Eva - Hedlund

Stockholm University | SU · Department of Biochemistry and Biophysics

Mutations in the RNA/DNA-binding proteins FUS and TDP-43 cause amyotrophic lateral sclerosis (ALS) with distinct neuropathological features. It is currently unclear how these gene mutations lead to selective motor neuron death and if there are common mechanisms across disease causations. Using single cell RNA sequencing of neurons derived from isog...

BACKGROUND Major depressive disorder (MDD) is a serious disease and a burden to patients, families and society. Rodent experiments and human studies suggest that several neuropeptide systems, including substance P(SP)/tachykinin, neuropeptide Y(NPY) and their G protein-coupled receptors are involved in mood regulation. METHODS We assessed the tran...

Background Spinal muscular atrophy (SMA) is a form of motor neuron disease affecting primarily children characterised by the loss of lower motor neurons (MNs). Breakdown of the neuromuscular junctions (NMJs) is an early pathological event in SMA. However, not all motor neurons are equally vulnerable, with some populations being lost early in the di...

Endurance exercise promotes skeletal muscle vascularization, oxidative metabolism, fiber-type switching, and neuromuscular junction integrity. Importantly, the metabolic and contractile properties of the muscle fiber must be coupled to the identity of the innervating motor neuron (MN). Here, we show that muscle-derived neurturin (NRTN) acts on musc...

Defining transcriptional profiles of substantia nigra pars compacta (SNc) and ventral tegmental area (VTA) dopamine neurons is critical to understanding their differential vulnerability in Parkinson’s Disease (PD). Here, we determine transcriptomes of human SNc and VTA dopamine neurons using LCM-seq on a large sample cohort. We apply a bootstrappin...

Endurance exercise promotes skeletal muscle vascularization, oxidative metabolism, fiber-type switching, and neuromuscular junction integrity. Importantly, the metabolic and contractile properties of the muscle fiber must be coupled to the identity of the innervating motor neuron (MN). Here, we show that muscle-derived neurturin (NRTN) acts on musc...

Apart from well-defined factors in neuronal cells¹, only a few reports consider that the variability of sporadic amyotrophic lateral sclerosis (ALS) progression can depend on less-defined contributions from glia2,3 and blood vessels⁴. In this study we use an expression-weighted cell-type enrichment method to infer cell activity in spinal cord sampl...

Mutations in the RNA-binding protein Fused in Sarcoma (FUS) cause early-onset amyotrophic lateral sclerosis (ALS). However, a detailed understanding of central RNA targets of FUS and their implications for disease remain elusive. Here, we use a unique blend of crosslinking and immunoprecipitation (CLIP) and NMR spectroscopy to identify and characte...

Somatic motor neurons are selectively vulnerable in spinal muscular atrophy (SMA), which is caused by a deficiency of the ubiquitously expressed survival of motor neuron protein. However, some motor neuron groups, including oculomotor and trochlear (ocular), which innervate eye muscles, are for unknown reasons spared. To reveal mechanisms of vulner...

Recent studies in neuron‐glial metabolic coupling have shown that, in the CNS, astrocytes and oligodendrocytes support neurons with energy‐rich lactate/pyruvate via monocarboxylate transporters (MCTs). The presence of such transporters in the PNS, in both Schwann cells and neurons, has prompted us to question if a similar interaction may be present...

Cranial irradiation (IR), an effective tool to treat malignant brain tumors, triggers a chronic pro-inflammatory microglial response, at least in the adult brain. Using single-cell and bulk RNA sequencing, combined with histology, we show that the microglial response in the juvenile mouse hippocampus is rapid but returns toward normal within 1 week...

In amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA), spinal and lower brainstem motor neurons degenerate, but some motor neuron subtypes are spared, including oculomotor neurons (OMNs). The mechanisms responsible for this selective degeneration are largely unknown, but the molecular signatures of resistant and vulnerable motor...

Defining transcriptional profiles of substantia nigra pars compacta (SNc) and ventral tegmental area (VTA) dopamine neurons in human is critical to understanding their differential vulnerability in Parkinson Disease. However, reported marker profiles for these neuron populations are derived predominantly from rodents, utilize small sample sizes and...

Neuronal processes have an RNA composition that is distinct from the cell body. Therefore, to fully understand neuronal biology in health and disease we need to study both somas, dendrites and axons. Here we describe a detailed protocol of a newly refined method, Axon-seq, for RNA sequencing of axons (and dendrites) grown in isolation using single...

Mutations in MORC2 lead to an axonal form of Charcot-Marie-Tooth (CMT) neuropathy type 2Z. To date, 31 families have been described with mutations in MORC2, indicating that this gene is frequently involved in axonal CMT cases. While the genetic data clearly establish the causative role of MORC2 in CMT2Z, the impact of its mutations on neuronal biol...

