting the function of par-5 in the checkpoint, we have observed a higher acc umulation of DNA repair foci (labeled with a HUS-1::GFP reporter) in germ cells after par-5 RNAi. This role in the DNA damage response is not general for par genes since we have observed that RNAi depletion of par-2 and par-3 do not affect the checkpoint response. The molecular mechanism ruling the function of PAR-5 in the HU-induced checkpoint will be discussed at the meeting.

Felix MA et al. (2002) Hermann, Editeurs des Sciences et des Arts. Paris, France. "Caenorhabditis elegans. Un organisme modele en biologie."

Labouesse M, Segalat L, Felix MA

[Hermann, Editeurs des Sciences et des Arts. Paris, France., 2002]

L'espce Caenorhabditis elegans fut dcrite en 1900 Alger par E. Maupas, qui s'intressait son mode de reproduction hermaphrodite. Plus tard, vers le milieu du vingtime sicle, V. Nigon et ses collaboratuers Lyon tudirent les reorganizations cellulaires accompagnant la fecundation et les premiers clivages. J. Brun isola les preiers mutants morpholgiques.

Dessauges C et al. (2017) Dev Cell "Developmental ERK Signaling Goes Digital."

Pertz O, Dessauges C

[Dev Cell, 2017]

Reporting in Developmental Cell, de la Cova etal. (2017) present a biosensor to measure ERK activity dynamics in C.elegans larvae. They find that fate decision signaling involves frequency-modulated, digital ERK activity pulses. These findings may explain how graded morphogen signals are converted into precise and robust cell fate patterns.

Chen TC et al. (2023) Prostaglandins Leukot Essent Fatty Acids "Effect of omega-6 linoleic acid on neurobehavioral development in Caenorhabditis elegans."

Chao HR, Lai YR, Chen TC, Tsai MH, Wu CY, Hsu WL, Chen CH

[Prostaglandins Leukot Essent Fatty Acids, 2023]

Linoleic acid (LA, omega-6), an essential polyunsaturated fatty acid, is supplied by vegetable oils such as corn, sunflower and soybean. Supplementary LA in infants and children is required for normal growth and brain development, but has also been reported to induce brain inflammation and neurodegenerative diseases. This controversial role of LA development requires further investigation. Our study utilized Caenorhabditis elegans (C. elegans) as a model to clarify the role of LA in regulating neurobehavioral development. A mere supplementary quantity of LA in C. elegans larval stage affected the worm's locomotive ability, intracellular ROS accumulation and lifespan. We found that more serotonergic neurons were activated by supplementing LA above 10 μM thereby promoting locomotive ability with upregulation of serotonin-related genes. Supplementation with LA above 10 μM also inhibited the expression of mtl-1, mtl-2 and ctl-3 to accelerate oxidative stress and attenuate lifespan in nematodes; however, enhancement of stress-related genes such as sod-1, sod-3, mtl-1, mtl-2 and cyp-35A2 by supplementary LA under 1 μM decreased oxidative stress and increased the worm's lifespan. In conclusion, our study reveals that supplementary LA possesses both pros and cons in worm physiology and provides new suggestions for LA intake administration in childhood.

Eberhardt AG et al. (2008) Vet Parasitol "The Strongyloides (Nematoda) of sheep and the predominant Strongyloides of cattle form at least two different, genetically isolated populations."

Mayer WE, Eberhardt AG, Streit A, Bonfoh B

[Vet Parasitol, 2008]

Strongyloides sp. (Nematoda) are very wide spread small intestinal parasites of vertebrates that can form a facultative free-living generation. Most authors considered all Strongyloides of farm ruminants to belong to the same species, namely Strongyloides papillosus (Wedl, 1856). Here we show that, at least in southern Germany, the predominant Strongyloides found in cattle and the Strongyloides found in sheep belong to separate, genetically isolated populations. While we did find mixed infections in cattle, one form clearly dominated. This variety, in turn, was never found in sheep, indicating that the two forms have different host preferences. We also present molecular tools for distinguishing the two varieties, and an analysis of their phylogenetic relationship with the human parasite Strongyloides stercoralis and the major laboratory model species Strongyloides ratti. Based on our findings we propose that Strongyloides from sheep and the predominant Strongyloides from cattle should be considered separate species as it had already been proposed by [Brumpt, E., 1921. Recherches sur le determinisme des sexes et de l''evolution des Anguillules parasites (Strongyloides). Comptes rendu hebdomadaires des seances et memoires de la Societe de Biologie et de ses filiales 85, 149-152], but was largely ignored by later authors. For nomenclature, we follow [Brumpt, E., 1921. Recherches sur le determinisme des sexes et de l''evolution des Anguillules parasites (Strongyloides). Comptes rendu hebdomadaires des seances et memoires de la Societe de Biologie et de ses filiales 85, 149-152] and use the name S. papillosus for the Strongyloides of sheep and the name Strongyloides vituli for the predominant Strongyloides of cattle.

