ADP-ribosylation of the   proteins induces growth inhibition, neurite outgrowth and acetylcholine esterase in cultured PC-12 cells

Nishiki, Tadaaki; Narumiya, Shuh; Morii, Narito; Yamamoto, Misaki; Fujiwara, Motohatsu; Kamata, Yoichi; Sakaguchi, Genji; Kozaki, Shunji

doi:10.1016/0006-291x(90)91760-p

Cited by 123 publications

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“…It has been reported that C3 enzyme induced shape changes and process formation in PC-12 cells [15], and that C3 enzyme and DBcAMP induced similar shape changes and process formation in NG108-15 cells [8]. In the present study, the morphological changes caused by C3 enzyme were slightly different from those caused by NGF or DBcAMP.…”

Section: Discussioncontrasting

confidence: 60%

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Botulinum C3 Enzyme Changes the Lactate Dehydrogenase Isozyme Pattern of Primary Culture of Neurons.

Watanabe

Morimatsu

Syuto

2000

The Journal of Veterinary Medical Science

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ABSTRACT. Changes in the lactate dehydrogenase (LDH) isozyme pattern of primary culture of neurons treated with botulinum C3 enzyme were examined in order to elucidate the functional changes accompanying the morphological change that follows ADP-ribosylation of Rho protein. Primary neurons were prepared from the cerebrum of ICR mouse embryos on day 15. Neurons were cultured in MEM with 10% fetal calf serum at 37°C. In the neurons treated with C3 enzyme, a typical morphological change was observed after 24 hr, and the LDH isozyme pattern was changed after 72 hr. The ratio of H-subunit to M-subunit in LDH was decreased by C3 treatment, suggesting the induction of a state of lower intracellular oxygen consumption in neurons in the primary cultures.-KEY WORDS: botulinum C3, C3 enzyme, LDH isozyme pattern, mouse neuron, primary culture.

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Section: Discussioncontrasting

confidence: 60%

“…In neural cell lines such as PC-12 [15], C6 and NG108-15 [8], the shape of the cell body is changed and neurite outgrowth is induced when C3 enzyme is added to the culture medium. Furthermore, it has been reported that the microinjection of Rho protein ADP-ribosylated with C3 enzyme into cultured cells results in morphological change [17].…”

Section: Methodsmentioning

confidence: 99%

Botulinum C3 Enzyme Changes the Lactate Dehydrogenase Isozyme Pattern of Primary Culture of Neurons.

Watanabe

Morimatsu

Syuto

2000

The Journal of Veterinary Medical Science

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“…C3 exoenzyme has been used to elucidate the role of rho p21 in various cellular functions [5][6][7][8][9][10][11][12][24][25][26][27][28][29]. There have been, however, no reports using site-directed enzyme mutants to demonstrate the relationship between its ADP-ribosylation activity and the biological response.…”

Section: Resultsmentioning

confidence: 99%

“…Clostridium botulinum C3 exoenzyme [3] specifically ADPribosylates the small molecular weight GTP binding protein rho p21 at the Asn 41 residue located in a putative effector binding domain [4]. Treatment of cells with C3 exoenzyme inhibits stimulus-induced actin filament organization and cell adhesion [5][6][7][8][9][10][11][12], indicating that rho p21 mediates these cellular responses as a molecular switch and that C3 exoenzyme inhibits these functions of rho p21. Identification of the amino acid residues of the C3 exoenzyme interacting with NAD is important in elucidating the structure of its catalytic site and in understanding a mechanism of the ADP-ribosylation.…”

Section: Introductionmentioning

confidence: 99%

Identification of Glu¹⁷³ as the critical amino acid residue for the ADP‐ribosyltransferase activity of Clostridium botulinum C3 exoenzyme

et al. 1995

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Clostridium botulinum C3 exoenzyme specifically ADP-ribosylates rho-p21 in eukaryotic cells. Trp Is and Glu 173 of this enzyme were substituted with other amino acids via sitedirected mutagenesis. All substitutions at Glu 173 caused a significant reduction in affinity for NAD and diminished ADPribosyltransferase activity. On the other hand, the activity of enzymes with the substitution at Trp Is remained intact. Swiss 3T3 cells treated with the enzyme with the Trp TM substitution showed the typical morphologic changes of the C3 exoenzyme phenotype. In contrast, no changes were found in cells incubated with the Glu~73-substituted enzyme. These results indicate that the Glu 173 residue of the C3 exoenzyme plays a key role in interacting with NAD and in expression of ADP-ribosyltransferase activity, which is essential for the phenotypic change by C3 exoenzyme treatment.