Key points: We have systematically investigated how defects in muscle contractile function contribute to weakness in ALS. Weakness in whole muscles from late stage SOD1G93A mice was explained by muscle atrophy as seen by reduced mass and maximal force. On the other hand, surviving single muscle fibres in late stage SOD1G93A have preserved intracel...

Oculomotor neurons, which regulate eye movement, are resilient to degeneration in the lethal motor neuron disease amyotrophic lateral sclerosis (ALS). It would be highly advantageous if motor neuron resilience could be modeled in vitro. Toward this goal, we generated a high proportion of oculomotor neurons from mouse embryonic stem cells through te...

Longitudinal bone growth in children is sustained by growth plates, narrow discs of cartilage that provide a continuous supply of chondrocytes for endochondral ossification ¹ . However, it remains unknown how this supply is maintained throughout childhood growth. Chondroprogenitors in the resting zone are thought to be gradually consumed as they su...

Spinal motor axons traverse large distances to innervate target muscles, thus requiring local control of cellular events for proper functioning. To interrogate axon-specific processes we developed Axon-seq, a refined method incorporating microfluidics, RNA sequencing (RNA-seq), and bioinformatic quality control. We show that the axonal transcriptom...

Table S1. Details the Antibodies Used in the Study, as well as the RNA-Seq Studies Used for Cross-Comparison in the Manuscript and Accompanies Data Presented in Figures 1, 2, 3, 4, and 5

contains the mouse motor axonal transcriptome, as defined by genes detected in three of five axonal samples, as well as the differentially expressed genes between mouse control axons and somas and accompanies Figures 1, 2, and 3.

contains the list of genes that are differentially expressed between mouse SOD1G93A somas and axons, with the associated fold change, p value, and RPKM levels in all samples. It also contains the differentially expressed genes between control and SOD1G93A axons, as well as the RPKM values of these 121 genes in the soma compartments of both lines.

contains the gene lists corresponding to the Venn diagrams in Figures S3A and S3B, showing comparisons of our mESC-derived motor axon transcriptome with primary motor axons (Briese et al., 2016) and primary DRG axons (Minis et al., 2014), as well as the associated full GO term analysis sheets.

contains the human motor axonal transcriptome, as defined by genes detected in three of six axonal samples, which accompanies Figure 4, as well as the full GO term analysis sheet of the overlapping genes between the mouse and the human motor axon transcriptome, accompanying the Venn diagram in Figure 4E.

Oculomotor neurons, which regulate eye movement, are resilient to degeneration in the lethal motor neuron disease amyotrophic lateral sclerosis (ALS). It would be highly advantageous if motor neuron resilience could be modeled in vitro. Towards this goal, we generated a high proportion of oculomotor neurons from mouse embryonic stem cells through t...

Somatic motor neurons are selectively vulnerable in spinal muscular atrophy (SMA), a lethal disease caused by a deficiency of the ubiquitously expressed survival of motor neuron (SMN) protein. However, some brainstem motor neuron groups, including oculomotor and trochlear (ocular), which innervate the muscles around the eyes, are for unknown reason...

Defining the transcriptional profiles of substantia nigra pars compacta and ventral tegmental area dopamine neurons may reveal mechanisms underlying their differential vulnerabilities in Parkinson disease. Existing studies display extensive variability due to small sample sizes, resulting in troublingly few reproducible subtype-specific markers. We...

Spinal motor axons traverse large distances to innervate target muscle, and thus require local control of cellular events for proper functioning of the distal axon. To interrogate axon-specific processes we developed Axon-seq, a refined method incorporating microfluidic devices and stringent bioinformatic quality controls. Axon-seq demonstrates imp...

The cytokine transforming growth factor-β (TGF-β) regulates the development and homeostasis of several tissue-resident macrophage populations, including microglia. TGF-β is not critical for microglia survival but is required for the maintenance of the microglia-specific homeostatic gene signature1,2. Under defined host conditions, circulating monoc...

CRISPR/Cas9-based genome editing offers the possibility to knock out almost any gene of interest in an affordable and simple manner. The most common strategy is the introduction of a frameshift into the open reading frame (ORF) of the target gene which truncates the coding sequence (CDS) and targets the corresponding transcript for degradation by n...

LCM-seq couples laser capture microdissection of cells from frozen tissues with polyA-based RNA sequencing and is applicable to single neurons. The method utilizes off-the-shelf reagents and direct lysis of the cells without RNA purification, making it a simple and relatively cheap method with high reproducibility and sensitivity compared to previo...

CRISPR/Cas9-based genome editing offers the possibility to knock out (KO) almost any gene of interest in an affordable and simple manner. The most common strategy is the introduction of a frameshift into the open reading frame (ORF) of the target gene which truncates the coding sequence (CDS) and targets the corresponding transcript for degradation...

