All Publications


  • Morphometric Analyses of Macrophages. Methods in molecular biology (Clifton, N.J.) Hansen, J. N. 2024; 2713: 519-541

    Abstract

    Cell morphology and motility drive the cellular capabilities to interact with the environment. For example, microglia, the longest known tissue-resident macrophages, show a highly branched process tree with which they continuously scan their environment. Computational image analysis allows to quantify morphology and/or motility from images of tissue-resident macrophages. Here, I describe a step-by-step protocol for analyzing the morphology (and motility) of macrophages with our recently described, freely available software MotiQ, which provides a broad band of parameters and thereby serves as a versatile tool for studies of morphology and motility.

    View details for DOI 10.1007/978-1-0716-3437-0_34

    View details for PubMedID 37639145

    View details for PubMedCentralID 7961142

  • Opposing roles of resident and infiltrating immune cells in the defence against Legionella longbeachae via IL-18R/IFN-γ/ROS axis in mice. Mucosal immunology Oberkircher, L. M., Scheiding, V. M., Linda Rafeld, H., Hanssen, E., Hansen, J. N., Fleischmann, M. J., Kessler, N., Pitsch, D., Wachten, D., Kastenmüller, W., Brown, A. S., Hartland, E. L., van Driel, I. R., Ng, G. Z., Garbi, N. 2024

    Abstract

    The immune response against Legionella longbeachae, a causative agent of the often-fatal Legionnaires' pneumonia, is poorly understood. Here we investigated the specific roles of tissue-resident alveolar macrophages (AM) and infiltrating phagocytes during infection with this pathogen. AM were the predominant cell type that internalized bacteria one day after infection. Three and five days after infection, AM numbers were greatly reduced while there was an influx of neutrophils and later monocyte-derived cells (MC) into lung tissue. AM carried greater numbers of viable L.longbeachae than neutrophils and MC, which correlated with a higher capacity of L.longbeachae to translocate bacterial effector proteins required for bacterial replication into the AM cytosol. Cell ablation experiments demonstrated that AM promoted infection whereas neutrophils and MC were required for efficient bacterial clearance. IL-18 was important for IFN-γ production by IL-18R+ NK cells and T cells which, in turn, stimulated ROS-mediated bactericidal activity in neutrophils resulting in restriction of L.longbeachae infection. Ciliated bronchiolar epithelial cells also expressed IL-18R but did not play a role in IL-18-mediated L.longbeachae clearance. Our results have identified opposing innate functions of tissue-resident and infiltrating immune cells during L.longbeachae infection that may be manipulated to improve protective responses.

    View details for DOI 10.1016/j.mucimm.2024.05.001

    View details for PubMedID 38750967

  • Human fertilization in vivo and in vitro requires the CatSper channel to initiate sperm hyperactivation. The Journal of clinical investigation Young, S., Schiffer, C., Wagner, A., Patz, J., Potapenko, A., Herrmann, L., Nordhoff, V., Pock, T., Krallmann, C., Stallmeyer, B., Ropke, A., Kierzek, M., Biagioni, C., Wang, T., Haalck, L., Deuster, D., Hansen, J. N., Wachten, D., Risse, B., Behre, H. M., Schlatt, S., Kliesch, S., Tuttelmann, F., Brenker, C., Strunker, T. 2024; 134 (1)

    Abstract

    The infertility of many couples rests on an enigmatic dysfunction of the man's sperm. To gain insight into the underlying pathomechanisms, we assessed the function of the sperm-specific multisubunit CatSper-channel complex in the sperm of almost 2,300 men undergoing a fertility workup, using a simple motility-based test. We identified a group of men with normal semen parameters but defective CatSper function. These men or couples failed to conceive naturally and upon medically assisted reproduction via intrauterine insemination and in vitro fertilization. Intracytoplasmic sperm injection (ICSI) was, ultimately, required to conceive a child. We revealed that the defective CatSper function was caused by variations in CATSPER genes. Moreover, we unveiled that CatSper-deficient human sperm were unable to undergo hyperactive motility and, therefore, failed to penetrate the egg coat. Thus, our study provides the experimental evidence that sperm hyperactivation is required for human fertilization, explaining the infertility of CatSper-deficient men and the need of ICSI for medically assisted reproduction. Finally, our study also revealed that defective CatSper function and ensuing failure to hyperactivate represents the most common cause of unexplained male infertility known thus far and that this sperm channelopathy can readily be diagnosed, enabling future evidence-based treatment of affected couples.

    View details for DOI 10.1172/JCI173564

    View details for PubMedID 38165034

  • Cylicins are a structural component of the sperm calyx being indispensable for male fertility in mice and human ELIFE Schneider, S., Kovacevic, A., Mayer, M., Dicke, A., Arevalo, L., Koser, S. A., Hansen, J. N., Young, S., Brenker, C., Kliesch, S., Wachten, D., Kirfel, G., Struenker, T., Tuettelmann, F., Schorle, H. 2023; 12

    Abstract

    Cylicins are testis-specific proteins, which are exclusively expressed during spermiogenesis. In mice and humans, two Cylicins, the gonosomal X-linked Cylicin 1 (Cylc1/CYLC1) and the autosomal Cylicin 2 (Cylc2/CYLC2) genes, have been identified. Cylicins are cytoskeletal proteins with an overall positive charge due to lysine-rich repeats. While Cylicins have been localized in the acrosomal region of round spermatids, they resemble a major component of the calyx within the perinuclear theca at the posterior part of mature sperm nuclei. However, the role of Cylicins during spermiogenesis has not yet been investigated. Here, we applied CRISPR/Cas9-mediated gene editing in zygotes to establish Cylc1- and Cylc2-deficient mouse lines as a model to study the function of these proteins. Cylc1 deficiency resulted in male subfertility, whereas Cylc2-/-, Cylc1-/yCylc2+/-, and Cylc1-/yCylc2-/- males were infertile. Phenotypical characterization revealed that loss of Cylicins prevents proper calyx assembly during spermiogenesis. This results in decreased epididymal sperm counts, impaired shedding of excess cytoplasm, and severe structural malformations, ultimately resulting in impaired sperm motility. Furthermore, exome sequencing identified an infertile man with a hemizygous variant in CYLC1 and a heterozygous variant in CYLC2, displaying morphological abnormalities of the sperm including the absence of the acrosome. Thus, our study highlights the relevance and importance of Cylicins for spermiogenic remodeling and male fertility in human and mouse, and provides the basis for further studies on unraveling the complex molecular interactions between perinuclear theca proteins required during spermiogenesis.

