Biomedical subjects

Cheolwha Jung

Publications and source records attributed to Cheolwha Jung.

9 recordsLinked to original sources

PubMed2005

The high and middle molecular weight neurofilament subunits regulate the association of neurofilaments with kinesin: inhibition by phosphorylation of the high molecular weight subunit.

Kinesin participates in axonal transport of neurofilaments (NFs), but the mode by which they attach to kinesin is unclear. We compared the association of NFs with kinesin in mice expressing or lacking NF-H or NF-M. In normal and M-/- mice, the leading edge of metabolically labeled NF subunits was selectively co-precipitated with kinesin. By contrast, the entire wave of radiolabeled subunits co-precipitated with kinesin in H-/- mice. Similar bulk levels of NFs co-precipitated with kinesin from normal and H-/- mice, but reduced levels co-precipitated from M-/- mice. These data suggest that both NF-H and NF-M regulate the association of NFs with kinesin. They further indicate that phosphorylation of NF-H dissociates NFs from kinesin and provides a mechanism by which NF-H phosphorylation can contribute to the slowing of NF axonal transport.

Animals↗

PubMed2005

Inhibition of chaperone activity is a shared property of several Cu,Zn-superoxide dismutase mutants that cause amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by progressive motor neuron degeneration, paralysis, and death. Mutant Cu,Zn-superoxide dismutase (SOD1) causes a subset of ALS by an unidentified toxic property. Increasing evidence suggests that chaperone dysfunction plays a role in motor neuron degeneration in ALS. To investigate the relationship between mutant SOD1 expression and chaperone dysfunction, we measured chaperone function in central nervous system tissue lysates from normal mice and transgenic mice expressing human SOD1 variants. We observed a significant decrease in chaperone activity in tissues from mice expressing ALS-linked mutant SOD1 but not control mice expressing human wild type SOD1. This decrease was detected only in the spinal cord, became apparent by 60 days of age (before the onset of muscle weakness and significant motor neuron loss), and persisted throughout the late stages. In addition, this impairment of chaperone activity occurred only in cytosolic but not in mitochondrial and nuclear fractions. Furthermore, multiple recombinant human SOD1 mutants with differing biochemical and biophysical properties inhibited chaperone function in a cell-free extract of normal mouse spinal cords. Thus, mutant SOD1 proteins may impair chaperone function independent of gene expression in vivo, and this inhibition may be a shared property of ALS-linked mutant SOD1 proteins.

Amyotrophic Lateral Sclerosis↗

PubMed2004

Neurofilament transport is dependent on actin and myosin.

Real-time analyses have revealed that some newly synthesized neurofilament (NF) subunits translocate into and along axonal neurites by moving along the inner plasma membrane surface, suggesting that they may translocate against the submembrane actin cortex. We therefore examined whether or not NF axonal transport was dependent on actin and myosin. Perturbation of filamentous actin in NB2a/d1 cells with cytochalasin B inhibited translocation of subunits into axonal neurites and inhibited bidirectional translocation of NF subunits within neurites. Intravitreal injection of cytochalasin B inhibited NF axonal transport in optic axons in a dose-response manner. NF subunits were coprecipitated from NB2a/d1 cells by an anti-myosin antibody, and myosin colocalized with NFs in immunofluorescent analyses. The myosin light chain kinase inhibitor ML-7 and the myosin ATPase inhibitor 2,3-butanedione-2-monoxime perturbed NF translocation within NB2a/d1 axonal neurites. These findings suggest that some NF subunits may undergo axonal transport via myosin-mediated interactions with the actin cortex.

Actins↗

PubMed2004

Neurofilament subunits undergo more rapid translocation within retinas than in optic axons.

Axonal transport of neurofilaments (NFs) has long been considered to be regulated by phosphorylation, although recent studies have challenged this hypothesis. Our prior analyses of axonal transport in optic axons demonstrated two distinct NF transport rates that spatially and temporally correlated with changes in NF phosphorylation. In our prior studies, we focused on subunits already within axons. Re-examination of these data using additional approaches and examining additional earlier time points have allowed us to calculate rates at which subunits transport out of retinas and into optic axons. NF subunits were radiolabeled by intravitreal injection of 35S-methionine. NF axonal transport was monitored by following the location of the front of radiolabeled subunits immunoprecipitated from retinas and segments of optic axons, which demonstrated four distinct transport rates. Subunits within retinas exhibited the fastest rate, and underwent a 50% slowing upon exiting the retina and entering optic axons. While this slowing could be due to a regional caliber increase and/or regional increase in NF phosphorylation within the first segment, prior studies indicated that inhibition of phosphatase activities increased NF phosphorylation within retinas and slowed NF subunit exit from retinas to a degree similar to that normally observed within the first segment of axons, suggesting that regional phosphorylation played a major role in slowing of NF transport following their exit from the retina. These findings provide additional support for the notion that phosphorylation regulates NF axonal transport.