HB9-GFP expressing motor neurons crossing central microchannel and innervating skeletal muscle myotubes. Myotubes displayed sustained and coordinated contractions under control conditions. Movie is displayed in real time over a 10 second period.

Phase contrast movie showing spontaneous and coordinated myotube contraction under control conditions. Video is displayed in real time over a 10 second period.

Phase contrast movie showing higher frequency myotube contraction following motor neurons stimulation with 50 mM KCl. Video is displayed in real time over a 10 second period.

Phase contrast movie showing KCl stimulated myotube contraction is inhibited following myotube treatment with tubocurarine. Video is displayed in real time over a 10 second period.

Objective: We examined whether skeletal muscle overexpression of PGC-1α1 or PGC-1α4 affected myokine secretion and neuromuscular junction (NMJ) formation. Methods: A microfluidic device was used to model endocrine signaling and NMJ formation between primary mouse myoblast-derived myotubes and embryonic stem cell-derived motor neurons. Difference...

Converting resident glia into functional and subtype-specific neurons in vivo by delivering reprogramming genes directly to the brain provides a step forward toward the possibility of treating brain injuries or diseases. To date, it has been possible to obtain GABAergic and glutamatergic neurons via in vivo conversion, but the precise phenotype of...

Converting resident glia into functional and subtype-specific neurons in vivo by delivering reprogramming genes directly to the brain provides a step forward toward the possibility of treating brain injuries or diseases. To date, it has been possible to obtain GABAergic and glutamatergic neurons via in vivo conversion, but the precise phenotype of...

Cells are the basic building blocks of organisms and each cell is unique. Single-cell RNA sequencing has emerged as an indispensable tool to dissect the cellular heterogeneity and decompose tissues into cell types and/or cell states, which offers enormous potential for de novo discovery. Single-cell transcriptomic atlases provide unprecedented reso...

In the fatal disease—amyotrophic lateral sclerosis (ALS)—upper (corticospinal) motor neurons (MNs) and lower somatic MNs, which innervate voluntary muscles, degenerate. Importantly, certain lower MN subgroups are relatively resistant to degeneration, even though pathogenic proteins are typically ubiquitously expressed. Ocular MNs (OMNs), including...

Pluripotency, differentiation and X chromosome inactivation (XCI) are key aspects of embryonic development. However, the underlying relationship and mechanisms among these processes remain unclear. Here we systematically dissected these features along developmental progression using mouse embryonic stem cells (mESCs) and single-cell RNA sequencing...

Laser capture microscopy (LCM) coupled with global transcriptome profiling could enable precise analyses of cell populations without the need for tissue dissociation, but has so far required relatively large numbers of cells. Here we report a robust and highly efficient strategy for LCM coupled with full-length mRNA-sequencing (LCM-seq) developed f...

Supplementary Figures 1-8 and Supplementary Tables 1-2

Differentially expressed genes between human spinal motor neurons and midbrain dopamine neurons.

Differentially expressed genes between human substantia nigra compacta (SNc) and ventral tegmental area (VTA) neurons.

Differentially expressed genes between motor neurons isolated from mouse cervical (cSC) and lumbar (lSC) spinal cord.

Degeneration of dopamine neurons in the midbrain causes symptoms of the movement disorder, Parkinson disease. Dopamine neurons are generated from proliferating progenitor cells localized in the embryonic ventral midbrain. However, it remains unclear for how long cells with dopamine progenitor character are retained and if there is any potential for...

The fatal disease amyotrophic lateral sclerosis (ALS) is characterized by the loss of somatic motor neurons leading to muscle wasting and paralysis. However, motor neurons in the oculomotor nucleus, controlling eye movement, are for unknown reasons spared. We found that insulin-like growth factor 2 (IGF-2) was maintained in oculomotor neurons in AL...

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with unknown origins. Neurodegeneration in ALS mouse models occurs together with signs of disrupted blood–spinal cord barrier (BSCB) and regressed capillary network, but the molecular pathways contributing to these vascular pathologies remain unknown. We show that motor neuron...

Neuromuscular junctions are primary pathological targets in the lethal motor neuron diseases spinal muscular atrophy (SMA) and amyotrophic lateral sclerosis (ALS). Synaptic pathology and denervation of target muscle fibers has been reported prior to the appearance of clinical symptoms in mouse models of both diseases, suggesting that neuromuscular...

The lethal disease amyotrophic lateral sclerosis (ALS) is characterized by the loss of somatic motor neurons. However, not all motor neurons are equally vulnerable to disease; certain groups are spared, including those in the oculomotor nucleus controlling eye movement. The reasons for this differential vulnerability remain unknown. Here we have id...