    View details for DOI 10.7554/eLife.86100

    View details for Web of Science ID 001134632000001

    View details for PubMedID 38013430

    View details for PubMedCentralID PMC10684152

  • Novel analytical tools reveal that local synchronization of cilia coincides with tissue-scale metachronal waves in zebrafish multiciliated epithelia. eLife Ringers, C., Bialonski, S., Ege, M., Solovev, A., Hansen, J. N., Jeong, I., Friedrich, B. M., Jurisch-Yaksi, N. 2023; 12

    Abstract

    Motile cilia are hair-like cell extensions that beat periodically to generate fluid flow along various epithelial tissues within the body. In dense multiciliated carpets, cilia were shown to exhibit a remarkable coordination of their beat in the form of traveling metachronal waves, a phenomenon which supposedly enhances fluid transport. Yet, how cilia coordinate their regular beat in multiciliated epithelia to move fluids remains insufficiently understood, particularly due to lack of rigorous quantification. We combine experiments, novel analysis tools, and theory to address this knowledge gap. To investigate collective dynamics of cilia, we studied zebrafish multiciliated epithelia in the nose and the brain. We focused mainly on the zebrafish nose, due to its conserved properties with other ciliated tissues and its superior accessibility for non-invasive imaging. We revealed that cilia are synchronized only locally and that the size of local synchronization domains increases with the viscosity of the surrounding medium. Even though synchronization is local only, we observed global patterns of traveling metachronal waves across the zebrafish multiciliated epithelium. Intriguingly, these global wave direction patterns are conserved across individual fish, but different for left and right noses, unveiling a chiral asymmetry of metachronal coordination. To understand the implications of synchronization for fluid pumping, we used a computational model of a regular array of cilia. We found that local metachronal synchronization prevents steric collisions, i.e., cilia colliding with each other, and improves fluid pumping in dense cilia carpets, but hardly affects the direction of fluid flow. In conclusion, we show that local synchronization together with tissue-scale cilia alignment coincide and generate metachronal wave patterns in multiciliated epithelia, which enhance their physiological function of fluid pumping.

    View details for DOI 10.7554/eLife.77701

    View details for PubMedID 36700548

    View details for PubMedCentralID PMC9940908

  • AdipoQ-a simple, open-source software to quantify adipocyte morphology and function in tissues and in vitro. Molecular biology of the cell Sieckmann, K., Winnerling, N., Huebecker, M., Leyendecker, P., Juliana Silva Ribeiro, D., Gnad, T., Pfeifer, A., Wachten, D., Hansen, J. N. 2022; 33 (12): br22

    Abstract

    The different adipose tissues (ATs) can be distinguished according to their function. For example, white AT stores energy in form of lipids, whereas brown AT dissipates energy in the form of heat. These functional differences are represented in the respective adipocyte morphology; whereas white adipocytes contain large, unilocular lipid droplets, brown adipocytes contain smaller, multilocular lipid droplets. However, an automated, image analysis pipeline to comprehensively analyze adipocytes in vitro in cell culture as well as ex vivo in tissue sections is missing. We here present AdipoQ, an open-source software implemented as ImageJ plugins that allows us to analyze adipocytes in tissue sections and in vitro after histological and/or immunofluorescent labeling. AdipoQ is compatible with different imaging modalities and staining methods, allows batch processing of large datasets and simple post-hoc analysis, provides a broad band of parameters, and allows combining multiple fluorescent readouts. Therefore AdipoQ is of immediate use not only for basic research but also for clinical diagnosis.

    View details for DOI 10.1091/mbc.E21-11-0592

    View details for PubMedID 35947507

    View details for PubMedCentralID PMC9635306

  • Measurement of ciliary beating and fluid flow in the zebrafish adult telencephalon. STAR protocols Jeong, I., Hansen, J. N., Wachten, D., Jurisch-Yaksi, N. 2022; 3 (3): 101542

    Abstract

    Motile cilia are hair-like structures that move and propel fluid, playing important roles in the physiology of organs. Here, we present a protocol to visualize and measure ciliary beating and cerebrospinal fluid (CSF) flow in the telencephalon of an adult zebrafish brain explant. We describe the preparation of brain explants, the recording of ciliary beating and CSF flow, and data analysis using ImageJ and MATLAB. These imaging and analysis techniques can be directly translated to other ciliated systems. For complete details on the use and execution of this protocol, please refer to D'Gama et al. (2021).

    View details for DOI 10.1016/j.xpro.2022.101542

    View details for PubMedID 35842868

    View details for PubMedCentralID PMC9294268

  • MotiQ: an open-source toolbox to quantify the cell motility and morphology of microglia. Molecular biology of the cell Hansen, J. N., Brückner, M., Pietrowski, M. J., Jikeli, J. F., Plescher, M., Beckert, H., Schnaars, M., Fülle, L., Reitmeier, K., Langmann, T., Förster, I., Boche, D., Petzold, G. C., Halle, A. 2022; 33 (11): ar99

    Abstract

    Microglia are the primary resident innate immune cells of the CNS. They possess branched, motile cell processes that are important for their cellular functions. To study the pathways that control microglial morphology and motility under physiological and disease conditions, it is necessary to quantify microglial morphology and motility precisely and reliably. Several image analysis approaches are available for the quantification of microglial morphology and motility. However, they are either not automated, not freely accessible, and/or limited in the number of morphology and motility parameters that can be assessed. Thus, we have developed MotiQ, an open-source, freely accessible software for automated quantification of microglial motility and morphology. MotiQ allows quantification of a diverse set of cellular motility and morphology parameters, including the parameters that have become the gold standard in the microglia field. We demonstrate that MotiQ can be applied to in vivo, ex vivo, and in vitro data from confocal, epifluorescence, or two-photon microscopy, and we compare its results to other analysis approaches. We suggest MotiQ as a versatile and customizable tool to study microglia.

    View details for DOI 10.1091/mbc.E21-11-0585

    View details for PubMedID 35731557

    View details for PubMedCentralID PMC9582802

  • A cAMP signalosome in primary cilia drives gene expression and kidney cyst formation. EMBO reports Hansen, J. N., Kaiser, F., Leyendecker, P., Stüven, B., Krause, J. H., Derakhshandeh, F., Irfan, J., Sroka, T. J., Preval, K. M., Desai, P. B., Kraut, M., Theis, H., Drews, A. D., De-Domenico, E., Händler, K., Pazour, G. J., Henderson, D. J., Mick, D. U., Wachten, D. 2022; 23 (8): e54315

    Abstract

    The primary cilium constitutes an organelle that orchestrates signal transduction independently from the cell body. Dysregulation of this intricate molecular architecture leads to severe human diseases, commonly referred to as ciliopathies. However, the molecular underpinnings how ciliary signaling orchestrates a specific cellular output remain elusive. By combining spatially resolved optogenetics with RNA sequencing and imaging, we reveal a novel cAMP signalosome that is functionally distinct from the cytoplasm. We identify the genes and pathways targeted by the ciliary cAMP signalosome and shed light on the underlying mechanisms and downstream signaling. We reveal that chronic stimulation of the ciliary cAMP signalosome transforms kidney epithelia from tubules into cysts. Counteracting this chronic cAMP elevation in the cilium by small molecules targeting activation of phosphodiesterase-4 long isoforms inhibits cyst growth. Thereby, we identify a novel concept of how the primary cilium controls cellular functions and maintains tissue integrity in a specific and spatially distinct manner and reveal novel molecular components that might be involved in the development of one of the most common genetic diseases, polycystic kidney disease.

