Center for Molecular Medicine Cologne

Wirth, Brunhilde | Piano, Valentina - C 18

Vesicular trafficking and axonal local translation in health and motor neuron disease

Introduction

Motoneurons (MNs) are highly polarized nondividing cells that innervate muscles via the neuromuscular junctions, the largest synapses in our body.  MNs must be maintained throughout life otherwise neuropathies and muscle atrophy will develop. During development, external cues - such as netrins - direct the axons to their ultimate destination by interacting with specific attractive or repulsive neuronal receptors. To accomplish this unique feature, axons and dendrites rely on highly sophisticated cellular processes - such as efficient local protein translation - that enable a prompt response to internal signals and external cues. In MNs, local protein translation requires a proper transport of mRNAs and cargos, a coordinated autophagy and protein degradation system, and functional mitochondria to provide enough energy locally for all these processes.

In this project we will make use of mouse and cellular models of Spinal Muscular Atrophy (SMA) -the most common recessively inherited MN disorder in humans - to understand why reduced levels of the house-keeping SMN protein primarily led to dysfunction of MNs. In addition, we will take advantage of our knowledge about three SMA protective modifiers (PLS3, NCALD and CHP1), which can counteract SMA development in humans and various SMA species by restoring impaired endocytosis (Ref 1-14). Nevertheless, how endocytosis is affected in SMA MNs and how it ultimately affects MN function, particularly axonal local translation, mitochondrial bioenergetics and neurotransmission, is still poorly understood and it will be the main objective of this project.

In particular, we will focus on these 3 specific questions:

  • (WP1) how low SMN levels affect and how the protective modifiers can rescue axonal local translation and vesicle trafficking in MNs;
  • (WP2) whether previously identified novel interactors of PLS3 influence receptor-ribosome mediated axonal translation and selective MN vulnerability;
  • (WP3) which key biochemical processes are dysregulated in specific sub-compartments of MNs by reduced SMN levels, and can be counteracted by the SMA modifiers
Figure 1

Clinical Relevance

Axonal vesicle trafficking and local translation are key regulatory mechanisms affected in a plethora of MN disorders, including SMA. In SMA, these processes are understudied, especially in the axons and pre-synapses. Recently, three FDA- and EMA-approved therapies, aimed at increasing the levels of functional SMN protein, showed impressive improvement in SMA patients. However, the current therapies are not efficient for severely affected SMA patients, who require very high levels of SMN protein postnatal, and for adult SMA patients, who require only low levels of SMN protein. The identification of new molecular mechanisms that can support MN function is needed to develop new therapies to reduce MN alterations and counteract their loss in SMA patients. Here, we aim to investigate these mechanisms and contribute to the development of SMN-independent therapies for SMA and MN disorders in general.

Approach

In this project we will use the following cutting-edge approaches (Figure 1) that will enable us to elucidate outstanding questions in the field of vesicle-trafficking, axonal translation and selective vulnerability in healthy and SMA motor neurons.

  • WP1: we will use MNs differentiated from human CRISPR-Cas9 modified iPSCs (Figure 2) and primary murine MNs from SMA and SMA-PLS3 overexpression (OE) embryos as model systems. To study endosomal local translation in proximity to mitochondria and endosomal/lysosomal vesicle distribution/trafficking in health and disease, we will uselive-cell imaging, immunofluorescence staining and high-resolution microscopy.
  • WP2: Our PLS3-and CHP1 Co-IP/MS proteome analysis from CRISPR-Cas9-mediated KO of Pls3 and Chp1 versus WT PC12 neuronal-like cells unraveled new possible modifiers. We will assess their role in axonal local translation in SMA, and how their interaction with PLS3 affects local translation, signaling and selective MN vulnerability.
  • WP3: To identify impaired components in the relevant sub-compartments of SMA MNs, we will apply a recently developed approach to separate soma from axon in combination with proteome, transcriptome and Ribo-Seq of WT and SMA-or SMA-PLS3 OE human and murine MN.
Figure 2
    • Strathmann E, et al (2023) Am J Hum Genet, in press
    • Zilio E, et al (2022) Int J Mol Sci, 23(18):10878
    • Wirth B (2021) Trends Neurosci 44(4):306-322;
    • Wolff L, et al (2021) Cell Mol Life Sci78(13):5275-5301
    • Buettner JM,et al (2021) iScience 24(11):103376.;
    • Wirth B, et al (2020) Annu Rev Genomics Hum Genet 21:231-261;
    • Thelen MP, et al (2020) Acta Neuropathol Commun 8(1):223;
    • Torres-Benito, et al (2019) Am J Hum Genet105(1):221-230
    • Janzen E, et al (2018)Brain141(8):2343-2361;
    • Riessland M,et al (2017) Am J Hum Genet100(2):297-315;
    • Hosseinibarkooie S, et al (2016) Am J Hum Genet 99(3):647-665;
    • Heesen L,et al. (2016) Cell Mol Life Sci73(10):2089-2104.
    • Ackermann B,et al. (2013) Hum Mol Genet 22(7):1328-1347.;
    • Oprea GE,et al. (2008) Science 320(5875):524-527;

