Mitochondrial activity is central to tissue homeostasis. Mitochondria dysfunction constitutes a hallmark of many genetic diseases and plays a key role in tumor progression. The essential role of mitochondria, added to their recently documented capacity to transfer from cell to cell, obviously contributes to their current interest. However, determining the proper role of mitochondria in defined biological contexts was hampered by the lack of suitable experimental tools. We designed a protocol (MitoCeption) to directly and quantitatively transfer mitochondria, isolated from cell type A, to recipient cell type B. We validated and quantified the effective mitochondria transfer by imaging, fluorescence-activated cell sorting (FACS) and mitochondrial DNA analysis. We show that the transfer of minute amounts of mesenchymal stem/stromal cell (MSC) mitochondria to cancer cells, a process otherwise occurring naturally in coculture, results in cancer cell enhanced oxidative phosphorylation (OXPHOS) activity and favors cancer cell proliferation and invasion. The MitoCeption technique, which can be applied to different cell systems, will therefore be a method of choice to analyze the metabolic modifications induced by exogenous mitochondria in host cells.
Low-threshold mechanoreceptor neurons (LTMs) of the dorsal root ganglia (DRG) are essential for touch sensation. They form highly specialized terminations in the skin and display stereotyped projections in the spinal cord. Functionally defined LTMs depend on neurotrophin signaling for their postnatal survival and functioning, but how these neurons arise during development is unknown. Here, we show that specific types of LTMs can be identified shortly after DRG genesis by unique expression of the MafA transcription factor, the Ret receptor and coreceptor GFRalpha2, and find that their specification is Ngn2 dependent. In mice lacking Ret, these LTMs display early differentiation defects, as revealed by reduced MafA expression, and at later stages their central and peripheral projections are compromised. Moreover, in MafA mutants, a discrete subset of LTMs display altered expression of neurotrophic factor receptors. Our results provide evidence that genetic interactions involving Ret and MafA progressively promote the differentiation and diversification of LTMs.
Eukaryotic cells harbor a small multiploid mitochondrial genome, organized in nucleoids spread within the mitochondrial network. Maintenance and distribution of mitochondrial DNA (mtDNA) are essential for energy metabolism, mitochondrial lineage in primordial germ cells, and to prevent mtDNA instability, which leads to many debilitating human diseases. Mounting evidence suggests that the actors of the mitochondrial network dynamics, among which is the intramitochondrial dynamin OPA1, might be involved in these processes. Here, using siRNAs specific to OPA1 alternate spliced exons, we evidenced that silencing of the OPA1 variants including exon 4b leads to mtDNA depletion, secondary to inhibition of mtDNA replication, and to marked alteration of mtDNA distribution in nucleoid and nucleoid distribution throughout the mitochondrial network. We demonstrate that a small hydrophobic 10-kDa peptide generated by cleavage of the OPA1-exon4b isoform is responsible for this process and show that this peptide is embedded in the inner membrane and colocalizes and coimmunoprecipitates with nucleoid components. We propose a novel synthetic model in which a peptide, including two transmembrane domains derived from the N terminus of the OPA1-exon4b isoform in vertebrates or from its ortholog in lower eukaryotes, might contribute to nucleoid attachment to the inner mitochondrial membrane and promotes mtDNA replication and distribution. Thus, this study places OPA1 as a direct actor in the maintenance of mitochondrial genome integrity.
and Frédérique Scamps. Axotomy-induced expression of calciumactivated chloride current in subpopulations of mouse dorsal root ganglion neurons. J Neurophysiol 90: 3764 -3773, 2003. First published August 27, 2003 10.1152/jn.00449.2003. Whole cell patchclamp recordings of calcium-activated chloride current [I Cl(Ca) ] were made from adult sensory neurons of naive and axotomized mouse L 4 -L 6 lumbar dorsal root ganglia after 1 day of culture in vitro. A basal I Cl(Ca) was specifically expressed in a subset of naive mediumdiameter neurons (30 -40 m). Prior nerve injury, induced by sciatic nerve transection 5 days before experiments, increased both I Cl(Ca) amplitude and its expression in medium-diameter neurons. Moreover, nerve injury also induced I Cl(Ca) expression in a new subpopulation of neurons, the large-diameter neurons (40 -50 m). Small-diameter neurons (inferior to 30 m) never expressed I Cl(Ca) . Regulated I Cl(Ca) expression was strongly correlated with injury-induced regenerative growth of sensory neurons in vitro and nerve regeneration in vivo. Cell culture on a substrate not permissive for growth, D,L-polyornithine, prevented both elongation growth and I Cl(Ca) expression in axotomized neurons. Regenerative growth and the induction of I Cl(Ca) expression take place 2 days after injury, peak after 5 days of conditioning in vivo, slowly declining thereafter to control values. The selective expression of I Cl(Ca) within medium-and large-diameter neurons conditioned for rapid, efficient growth suggests that these channels play a specific role in postinjury behavior of sensory neuron subpopulations such as neuropathic pain and/or axonal regeneration.
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