    View details for DOI 10.15252/embr.202154315

    View details for PubMedID 35695071

    View details for PubMedCentralID PMC9346484

  • Cannabinoid receptor 2 is necessary to induce toll-like receptor-mediated microglial activation. Glia Reusch, N., Ravichandran, K. A., Olabiyi, B. F., Komorowska-Müller, J. A., Hansen, J. N., Ulas, T., Beyer, M., Zimmer, A., Schmöle, A. C. 2022; 70 (1): 71-88

    Abstract

    The tight regulation of microglia activity is key for precise responses to potential threats, while uncontrolled and exacerbated microglial activity is neurotoxic. Microglial toll-like receptors (TLRs) are indispensable for sensing different types of assaults and triggering an innate immune response. Cannabinoid receptor 2 (CB2) signaling is a key pathway to control microglial homeostasis and activation, and its activation is connected to changes in microglial activity. We aimed to investigate how CB2 signaling impacts TLR-mediated microglial activation. Here, we demonstrate that deletion of CB2 causes a dampened transcriptional response to prototypic TLR ligands in microglia. Loss of CB2 results in distinct microglial gene expression profiles, morphology, and activation. We show that the CB2-mediated attenuation of TLR-induced microglial activation is mainly p38 MAPK-dependent. Taken together, we demonstrate that CB2 expression and signaling are necessary to fine-tune TLR-induced activation programs in microglia.

    View details for DOI 10.1002/glia.24089

    View details for PubMedID 34499767

  • Diversity and function of motile ciliated cell types within ependymal lineages of the zebrafish brain. Cell reports D'Gama, P. P., Qiu, T., Cosacak, M. I., Rayamajhi, D., Konac, A., Hansen, J. N., Ringers, C., Acuña-Hinrichsen, F., Hui, S. P., Olstad, E. W., Chong, Y. L., Lim, C. K., Gupta, A., Ng, C. P., Nilges, B. S., Kashikar, N. D., Wachten, D., Liebl, D., Kikuchi, K., Kizil, C., Yaksi, E., Roy, S., Jurisch-Yaksi, N. 2021; 37 (1): 109775

    Abstract

    Motile cilia defects impair cerebrospinal fluid (CSF) flow and can cause brain and spine disorders. The development of ciliated cells, their impact on CSF flow, and their function in brain and axial morphogenesis are not fully understood. We have characterized motile ciliated cells within the zebrafish brain ventricles. We show that the ventricles undergo restructuring through development, involving a transition from mono- to multiciliated cells (MCCs) driven by gmnc. MCCs co-exist with monociliated cells and generate directional flow patterns. These ciliated cells have different developmental origins and are genetically heterogenous with respect to expression of the Foxj1 family of ciliary master regulators. Finally, we show that cilia loss from the tela choroida and choroid plexus or global perturbation of multiciliation does not affect overall brain or spine morphogenesis but results in enlarged ventricles. Our findings establish that motile ciliated cells are generated by complementary and sequential transcriptional programs to support ventricular development.

    View details for DOI 10.1016/j.celrep.2021.109775

    View details for PubMedID 34610312

    View details for PubMedCentralID PMC8524669

  • Soluble adenylyl cyclase inhibition prevents human sperm functions essential for fertilization. Molecular human reproduction Balbach, M., Ghanem, L., Rossetti, T., Kaur, N., Ritagliati, C., Ferreira, J., Krapf, D., Puga Molina, L. C., Santi, C. M., Hansen, J. N., Wachten, D., Fushimi, M., Meinke, P. T., Buck, J., Levin, L. R. 2021; 27 (9)

    Abstract

    Soluble adenylyl cyclase (sAC: ADCY10) has been genetically confirmed to be essential for male fertility in mice and humans. In mice, ex vivo studies of dormant, caudal epididymal sperm demonstrated that sAC is required for initiating capacitation and activating motility. We now use an improved sAC inhibitor, TDI-10229, for a comprehensive analysis of sAC function in mouse and human sperm. In contrast to caudal epididymal mouse sperm, human sperm are collected post-ejaculation, after sAC activity has already been stimulated. In addition to preventing the capacitation-induced stimulation of sAC and protein kinase A activities, tyrosine phosphorylation, alkalinization, beat frequency and acrosome reaction in dormant mouse sperm, sAC inhibitors interrupt each of these capacitation-induced changes in ejaculated human sperm. Furthermore, we show for the first time that sAC is required during acrosomal exocytosis in mouse and human sperm. These data define sAC inhibitors as candidates for non-hormonal, on-demand contraceptives suitable for delivery via intravaginal devices in women.

    View details for DOI 10.1093/molehr/gaab054

    View details for PubMedID 34463764

    View details for PubMedCentralID PMC8473925

  • Nucleo-cytoplasmic shuttling of splicing factor SRSF1 is required for development and cilia function. eLife Haward, F., Maslon, M. M., Yeyati, P. L., Bellora, N., Hansen, J. N., Aitken, S., Lawson, J., von Kriegsheim, A., Wachten, D., Mill, P., Adams, I. R., Caceres, J. F. 2021; 10

    Abstract

    Shuttling RNA-binding proteins coordinate nuclear and cytoplasmic steps of gene expression. The SR family proteins regulate RNA splicing in the nucleus and a subset of them, including SRSF1, shuttles between the nucleus and cytoplasm affecting post-splicing processes. However, the physiological significance of this remains unclear. Here, we used genome editing to knock-in a nuclear retention signal (NRS) in Srsf1 to create a mouse model harboring an SRSF1 protein that is retained exclusively in the nucleus. Srsf1NRS/NRS mutants displayed small body size, hydrocephalus, and immotile sperm, all traits associated with ciliary defects. We observed reduced translation of a subset of mRNAs and decreased abundance of proteins involved in multiciliogenesis, with disruption of ciliary ultrastructure and motility in cells and tissues derived from this mouse model. These results demonstrate that SRSF1 shuttling is used to reprogram gene expression networks in the context of high cellular demands, as observed here, during motile ciliogenesis.