Fontrodona, Laura et al. (2010) C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany "Diverse functions in development of RSR-2, the Caenorhabditis elegans homolog of the human spliceosomal protein SRm300/SRRM2"

CERON, Julian, Villanueva, Alberto, Moran, Tomas, Porta-de-la-Riva, Montserrat, Schwartz, Simo, FONTRODONA, Laura, Diaz, Monica

[C. elegans: Development and Gene Expression, EMBL, Heidelberg, Germany, 2010]

Protein components of the splicing machinery are highly conserved in eukaryotes and regulate cellular and developmental processes at multiple levels. Despite the macromolecular complex nature of the spliceosome, there is a vast amount of evidence supporting the specific functions of its individual components. Previously, rsr-2, a gene encoding an SR-like protein homologous to the human splicing factor SR m300, was identified as one of the genes genetically interacting with lin-35 Rb. Recent studies in yeast have demonstrated that depletion of the SRm300 ortholog (Cwc21) produces the reduction of general transcript levels and that Cwc21p binds to several other splicing-related proteins. Through RNAi assays, we found that rsr-2 inactivation causes masculinisation of the germline. Such defect is produced by the failure to switch the germ cell fate from sperm to oocytes during germline development, consequently leading to the excess of sperm and the absence of oocytes. Our genetic epistasis analysis and reporter assays unraveled the location of rsr-2 in the functional network regulating germline sex determination. To further investigate the functional mechanisms of rsr-2, we have observed by in situ hybridization that rsr-2 mRNA is present all along the germline but enriched in its proximal part. We have also generated several transgenic reporter strains to study the regulation of rsr-2 expression in detail. Additionally, we extracted RNA from synchronized rsr-2(RNAi) animals and hybridized the corresponding cDNA on tiling arrays. Preliminary analysis of these arrays has sorted out a subset of genes whose transcript levels are significantly altered. Finally, upon our request, the National Bio-resource Project has isolated two mutations in rsr-2, tm2607 and tm2625, which are deletions producing viable and early larval arrested animals respectively. Characterization of rsr-2(tm2625) larvae indicates that rsr-2 is a gene required for cell differentiation and animal growth.

Lavatelli A et al. (2020) J Biol Chem "Defining Caenorhabditis elegans as a model system to investigate lipoic acid metabolism."

de Mendoza D, Lavatelli A, Mansilla MC

[J Biol Chem, 2020]

Lipoic acid (LA) is a sulfur-containing cofactor that covalently binds to a variety of cognate enzymes that are essential for redox reactions in all three domains of life. Inherited mutations in the enzymes that make LA, namely lipoyl synthase, octanoyltransferase and amidotransferase, result in devastating human metabolic disorders. Unfortunately, because many aspects of this essential pathway are still obscure, available treatments only serve to alleviate symptoms. We envisioned that the development of an organismal model system might provide new opportunities to interrogate LA biochemistry, biology, and physiology. Here we report our investigations on three <i>Caenorhabditis elegans</i> orthologous proteins involved in this post-translational modification. We established that M01F1.3 encodes a lipoyl synthase, ZC410.7 an octanoyltransferase and C45G3.3 an amidotransferase. Worms subjected to RNAi against <i>M01F1.3</i> and <i>ZC410.7</i> manifest larval arrest in the second generation. The arrest was not rescued by LA supplementation, indicating that endogenous synthesis of LA is essential for <i>C. elegans</i> development. Expression of the enzymes M01F1.3, ZC410.7 and C45G3.3 completely rescue bacterial or yeast mutants affected in different steps of the lipoylation pathway, indicating functional overlap. Thus, we demonstrate that, similarly to humans, <i>C. elegans</i> is able to synthesize LA de novo via a lipoyl-relay pathway, and suggest this nematode could be a valuable model to dissect the role of protein mislipoylation and to develop new therapies.

Das, Ravi et al. (2021) International Worm Meeting "Local compression mechanosensing by DVA proprioceptors curbs body bends during locomotion"

Sanfeliu-Cerdan, Neus, Porta-de-la-Riva, Montserrat, Catala-Castro, Frederic, Krieg, Michael, Pidde, Aleksandra, Das, Ravi, Lin, Li-Chun

[International Worm Meeting, 2021]

Coordinated cell shape changes is the basis for a myriad of dynamic processes in animals, from limb movement to locomotion, but also for visceral morphodynamics such as lung expansion. These processes are supervised by specialized, proprioceptive mechanosensors, which are capable to accurately transduce mechanical information from organ deformation into biochemical signals. However, we still have little knowledge about the physiologically relevant mechanical stresses and deformations that lead to the activation of mechanosensitive neurons during proprioception or visceral mechanosensation. The stereotypical locomotion phenotype of C elegans can help us elucidate several mechanotransduction pathways coming into play for proprioception. We used a conditional knock-out strategy and found that unc-70 beta-spectrin has cell-specific roles in the DVA neuron in limiting body bending amplitudes during locomotion. By means of calcium imaging, microfluidic manipulation and genetically encoded tension sensors we found that the spectrin cytoskeleton is able to bear compressive stresses that are transduced by the trp-4 ion channel of the TRPN/NOMPC family. In conjunction with optical tweezer based force spectroscopy assays of cultured DVA neurons, we found that the potassium leak channel twk-16 is responsible for inhibiting depolarization during axonal stretch, which adjusts the proprioceptive mechanism of DVA within compression. Finally, we formulate our observations within a neuromechanical framework that shows that compartmentalized, compressive mechanosensitivity triggers local muscle contraction that is critical for animal locomotion.