Animals↗

PubMed2003

ALS-associated mutant SOD1G93A causes mitochondrial vacuolation by expansion of the intermembrane space and by involvement of SOD1 aggregation and peroxisomes.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is an age-dependent neurodegenerative disease that causes motor neuron degeneration, paralysis and death. Mutations in Cu, Zn superoxide dismutase (SOD1) are one cause for the familial form of this disease. Transgenic mice expressing mutant SOD1 develop age-dependent motor neuron degeneration, skeletal muscle weakness, paralysis and death similar to humans. The mechanism whereby mutant SOD1 induces motor neuron degeneration is not understood but widespread mitochondrial vacuolation has been observed during early phases of motor neuron degeneration. How this vacuolation develops is not clear, but could involve autophagic vacuolation, mitochondrial permeability transition (MPT) or uncharacterized mechanisms. To determine which of these possibilities are true, we examined the vacuolar patterns in detail in transgenic mice expressing mutant SOD1G93A. RESULTS: Vacuolar patterns revealed by electron microscopy (EM) suggest that vacuoles originate from the expansion of the mitochondrial intermembrane space and extension of the outer mitochondrial membrane. Immunofluorescence microscopy and immuno-gold electron microscopy reveal that vacuoles are bounded by SOD1 and mitochondrial outer membrane markers, but the inner mitochondrial membrane marker is located in focal areas inside the vacuoles. Small vacuoles contain cytochrome c while large vacuoles are porous and lack cytochrome c. Vacuoles lack lysosomal signal but contain abundant peroxisomes and SOD1 aggregates. CONCLUSION: These findings demonstrate that mutant SOD1, possibly by toxicity associated with its aggregation, causes mitochondrial degeneration by inducing extension and leakage of the outer mitochondrial membrane, and expansion of the intermembrane space. This could release the pro-cell death molecules normally residing in the intermembrane space and initiate motor neuron degeneration. This Mitochondrial Vacuolation by Intermembrane Space Expansion (MVISE) fits neither MPT nor autophagic vacuolation mechanisms, and thus, is a previously uncharacterized mechanism of mitochondrial degeneration in mammalian CNS.

Amyotrophic Lateral Sclerosis↗

PubMed2003

Does neurofilament phosphorylation regulate axonal transport?

Phosphorylation of neurofilaments has long been considered to regulate their axonal transport rate and, in doing so, to provide stability to mature axons. Interpretation of data recently obtained following C-terminal deletion experiments has prompted a challenge to this hypothesis. We present evidence that these deletion studies remain consistent with, rather than refute, a role for C-terminal phosphorylation in regulation of neurofilament axonal transport.

Amino Acid Sequence↗

PubMed2002

A quantitative histochemical assay for activities of mitochondrial electron transport chain complexes in mouse spinal cord sections.

Mitochondrial dysfunction and degeneration are associated with neurodegenerative disorders. A dysfunctional mitochondrial electron transport chain (ETC) impairs ATP production and accelerates the generation of free radicals. To quantify ETC activity, solution-spectrophotometric assays and histochemical reactions on blue native polyacrylamide gel electrophoresis (BN-PAGE) gels have been used. These methods, however, do not provide information regarding mitochondrial ETC activities associated with specific regions in the central nervous system (CNS). Because neurodegenerative diseases often strike a specific subset of neurons within specific regions in the CNS, reliable methods for quantifying mitochondrial ETC activities in selected CNS regions are needed. We have studied the quantitative range of in situ histochemical assays for ETC complex I, II and IV and determined the optimal conditions for quantification of these ETC complex activities. We also demonstrate that these assays can detect a decrease in mitochondrial ETC activities in the ventral horn of spinal cords isolated from a transgenic mouse model for amyotrophic lateral sclerosis (ALS), a fatal neurodegenerative disease.

Amyotrophic Lateral Sclerosis↗

PubMed2002

Mutant Cu, Zn superoxide dismutase that causes motoneuron degeneration is present in mitochondria in the CNS.

Mutations in Cu, Zn superoxide dismutase (SOD1) cause a fraction of amyotrophic lateral sclerosis (ALS), which involves motoneuron degeneration, paralysis, and death. An acquired activity by mutant SOD1 is responsible for the cellular toxicity, but how mutant SOD1 kills motoneurons is unclear. In transgenic mouse models of ALS, mitochondrial degeneration occurs early, before disease onset, raising the question of how mutant SOD1 damages mitochondria. Here we investigate the intracellular localization of SOD1 in the CNS to determine whether SOD1 is present in mitochondria, where it could directly damage this organelle. We show that endogenous mouse SOD1, wild-type human, and mutant human SOD1 (G93A), when expressed as transgenes, are colocalized with mitochondria in spinal cord by immunofluorescence confocal microscopy. By immunoelectron microscopy, we show that SOD1 is present within mitochondria at similar concentrations as in the cytoplasm. Thus SOD1, in addition to being a cytosolic enzyme, is present inside mitochondria in the CNS.

Amino Acid Substitution↗

PubMed2002

Mitochondrial electron transport chain complex dysfunction in a transgenic mouse model for amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis is a fatal neurodegenerative disease that causes degeneration of motoneurons. Mutation of Cu,Zn superoxide dismutase (SOD1) is one cause for this disease. In mice, expression of mutant protein causes motoneuron degeneration and paralysis resembling the human disease. Morphological change, indicative of mitochondrial damage, occurs at early stages of the disease. To determine whether mitochondrial function changes during the course of disease progression, enzyme activities of mitochondrial electron transport chain in spinal cords from mice at different disease stages were measured using three different methods: spectrophotometric assay, in situ histochemical enzyme assay, and blue native gel electrophoresis combined with in-gel histochemical reaction. The enzyme activities were decreased in the spinal cord, particularly in the ventral horn, beginning at early disease stages. This decrease persisted throughout the course of disease progression. This decrease was not detected in the spinal cords of non-transgenic animals, of mice expressing the wild-type protein, and in cerebellum and dorsal horn of the spinal cords from mice expressing mutant protein. These results demonstrate a functional defect in mitochondria in the ventral horn region and support the view that mitochondrial damage plays a role in mutant SOD1-induced motoneuron degeneration pathway.

Amyotrophic Lateral Sclerosis↗