    View details for DOI 10.7554/eLife.65104

    View details for PubMedID 34338635

    View details for PubMedCentralID PMC8352595

  • Multifocal imaging for precise, label-free tracking of fast biological processes in 3D. Nature communications Hansen, J. N., Gong, A., Wachten, D., Pascal, R., Turpin, A., Jikeli, J. F., Kaupp, U. B., Alvarez, L. 2021; 12 (1): 4574

    Abstract

    Many biological processes happen on a nano- to millimeter scale and within milliseconds. Established methods such as confocal microscopy are suitable for precise 3D recordings but lack the temporal or spatial resolution to resolve fast 3D processes and require labeled samples. Multifocal imaging (MFI) allows high-speed 3D imaging but is limited by the compromise between high spatial resolution and large field-of-view (FOV), and the requirement for bright fluorescent labels. Here, we provide an open-source 3D reconstruction algorithm for multi-focal images that allows using MFI for fast, precise, label-free tracking spherical and filamentous structures in a large FOV and across a high depth. We characterize fluid flow and flagellar beating of human and sea urchin sperm with a z-precision of 0.15 µm, in a volume of 240 × 260 × 21 µm, and at high speed (500 Hz). The sampling volume allowed to follow sperm trajectories while simultaneously recording their flagellar beat. Our MFI concept is cost-effective, can be easily implemented, and does not rely on object labeling, which renders it broadly applicable.

    View details for DOI 10.1038/s41467-021-24768-4

    View details for PubMedID 34321468

    View details for PubMedCentralID PMC8319204

  • Cannabinoid receptor 1 signalling modulates stress susceptibility and microglial responses to chronic social defeat stress. Translational psychiatry Beins, E. C., Beiert, T., Jenniches, I., Hansen, J. N., Leidmaa, E., Schrickel, J. W., Zimmer, A. 2021; 11 (1): 164

    Abstract

    Psychosocial stress is one of the main environmental factors contributing to the development of psychiatric disorders. In humans and rodents, chronic stress is associated with elevated inflammatory responses, indicated by increased numbers of circulating myeloid cells and activation of microglia, the brain-resident immune cells. The endocannabinoid system (ECS) regulates neuronal and endocrine stress responses via the cannabinoid receptor 1 (CB1). CB1-deficient mice (Cnr1-/-) are highly sensitive to stress, but if this involves altered inflammatory responses is not known. To test this, we exposed Cnr1+/+ and Cnr1-/- mice to chronic social defeat stress (CSDS). Cnr1-/- mice were extremely sensitive to a standard protocol of CSDS, indicated by an increased mortality rate. Therefore, a mild CSDS protocol was established, which still induced a behavioural phenotype in susceptible Cnr1-/- mice. These mice also showed altered glucocorticoid levels after mild CSDS, suggesting dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis. Mild CSDS induced weak myelopoiesis in the periphery, but no recruitment of myeloid cells to the brain. In contrast, mild CSDS altered microglial activation marker expression and morphology in Cnr1-/- mice. These microglial changes correlated with the severity of the behavioural phenotype. Furthermore, microglia of Cnr1-/- mice showed increased expression of Fkbp5, an important regulator of glucocorticoid signalling. Overall, the results confirm that CB1 signalling protects the organism from the physical and emotional harm of social stress and implicate endocannabinoid-mediated modulation of microglia in the development of stress-related pathologies.

    View details for DOI 10.1038/s41398-021-01283-0

    View details for PubMedID 33723234

    View details for PubMedCentralID PMC7961142

  • CiliaQ: a simple, open-source software for automated quantification of ciliary morphology and fluorescence in 2D, 3D, and 4D images. The European physical journal. E, Soft matter Hansen, J. N., Rassmann, S., Stüven, B., Jurisch-Yaksi, N., Wachten, D. 2021; 44 (2): 18

    Abstract

    Cilia are hair-like membrane protrusions that emanate from the surface of most vertebrate cells and are classified into motile and primary cilia. Motile cilia move fluid flow or propel cells, while also fulfill sensory functions. Primary cilia are immotile and act as a cellular antenna, translating environmental cues into cellular responses. Ciliary dysfunction leads to severe diseases, commonly termed ciliopathies. The molecular details underlying ciliopathies and ciliary function are, however, not well understood. Since cilia are small subcellular compartments, imaging-based approaches have been used to study them. However, tools to comprehensively analyze images are lacking. Automatic analysis approaches require commercial software and are limited to 2D analysis and only a few parameters. The widely used manual analysis approaches are time consuming, user-biased, and difficult to compare. Here, we present CiliaQ, a package of open-source, freely available, and easy-to-use ImageJ plugins. CiliaQ allows high-throughput analysis of 2D and 3D, static or time-lapse images from fluorescence microscopy of cilia in cell culture or tissues, and outputs a comprehensive list of parameters for ciliary morphology, length, bending, orientation, and fluorescence intensity, making it broadly applicable. We envision CiliaQ as a resource and platform for reproducible and comprehensive analysis of ciliary function in health and disease.

    View details for DOI 10.1140/epje/s10189-021-00031-y

    View details for PubMedID 33683488

    View details for PubMedCentralID PMC7940315

  • Tubulin glycylation controls axonemal dynein activity, flagellar beat, and male fertility. Science (New York, N.Y.) Gadadhar, S., Alvarez Viar, G., Hansen, J. N., Gong, A., Kostarev, A., Ialy-Radio, C., Leboucher, S., Whitfield, M., Ziyyat, A., Touré, A., Alvarez, L., Pigino, G., Janke, C. 2021; 371 (6525)

    Abstract

    Posttranslational modifications of the microtubule cytoskeleton have emerged as key regulators of cellular functions, and their perturbations have been linked to a growing number of human pathologies. Tubulin glycylation modifies microtubules specifically in cilia and flagella, but its functional and mechanistic roles remain unclear. In this study, we generated a mouse model entirely lacking tubulin glycylation. Male mice were subfertile owing to aberrant beat patterns of their sperm flagella, which impeded the straight swimming of sperm cells. Using cryo-electron tomography, we showed that lack of glycylation caused abnormal conformations of the dynein arms within sperm axonemes, providing the structural basis for the observed dysfunction. Our findings reveal the importance of microtubule glycylation for controlled flagellar beating, directional sperm swimming, and male fertility.

    View details for DOI 10.1126/science.abd4914

    View details for PubMedID 33414192

    View details for PubMedCentralID PMC7612590

  • Molecular Mechanism Underlying the Action of Zona-pellucida Glycoproteins on Mouse Sperm. Frontiers in cell and developmental biology Balbach, M., Hamzeh, H., Jikeli, J. F., Brenker, C., Schiffer, C., Hansen, J. N., Neugebauer, P., Trötschel, C., Jovine, L., Han, L., Florman, H. M., Kaupp, U. B., Strünker, T., Wachten, D. 2020; 8: 572735

    Abstract

    Mammalian oocytes are enveloped by the zona pellucida (ZP), an extracellular matrix of glycoproteins. In sperm, stimulation with ZP proteins evokes a rapid Ca2+ influx via the sperm-specific, pH-sensitive Ca2+ channel CatSper. However, the physiological role and molecular mechanisms underlying ZP-dependent activation of CatSper are unknown. Here, we delineate the sequence of ZP-signaling events in mouse sperm. We show that ZP proteins evoke a rapid intracellular pH i increase that rests predominantly on Na+/H+ exchange by NHA1 and requires cAMP synthesis by the soluble adenylyl cyclase sAC as well as a sufficiently negative membrane potential set by the spem-specific K+ channel Slo3. The alkaline-activated CatSper channel translates the ZP-induced pH i increase into a Ca2+ response. Our findings reveal the molecular components underlying ZP action on mouse sperm, opening up new avenues for understanding the basic principles of sperm function and, thereby, mammalian fertilization.