    Overview of publications generated during the current funding period (1/2023-12/2025) with CMMC affiliation

    For a complete list of B. Wirth's publications, please visit - pubmed_Wirth
    For a complete list of V. Piano's publications, please visit - pubmed_Piano

    • Ermanoska B, Asselbergh B, Morant L, Petrovic-Erfurth ML, Hosseinibarkooie S, Leitao-Goncalves R, Almeida-Souza L, Bervoets S, Sun L, Lee L, Atkinson D, Khanghahi A, Tournev I, Callaerts P, Verstreken P, Yang XL, Wirth B, Rodal AA, Timmerman V, Goode BL, Godenschwege TA, and Jordanova A (2023). Tyrosyl-tRNA synthetase has a noncanonical function in actin bundling. Nat Commun 14, 999. doi:10.1038/s41467-023-35908-3.
       
    • Fortuna TR, Kour S, Chimata AV, Muinos-Buhl A, Anderson EN, Nelson Iv CH, Ward C, Chauhan O, O'Brien C, Rajasundaram D, Rajan DS, Wirth B, Singh A, and Pandey UB (2023). SMN regulates GEMIN5 expression and acts as a modifier of GEMIN5-mediated neurodegeneration. Acta Neuropathol 146, 477-498. doi:10.1007/s00401-023-02607-8.
       
    • Kruse T, Shamai S, Leflerova D, Wirth B, Heller R, Schloss N, Lehmann HC, Brakemeier S, Hagenacker T, Braumann B, and Wunderlich G (2023). Objective measurement of oral function in adults with spinal muscular atrophy. Orphanet J Rare Dis 18, 103. doi:10.1186/s13023-023-02688-4.
       
    • Muinos-Buhl A, Rombo R, Ling KK, Zilio E, Rigo F, Bennett CF, and Wirth B (2023). Long-Term SMN- and Ncald-ASO Combinatorial Therapy in SMA Mice and NCALD-ASO Treatment in hiPSC-Derived Motor Neurons Show Protective Effects. Int J Mol Sci 24. doi:10.3390/ijms24044198.
       
    • Pavinato L, Delle Vedove A, Carli D, Ferrero M, Carestiato S, Howe JL, Agolini E, Coviello DA, van de Laar I, Au PYB, Di Gregorio E, Fabbiani A, Croci S, Mencarelli MA, Bruno LP, Renieri A, Veltra D, Sofocleous C, Faivre L, Mazel B, Safraou H, Denomme-Pichon AS, van Slegtenhorst MA, Giesbertz N, van Jaarsveld RH, Childers A, Rogers RC, Novelli A, De Rubeis S, Buxbaum JD, Scherer SW, Ferrero GB, Wirth B, and Brusco A (2023). CAPRIN1 haploinsufficiency causes a neurodevelopmental disorder with language impairment, ADHD and ASD. Brain 146, 534-548. doi:10.1093/brain/awac278.
       
    • Strathmann EA, Holker I, Tschernoster N, Hosseinibarkooie S, Come J, Martinat C, Altmuller J, and Wirth B (2023). Epigenetic regulation of plastin 3 expression by the macrosatellite DXZ4 and the transcriptional regulator CHD4. Am J Hum Genet 110, 442-459. doi:10.1016/j.ajhg.2023.02.004.
       
    • Winter NC, Karakaya M, Mosen P, Brusius I, Anlar B, Haliloglu G, Winter D, and Wirth B (2023). Proteomic Investigation of Differential Interactomes of Glypican 1 and a Putative Disease-Modifying Variant of Ataxia. Journal of Proteome Research 22, 3081-3095. doi:10.1021/acs.jproteome.3c00402.
    Prof. Dr. Brunhilde Wirth CMMC Cologne
    Prof. Dr. Brunhilde Wirth

    Institute of Human Genetics - CMMC Research Building

    CMMC - PI - C 18

    Executive Board Member

    +49 221 478 86464

    +49 221 478 86465

    Institute of Human Genetics - CMMC Research Building

    Kerpener Str. 34

    50931 Cologne

    https://humangenetik.uk-koeln.de/forschung/ag-neuromuskulaere-und-skelettale-erkrankungen/

    CMMC Profile Page

    Curriculum Vitae (CV)

    Publications on PubMed

    Dr. Valentina Piano CMMC Cologne
    Dr. Valentina Piano

    Institute of Human Genetics - CMMC Research Building

    CMMC - Co-PI - C 18 I CAP 32
    JRG Leader - Institute for Human Genetics Cologne

    +49 221 478 89523

    Institute of Human Genetics - CMMC Research Building

    Robert-Koch-Str. 21

    50931 Cologne

    https://pianolab.wixsite.com/pianolab

    CMMC Profile Page

    Curriculum Vitae (CV)

    Publications on PubMed