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“…Since it has been reported that C3 enzyme modifies Rho proteins, and induces differentiation-like morphological change in PC-12, NG108-15 and C-6 cells [12,18], we examined the potential of C3 enzyme for medical use, especially in dendrite induction. We first examined the C3 act ivity for the induction of differentia tion-like morphological changes of primary cultured neurons.…”

Section: Discussionmentioning

confidence: 99%

The Evaluation of the Potential of Botulinum C3 Enzyme as an Exogenous Differentiation Inducing Factor to Neurons.

Watanabe

Morimatsu

Syuto

2000

The Journal of Veterinary Medical Science

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ABSTRACT. Botulinum C3 enzyme produced by Clostridium botulinum type C and D strains modifies Rho proteins. In a previous study, we observed that the LDH isozyme pattern of neurons treated with C3 enzyme was different from that induced with endogenous growth factor of neurons such as NGF [21]. This type of change is considered to have an advantage in the medical use of C3 enzyme for neural disorder. To determine the functional similarity of C3-treated neurons to control and NGF-treated neurons, we examined the responses of C3-treated neurons to various drugs, including some neurotransmitters, by measuring the rise of intracellular Ca ions into the neurons. The time course of the rise of intracellular Ca ions induced by high concentration of potassium in the C3-treated neurons was similar to that in the NGF-treated neurons. The C3-treated neurons responded to glutamic acid, aspartic acid, kainic acid, γ-aminobutylic acid, muscarine and ACh with similar time courses and magnitudes as the control neurons. These results suggest that the C3 enzyme induces the functional differentiation of neurons, and that C3 enzyme has the potential for the medical use as an exogenous differentiation-inducing factor of neurons.-KEY WORDS: botulinum C3 enzyme, Ca 2+ , differentiation, neuron, neurotransmitter.Botulinum C3 enzyme was first reported as a new type of botulinum toxin with an ADP-ribosyl transferase activity [1]. C3 enzyme has the unique function of inducing a morphological change in cells when they are incubated with it. Since the morphological change of neural cells caused by C3 enzyme seems to be a differentiation of cells, the effects of C3 enzyme on induction of differentiation marker enzymes have been examined. C3 enzyme increases choline acetyl transferase, acetylcholine esterase and 2', 3'-cyclic-nucleotide 3'-phosphohydrolase in NG108-15 and C6 cell lines [12]. These facts suggest that C3 enzyme has a differentiation-inducing activity on neurons.We obtained results in previous studies on the change of the lactate dehydrogenase (LDH) isozyme pattern; the ratio of H-subunit to M-subunit in LDH was decreased significantly by C3 enzyme treatment in primary cultured neurons. Also, the ratio of H/M in LDH was increased by nerve growth factor (NGF) and N6, 2'-O-dibutyryl 3', 5'-cyclic monophosphate (DBcAMP) [21]. These results suggest that the C3 enzyme induces an intracellular state in which less oxygen is consumed in primary cultured neurons. The change in the characteristics of the neuron to the intracellular reduced oxygen consumption state has important benefits for the treatment of ischemia. These results motivated us to study the potential of the C3 enzyme as an exogenous differentiation-inducing factor for neurons. To evaluate the potential of C3 enzyme, it is necessary to determine the response of the C3-treated neurons to various physiological and pharmacological substances.In this study, we examined the responses of C3-treated neurons to various drugs, including some neurotransmitters, by measuring the rise o...

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ADP-ribosylation of the proteins induces growth inhibition, neurite outgrowth and acetylcholine esterase in cultured PC-12 cells

Cited by 123 publications

References 19 publications

Botulinum C3 Enzyme Changes the Lactate Dehydrogenase Isozyme Pattern of Primary Culture of Neurons.

Botulinum C3 Enzyme Changes the Lactate Dehydrogenase Isozyme Pattern of Primary Culture of Neurons.

Identification of Glu¹⁷³ as the critical amino acid residue for the ADP‐ribosyltransferase activity of Clostridium botulinum C3 exoenzyme

The Evaluation of the Potential of Botulinum C3 Enzyme as an Exogenous Differentiation Inducing Factor to Neurons.

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ADP-ribosylation of the proteins induces growth inhibition, neurite outgrowth and acetylcholine esterase in cultured PC-12 cells

Cited by 123 publications

References 19 publications

Botulinum C3 Enzyme Changes the Lactate Dehydrogenase Isozyme Pattern of Primary Culture of Neurons.

Botulinum C3 Enzyme Changes the Lactate Dehydrogenase Isozyme Pattern of Primary Culture of Neurons.

Identification of Glu173 as the critical amino acid residue for the ADP‐ribosyltransferase activity of Clostridium botulinum C3 exoenzyme

The Evaluation of the Potential of Botulinum C3 Enzyme as an Exogenous Differentiation Inducing Factor to Neurons.

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Identification of Glu¹⁷³ as the critical amino acid residue for the ADP‐ribosyltransferase activity of Clostridium botulinum C3 exoenzyme