    View details for DOI 10.3389/fcell.2020.572735

    View details for PubMedID 32984353

    View details for PubMedCentralID PMC7487327

  • Cfap97d1 is important for flagellar axoneme maintenance and male mouse fertility. PLoS genetics Oura, S., Kazi, S., Savolainen, A., Nozawa, K., Castañeda, J., Yu, Z., Miyata, H., Matzuk, R. M., Hansen, J. N., Wachten, D., Matzuk, M. M., Prunskaite-Hyyryläinen, R. 2020; 16 (8): e1008954

    Abstract

    The flagellum is essential for sperm motility and fertilization in vivo. The axoneme is the main component of the flagella, extending through its entire length. An axoneme is comprised of two central microtubules surrounded by nine doublets, the nexin-dynein regulatory complex, radial spokes, and dynein arms. Failure to properly assemble components of the axoneme in a sperm flagellum, leads to fertility alterations. To understand this process in detail, we have defined the function of an uncharacterized gene, Cfap97 domain containing 1 (Cfap97d1). This gene is evolutionarily conserved in mammals and multiple other species, including Chlamydomonas. We have used two independently generated Cfap97d1 knockout mouse models to study the gene function in vivo. Cfap97d1 is exclusively expressed in testes starting from post-natal day 20 and continuing throughout adulthood. Deletion of the Cfap97d1 gene in both mouse models leads to sperm motility defects (asthenozoospermia) and male subfertility. In vitro fertilization (IVF) of cumulus-intact oocytes with Cfap97d1 deficient sperm yielded few embryos whereas IVF with zona pellucida-free oocytes resulted in embryo numbers comparable to that of the control. Knockout spermatozoa showed abnormal motility characterized by frequent stalling in the anti-hook position. Uniquely, Cfap97d1 loss caused a phenotype associated with axonemal doublet heterogeneity linked with frequent loss of the fourth doublet in the sperm stored in the epididymis. This study demonstrates that Cfap97d1 is required for sperm flagellum ultra-structure maintenance, thereby playing a critical role in sperm function and male fertility in mice.

    View details for DOI 10.1371/journal.pgen.1008954

    View details for PubMedID 32785227

    View details for PubMedCentralID PMC7444823

  • Nanobody-directed targeting of optogenetic tools to study signaling in the primary cilium. eLife Hansen, J. N., Kaiser, F., Klausen, C., Stüven, B., Chong, R., Bönigk, W., Mick, D. U., Möglich, A., Jurisch-Yaksi, N., Schmidt, F. I., Wachten, D. 2020; 9

    Abstract

    Compartmentalization of cellular signaling forms the molecular basis of cellular behavior. The primary cilium constitutes a subcellular compartment that orchestrates signal transduction independent from the cell body. Ciliary dysfunction causes severe diseases, termed ciliopathies. Analyzing ciliary signaling has been challenging due to the lack of tools to investigate ciliary signaling. Here, we describe a nanobody-based targeting approach for optogenetic tools in mammalian cells and in vivo in zebrafish to specifically analyze ciliary signaling and function. Thereby, we overcome the loss of protein function observed after fusion to ciliary targeting sequences. We functionally localized modifiers of cAMP signaling, the photo-activated adenylyl cyclase bPAC and the light-activated phosphodiesterase LAPD, and the cAMP biosensor mlCNBD-FRET to the cilium. Using this approach, we studied the contribution of spatial cAMP signaling in controlling cilia length. Combining optogenetics with nanobody-based targeting will pave the way to the molecular understanding of ciliary function in health and disease.

    View details for DOI 10.7554/eLife.57907

    View details for PubMedID 32579112

    View details for PubMedCentralID PMC7338050

  • Reduced sialylation triggers homeostatic synapse and neuronal loss in middle-aged mice. Neurobiology of aging Klaus, C., Hansen, J. N., Ginolhac, A., Gérard, D., Gnanapragassam, V. S., Horstkorte, R., Rossdam, C., Buettner, F. F., Sauter, T., Sinkkonen, L., Neumann, H., Linnartz-Gerlach, B. 2020; 88: 91-107

    Abstract

    Sialic acid-binding Ig-like lectin (Siglec) receptors are linked to neurodegenerative processes, but the role of sialic acids in physiological aging is still not fully understood. We investigated the impact of reduced sialylation in the brain of mice heterozygous for the enzyme glucosamine-2-epimerase/N-acetylmannosamine kinase (GNE+/-) that is essential for sialic acid biosynthesis. We demonstrate that GNE+/- mice have hyposialylation in different brain regions, less synapses in the hippocampus and reduced microglial arborization already at 6 months followed by increased loss of neurons at 12 months. A transcriptomic analysis revealed no pro-inflammatory changes indicating an innate homeostatic immune process leading to the removal of synapses and neurons in GNE+/- mice during aging. Crossbreeding with complement C3-deficient mice rescued the earlier onset of neuronal and synaptic loss as well as the changes in microglial arborization. Thus, sialic acids of the glycocalyx contribute to brain homeostasis and act as a recognition system for the innate immune system in the brain.

    View details for DOI 10.1016/j.neurobiolaging.2020.01.008

    View details for PubMedID 32087947

  • Elucidating cyclic AMP signaling in subcellular domains with optogenetic tools and fluorescent biosensors. Biochemical Society transactions Klausen, C., Kaiser, F., Stüven, B., Hansen, J. N., Wachten, D. 2019; 47 (6): 1733-1747

    Abstract

    The second messenger 3',5'-cyclic nucleoside adenosine monophosphate (cAMP) plays a key role in signal transduction across prokaryotes and eukaryotes. Cyclic AMP signaling is compartmentalized into microdomains to fulfil specific functions. To define the function of cAMP within these microdomains, signaling needs to be analyzed with spatio-temporal precision. To this end, optogenetic approaches and genetically encoded fluorescent biosensors are particularly well suited. Synthesis and hydrolysis of cAMP can be directly manipulated by photoactivated adenylyl cyclases (PACs) and light-regulated phosphodiesterases (PDEs), respectively. In addition, many biosensors have been designed to spatially and temporarily resolve cAMP dynamics in the cell. This review provides an overview about optogenetic tools and biosensors to shed light on the subcellular organization of cAMP signaling.

    View details for DOI 10.1042/BST20190246

    View details for PubMedID 31724693

  • Exacerbation of C1q dysregulation, synaptic loss and memory deficits in tau pathology linked to neuronal adenosine A2A receptor. Brain : a journal of neurology Carvalho, K., Faivre, E., Pietrowski, M. J., Marques, X., Gomez-Murcia, V., Deleau, A., Huin, V., Hansen, J. N., Kozlov, S., Danis, C., Temido-Ferreira, M., Coelho, J. E., Mériaux, C., Eddarkaoui, S., Gras, S. L., Dumoulin, M., Cellai, L., Landrieu, I., Chern, Y., Hamdane, M., Buée, L., Boutillier, A. L., Levi, S., Halle, A., Lopes, L. V., Blum, D. 2019; 142 (11): 3636-3654

    Abstract

    Accumulating data support the role of tau pathology in cognitive decline in ageing and Alzheimer's disease, but underlying mechanisms remain ill-defined. Interestingly, ageing and Alzheimer's disease have been associated with an abnormal upregulation of adenosine A2A receptor (A2AR), a fine tuner of synaptic plasticity. However, the link between A2AR signalling and tau pathology has remained largely unexplored. In the present study, we report for the first time a significant upregulation of A2AR in patients suffering from frontotemporal lobar degeneration with the MAPT P301L mutation. To model these alterations, we induced neuronal A2AR upregulation in a tauopathy mouse model (THY-Tau22) using a new conditional strain allowing forebrain overexpression of the receptor. We found that neuronal A2AR upregulation increases tau hyperphosphorylation, potentiating the onset of tau-induced memory deficits. This detrimental effect was linked to a singular microglial signature as revealed by RNA sequencing analysis. In particular, we found that A2AR overexpression in THY-Tau22 mice led to the hippocampal upregulation of C1q complement protein-also observed in patients with frontotemporal lobar degeneration-and correlated with the loss of glutamatergic synapses, likely underlying the observed memory deficits. These data reveal a key impact of overactive neuronal A2AR in the onset of synaptic loss in tauopathies, paving the way for new therapeutic approaches.

    View details for DOI 10.1093/brain/awz288

    View details for PubMedID 31599329

    View details for PubMedCentralID PMC6821333

  • Neural sphingosine 1-phosphate accumulation activates microglia and links impaired autophagy and inflammation. Glia Karunakaran, I., Alam, S., Jayagopi, S., Frohberger, S. J., Hansen, J. N., Kuehlwein, J., Hölbling, B. V., Schumak, B., Hübner, M. P., Gräler, M. H., Halle, A., van Echten-Deckert, G. 2019; 67 (10): 1859-1872

    Abstract

    Microglia mediated responses to neuronal damage in the form of neuroinflammation is a common thread propagating neuropathology. In this study, we investigated the microglial alterations occurring as a result of sphingosine 1-phosphate (S1P) accumulation in neural cells. We evidenced increased microglial activation in the brains of neural S1P-lyase (SGPL1) ablated mice (SGPL1fl/fl/Nes ) as shown by an activated and deramified morphology and increased activation markers on microglia. In addition, an increase of pro-inflammatory cytokines in sorted and primary cultured microglia generated from SGPL1 deficient mice was noticed. Further, we assessed autophagy, one of the major mechanisms in the brain that keeps inflammation in check. Indeed, microglial inflammation was accompanied by defective microglial autophagy in SGPL1 ablated mice. Rescuing autophagy by treatment with rapamycin was sufficient to decrease interleukin 6 (IL-6) but not tumor necrosis factor (TNF) secretion in cultured microglia. Rapamycin mediated decrease of IL-6 secretion suggests a particular mechanistic target of rapamycin (mTOR)-IL-6 link and appeared to be microglia specific. Using pharmacological inhibitors of the major receptors of S1P expressed in the microglia, we identified S1P receptor 2 (S1PR2) as the mediator of both impaired autophagy and proinflammatory effects. In line with these results, the addition of exogenous S1P to BV2 microglial cells showed similar effects as those observed in the genetic knock out of SGPL1 in the neural cells. In summary, we show a novel role of the S1P-S1PR2 axis in the microglia of mice with neural-targeted SGPL1 ablation and in BV2 microglial cell line exogenously treated with S1P.

    View details for DOI 10.1002/glia.23663

    View details for PubMedID 31231866

  • Revisiting and Redesigning Light-Activated Cyclic-Mononucleotide Phosphodiesterases. Journal of molecular biology Stabel, R., Stüven, B., Hansen, J. N., Körschen, H. G., Wachten, D., Möglich, A. 2019; 431 (17): 3029-3045

    Abstract

    As diffusible second messengers, cyclic nucleoside monophosphates (cNMPs) relay and amplify molecular signals in myriad cellular pathways. The triggering of downstream physiological responses often requires defined cNMP gradients in time and space, generated through the concerted action of nucleotidyl cyclases and phosphodiesterases (PDEs). In an approach denoted optogenetics, sensory photoreceptors serve as genetically encoded, light-responsive actuators to enable the noninvasive, reversible, and spatiotemporally precise control of manifold cellular processes, including cNMP metabolism. Although nature provides efficient photoactivated nucleotidyl cyclases, light-responsive PDEs are scarce. Through modular recombination of a bacteriophytochrome photosensor and the effector of human PDE2A, we previously generated the light-activated, cNMP-specific PDE LAPD. By pursuing parallel design strategies, we here report a suite of derivative PDEs with enhanced amplitude and reversibility of photoactivation. Opposite to LAPD, far-red light completely reverts prior activation by red light in several PDEs. These improved PDEs thus complement photoactivated nucleotidyl cyclases and extend the sensitivity of optogenetics to red and far-red light. More generally, our study informs future efforts directed at designing bacteriophytochrome photoreceptors.

    View details for DOI 10.1016/j.jmb.2019.07.011

    View details for PubMedID 31301407

  • Cyclic Nucleotide-Specific Optogenetics Highlights Compartmentalization of the Sperm Flagellum into cAMP Microdomains. Cells Raju, D. N., Hansen, J. N., Rassmann, S., Stüven, B., Jikeli, J. F., Strünker, T., Körschen, H. G., Möglich, A., Wachten, D. 2019; 8 (7)

    Abstract

    Inside the female genital tract, mammalian sperm undergo a maturation process called capacitation, which primes the sperm to navigate across the oviduct and fertilize the egg. Sperm capacitation and motility are controlled by 3',5'-cyclic adenosine monophosphate (cAMP). Here, we show that optogenetics, the control of cellular signaling by genetically encoded light-activated proteins, allows to manipulate cAMP dynamics in sperm flagella and, thereby, sperm capacitation and motility by light. To this end, we used sperm that express the light-activated phosphodiesterase LAPD or the photo-activated adenylate cyclase bPAC. The control of cAMP by LAPD or bPAC combined with pharmacological interventions provides spatiotemporal precision and allows to probe the physiological function of cAMP compartmentalization in mammalian sperm.

    View details for DOI 10.3390/cells8070648

    View details for PubMedID 31252584

    View details for PubMedCentralID PMC6679001

  • Species-specific differences in nonlysosomal glucosylceramidase GBA2 function underlie locomotor dysfunction arising from loss-of-function mutations. The Journal of biological chemistry Woeste, M. A., Stern, S., Raju, D. N., Grahn, E., Dittmann, D., Gutbrod, K., Dörmann, P., Hansen, J. N., Schonauer, S., Marx, C. E., Hamzeh, H., Körschen, H. G., Aerts, J. M., Bönigk, W., Endepols, H., Sandhoff, R., Geyer, M., Berger, T. K., Bradke, F., Wachten, D. 2019; 294 (11): 3853-3871

    Abstract

    The nonlysosomal glucosylceramidase β2 (GBA2) catalyzes the hydrolysis of glucosylceramide to glucose and ceramide. Mutations in the human GBA2 gene have been associated with hereditary spastic paraplegia (HSP), autosomal-recessive cerebellar ataxia (ARCA), and the Marinesco-Sjögren-like syndrome. However, the underlying molecular mechanisms are ill-defined. Here, using biochemistry, immunohistochemistry, structural modeling, and mouse genetics, we demonstrate that all but one of the spastic gait locus #46 (SPG46)-connected mutations cause a loss of GBA2 activity. We demonstrate that GBA2 proteins form oligomeric complexes and that protein-protein interactions are perturbed by some of these mutations. To study the pathogenesis of GBA2-related HSP and ARCA in vivo, we investigated GBA2-KO mice as a mammalian model system. However, these mice exhibited a high phenotypic variance and did not fully resemble the human phenotype, suggesting that mouse and human GBA2 differ in function. Whereas some GBA2-KO mice displayed a strong locomotor defect, others displayed only mild alterations of the gait pattern and no signs of cerebellar defects. On a cellular level, inhibition of GBA2 activity in isolated cerebellar neurons dramatically affected F-actin dynamics and reduced neurite outgrowth, which has been associated with the development of neurological disorders. Our results shed light on the molecular mechanism underlying the pathogenesis of GBA2-related HSP and ARCA and reveal species-specific differences in GBA2 function in vivo.

    View details for DOI 10.1074/jbc.RA118.006311

    View details for PubMedID 30662006

    View details for PubMedCentralID PMC6422076

  • Ciliary Beating Compartmentalizes Cerebrospinal Fluid Flow in the Brain and Regulates Ventricular Development. Current biology : CB Olstad, E. W., Ringers, C., Hansen, J. N., Wens, A., Brandt, C., Wachten, D., Yaksi, E., Jurisch-Yaksi, N. 2019; 29 (2): 229-241.e6

    Abstract

    Motile cilia are miniature, propeller-like extensions, emanating from many cell types across the body. Their coordinated beating generates a directional fluid flow, which is essential for various biological processes, from respiration to reproduction. In the nervous system, ependymal cells extend their motile cilia into the brain ventricles and contribute to cerebrospinal fluid (CSF) flow. Although motile cilia are not the only contributors to CSF flow, their functioning is crucial, as patients with motile cilia defects develop clinical features, like hydrocephalus and scoliosis. CSF flow was suggested to primarily deliver nutrients and remove waste, but recent studies emphasized its role in brain development and function. Nevertheless, it remains poorly understood how ciliary beating generates and organizes CSF flow to fulfill these roles. Here, we study motile cilia and CSF flow in the brain ventricles of larval zebrafish. We identified that different populations of motile ciliated cells are spatially organized and generate a directional CSF flow powered by ciliary beating. Our investigations revealed that CSF flow is confined within individual ventricular cavities, with little exchange of fluid between ventricles, despite a pulsatile CSF displacement caused by the heartbeat. Interestingly, our results showed that the ventricular boundaries supporting this compartmentalized CSF flow are abolished during bodily movement, highlighting that multiple physiological processes regulate the hydrodynamics of CSF flow. Finally, we showed that perturbing cilia reduces hydrodynamic coupling between the brain ventricles and disrupts ventricular development. We propose that motile-cilia-generated flow is crucial in regulating the distribution of CSF within and across brain ventricles.

    View details for DOI 10.1016/j.cub.2018.11.059

    View details for PubMedID 30612902

    View details for PubMedCentralID PMC6345627

  • SpermQ⁻A Simple Analysis Software to Comprehensively Study Flagellar Beating and Sperm Steering. Cells Hansen, J. N., Rassmann, S., Jikeli, J. F., Wachten, D. 2018; 8 (1)

    Abstract

    Motile cilia, also called flagella, are found across a broad range of species; some cilia propel prokaryotes and eukaryotic cells like sperm, while cilia on epithelial surfaces create complex fluid patterns e.g., in the brain or lung. For sperm, the picture has emerged that the flagellum is not only a motor but also a sensor that detects stimuli from the environment, computing the beat pattern according to the sensory input. Thereby, the flagellum navigates sperm through the complex environment in the female genital tract. However, we know very little about how environmental signals change the flagellar beat and, thereby, the swimming behavior of sperm. It has been proposed that distinct signaling domains in the flagellum control the flagellar beat. However, a detailed analysis has been mainly hampered by the fact that current comprehensive analysis approaches rely on complex microscopy and analysis systems. Thus, knowledge on sperm signaling regulating the flagellar beat is based on custom quantification approaches that are limited to only a few aspects of the beat pattern, do not resolve the kinetics of the entire flagellum, rely on manual, qualitative descriptions, and are only a little comparable among each other. Here, we present SpermQ, a ready-to-use and comprehensive analysis software to quantify sperm motility. SpermQ provides a detailed quantification of the flagellar beat based on common time-lapse images acquired by dark-field or epi-fluorescence microscopy, making SpermQ widely applicable. We envision SpermQ becoming a standard tool in flagellar and motile cilia research that allows to readily link studies on individual signaling components in sperm and distinct flagellar beat patterns.

    View details for DOI 10.3390/cells8010010

    View details for PubMedID 30587820

    View details for PubMedCentralID PMC6357160

  • CCL17 exerts a neuroimmune modulatory function and is expressed in hippocampal neurons. Glia Fülle, L., Offermann, N., Hansen, J. N., Breithausen, B., Erazo, A. B., Schanz, O., Radau, L., Gondorf, F., Knöpper, K., Alferink, J., Abdullah, Z., Neumann, H., Weighardt, H., Henneberger, C., Halle, A., Förster, I. 2018; 66 (10): 2246-2261

    Abstract

    Chemokines are important signaling molecules in the immune and nervous system. Using a fluorescence reporter mouse model, we demonstrate that the chemokine CCL17, a ligand of the chemokine receptor CCR4, is produced in the murine brain, particularly in a subset of hippocampal CA1 neurons. We found that basal expression of Ccl17 in hippocampal neurons was strongly enhanced by peripheral challenge with lipopolysaccharide (LPS). LPS-mediated induction of Ccl17 in the hippocampus was dependent on local tumor necrosis factor (TNF) signaling, whereas upregulation of Ccl22 required granulocyte-macrophage colony-stimulating factor (GM-CSF). CCL17 deficiency resulted in a diminished microglia density under homeostatic and inflammatory conditions. Further, microglia from naïve Ccl17-deficient mice possessed a reduced cellular volume and a more polarized process tree as assessed by computer-assisted imaging analysis. Regarding the overall branching, cell surface area, and total tree length, the morphology of microglia from naïve Ccl17-deficient mice resembled that of microglia from wild-type mice after LPS stimulation. In line, electrophysiological recordings indicated that CCL17 downmodulates basal synaptic transmission at CA3-CA1 Schaffer collaterals in acute slices from naïve but not LPS-treated animals. Taken together, our data identify CCL17 as a homeostatic and inducible neuromodulatory chemokine affecting the presence and morphology of microglia and synaptic transmission in the hippocampus.

    View details for DOI 10.1002/glia.23507

    View details for PubMedID 30277599

  • Plaque-dependent morphological and electrophysiological heterogeneity of microglia in an Alzheimer's disease mouse model. Glia Plescher, M., Seifert, G., Hansen, J. N., Bedner, P., Steinhäuser, C., Halle, A. 2018; 66 (7): 1464-1480

    Abstract

    Microglia, the central nervous system resident innate immune cells, cluster around Aβ plaques in Alzheimer's disease (AD). The activation phenotype of these plaque-associated microglial cells, and their differences to microglia distant to Aβ plaques, are incompletely understood. We used novel three-dimensional cell analysis software to comprehensively analyze the morphological properties of microglia in the TgCRND8 mouse model of AD in spatial relation to Aβ plaques. We found strong morphological changes exclusively in plaque-associated microglia, whereas plaque-distant microglia showed only minor changes. In addition, patch-clamp recordings of microglia in acute cerebral slices of TgCRND8 mice revealed increased K+ currents in plaque-associated but not plaque-distant microglia. Within the subgroup of plaque-associated microglia, two different current profiles were detected. One subset of cells displayed only increased inward currents, while a second subset showed both increased inward and outward currents, implicating that the plaque microenvironment differentially impacts microglial ion channel expression. Using pharmacological channel blockers, multiplex single-cell PCR analysis and RNA fluorescence in situ hybridization, we identified Kir and Kv channel types contributing to the in- and outward K+ conductance in plaque-associated microglia. In summary, we have identified a previously unrecognized level of morphological and electrophysiological heterogeneity of microglia in relation to amyloid plaques, suggesting that microglia may display multiple activation states in AD.

    View details for DOI 10.1002/glia.23318

    View details for PubMedID 29493017

  • Shedding light on the role of cAMP in mammalian sperm physiology. Molecular and cellular endocrinology Balbach, M., Beckert, V., Hansen, J. N., Wachten, D. 2018; 468: 111-120

    Abstract

    Mammalian fertilization relies on sperm finding the egg and penetrating the egg vestments. All steps in a sperm's lifetime crucially rely on changes in the second messenger cAMP (cyclic adenosine monophosphate). In recent years, it has become clear that signal transduction in sperm is not a continuum, but rather organized in subcellular domains, e.g. the sperm head and the sperm flagellum, with the latter being further separated into the midpiece, principal piece, and endpiece. To understand the underlying signaling pathways controlling sperm function in more detail, experimental approaches are needed that allow to study sperm signaling with spatial and temporal precision. Here, we will give a comprehensive overview on cAMP signaling in mammalian sperm, describing the molecular players involved in these pathways and the sperm functions that are controlled by cAMP. Furthermore, we will highlight recent advances in analyzing and manipulating sperm signaling with spatio-temporal precision using light.

    View details for DOI 10.1016/j.mce.2017.11.008

    View details for PubMedID 29146556

  • Cannabinoid Receptor 2-Deficiency Ameliorates Disease Symptoms in a Mouse Model with Alzheimer's Disease-Like Pathology. Journal of Alzheimer's disease : JAD Schmöle, A. C., Lundt, R., Toporowski, G., Hansen, J. N., Beins, E., Halle, A., Zimmer, A. 2018; 64 (2): 379-392

    Abstract

    It is widely accepted that the endocannabinoid system (ECS) is a modulator of neuroinflammation associated with neurodegenerative disorders, including Alzheimer's disease (AD). Thus, expression of the cannabinoid receptor 2 (CB2) is induced in plaque-associated microglia and astrocytes in brain tissues from AD patients and in genetic mouse models expressing pathogenic variants of the amyloid precursor protein (APP). However, the exact mechanism of CB2 signaling in this mouse model remains elusive, because the genetic deletion of CB2 and the pharmacological activation of CB2 both reduced neuroinflammation. Here, we demonstrate that CB2 deletion also improved cognitive and learning deficits in APP/PS1*CB2-/- mice. This was accompanied by reduced neuronal loss and decreased plaque levels and coincided with increased expression of Aβ degrading enzymes. Interestingly, plaque-associated microglia in APP/PS1*CB2-/- mice showed a less activated morphology, while plaques were smaller and more condensed than in APP/PS1 mice. Taken together, these results indicate a beneficial effect of CB2-deficiency in APP transgenic mice. CB2 appears to be part of a protective system that may be detrimental when engaged continuously.

    View details for DOI 10.3233/JAD-180230

    View details for PubMedID 29865078

  • P2Y1 receptor blockade normalizes network dysfunction and cognition in an Alzheimer's disease model. The Journal of experimental medicine Reichenbach, N., Delekate, A., Breithausen, B., Keppler, K., Poll, S., Schulte, T., Peter, J., Plescher, M., Hansen, J. N., Blank, N., Keller, A., Fuhrmann, M., Henneberger, C., Halle, A., Petzold, G. C. 2018; 215 (6): 1649-1663

    Abstract

    Astrocytic hyperactivity is an important contributor to neuronal-glial network dysfunction in Alzheimer's disease (AD). We have previously shown that astrocyte hyperactivity is mediated by signaling through the P2Y1 purinoreceptor (P2Y1R) pathway. Using the APPPS1 mouse model of AD, we here find that chronic intracerebroventricular infusion of P2Y1R inhibitors normalizes astroglial and neuronal network dysfunction, as measured by in vivo two-photon microscopy, augments structural synaptic integrity, and preserves hippocampal long-term potentiation. These effects occur independently from β-amyloid metabolism or plaque burden but are associated with a higher morphological complexity of periplaque reactive astrocytes, as well as reduced dystrophic neurite burden and greater plaque compaction. Importantly, APPPS1 mice chronically treated with P2Y1R antagonists, as well as APPPS1 mice carrying an astrocyte-specific genetic deletion (Ip3r2-/-) of signaling pathways downstream of P2Y1R activation, are protected from the decline of spatial learning and memory. In summary, our study establishes the restoration of network homoeostasis by P2Y1R inhibition as a novel treatment target in AD.

    View details for DOI 10.1084/jem.20171487

    View details for PubMedID 29724785

    View details for PubMedCentralID PMC5987918