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Impaired Coenzyme A metabolism affects histone and tubulin acetylation in Drosophila and human cell models of pantothenate kinase associated neurodegeneration.

Siudeja K, Srinivasan B, Xu L, Rana A, de Jong J, Nollen EA, Jackowski S, Sanford L, Hayflick S, Sibon OC - EMBO Mol Med (2011)

Bottom Line: Previously, we observed reduced Coenzyme A levels in a Drosophila model for PKAN.Here we investigate whether decreased levels of the central metabolite Coenzyme A induce alterations in protein acetylation and whether this correlates with specific phenotypes of PKAN models.We show that in various organisms proper Coenzyme A metabolism is required for maintenance of histone- and tubulin acetylation, and decreased acetylation of these proteins is associated with an impaired DNA damage response, decreased locomotor function and decreased survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Radiation and Stress Cell Biology, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.

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Levels of acetylated tubulin and histones are decreased in dPANK/Fbl-depleted cellsCell extracts of control cells and dPANK/Fbl-depleted cells (by RNAi) were analysed by Western blot using antibodies specifically recognizing acetylated-tubulin. Efficiency of RNAi was determined by using a dPANK/Fbl antibody and tubulin was used as a loading control. Control cells and dPANK/Fbl-depleted cells were left untreated, treated with pantethine, with TSA or with TSA and pantethine.Quantification of the relative levels of tubulin acetylation under the conditions presented in A compared to control cells.Cell extracts of control cells and dPANK/Fbl-depleted cells were analysed using Western blot to determine acetylation levels of specific histones. Specific antibodies were used to detect levels of acetylated histone 3 and acetylated histone 4. Control and dPANK/Fbl-depleted cells were left untreated or were treated with pantethine, with TSA or with TSA and pantethine. The efficiency of the RNAi treatment was investigated by the use of an antibody against dPANK/Fbl. H2A was used as a loading control.Quantification of the relative levels of histone acetylation under the conditions presented in C compared to control cells.
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fig02: Levels of acetylated tubulin and histones are decreased in dPANK/Fbl-depleted cellsCell extracts of control cells and dPANK/Fbl-depleted cells (by RNAi) were analysed by Western blot using antibodies specifically recognizing acetylated-tubulin. Efficiency of RNAi was determined by using a dPANK/Fbl antibody and tubulin was used as a loading control. Control cells and dPANK/Fbl-depleted cells were left untreated, treated with pantethine, with TSA or with TSA and pantethine.Quantification of the relative levels of tubulin acetylation under the conditions presented in A compared to control cells.Cell extracts of control cells and dPANK/Fbl-depleted cells were analysed using Western blot to determine acetylation levels of specific histones. Specific antibodies were used to detect levels of acetylated histone 3 and acetylated histone 4. Control and dPANK/Fbl-depleted cells were left untreated or were treated with pantethine, with TSA or with TSA and pantethine. The efficiency of the RNAi treatment was investigated by the use of an antibody against dPANK/Fbl. H2A was used as a loading control.Quantification of the relative levels of histone acetylation under the conditions presented in C compared to control cells.

Mentions: Next we identified the indicated proteins starting with the protein migrating at 55 kD. Previously it has been shown that tubulin is a protein that can be acetylated (Akella et al, 2010; LeDizet and Piperno, 1987) and its molecular weight matches with the protein indicated by the upper asterisk in Fig 1B. Western blot analysis using antibodies that recognize acetylated-tubulin confirmed that indeed in dPANK/Fbl depleted cells; levels of acetylated-tubulin, but not the total levels of tubulin are decreased (Fig 2A compare lane 1 with lane 3, see Fig 2B for quantification). Acetylation levels of tubulin in dPANK/Fbl depleted cells are restored by addition of pantethine and by addition of the HDAC inhibitor TSA (Fig 2A and B) confirming further the results of Fig 1B and C. Altogether these data demonstrate that a decrease in CoA levels coincides with decreased levels of acetylated-tubulin.


Impaired Coenzyme A metabolism affects histone and tubulin acetylation in Drosophila and human cell models of pantothenate kinase associated neurodegeneration.

Siudeja K, Srinivasan B, Xu L, Rana A, de Jong J, Nollen EA, Jackowski S, Sanford L, Hayflick S, Sibon OC - EMBO Mol Med (2011)

Levels of acetylated tubulin and histones are decreased in dPANK/Fbl-depleted cellsCell extracts of control cells and dPANK/Fbl-depleted cells (by RNAi) were analysed by Western blot using antibodies specifically recognizing acetylated-tubulin. Efficiency of RNAi was determined by using a dPANK/Fbl antibody and tubulin was used as a loading control. Control cells and dPANK/Fbl-depleted cells were left untreated, treated with pantethine, with TSA or with TSA and pantethine.Quantification of the relative levels of tubulin acetylation under the conditions presented in A compared to control cells.Cell extracts of control cells and dPANK/Fbl-depleted cells were analysed using Western blot to determine acetylation levels of specific histones. Specific antibodies were used to detect levels of acetylated histone 3 and acetylated histone 4. Control and dPANK/Fbl-depleted cells were left untreated or were treated with pantethine, with TSA or with TSA and pantethine. The efficiency of the RNAi treatment was investigated by the use of an antibody against dPANK/Fbl. H2A was used as a loading control.Quantification of the relative levels of histone acetylation under the conditions presented in C compared to control cells.
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Related In: Results  -  Collection

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fig02: Levels of acetylated tubulin and histones are decreased in dPANK/Fbl-depleted cellsCell extracts of control cells and dPANK/Fbl-depleted cells (by RNAi) were analysed by Western blot using antibodies specifically recognizing acetylated-tubulin. Efficiency of RNAi was determined by using a dPANK/Fbl antibody and tubulin was used as a loading control. Control cells and dPANK/Fbl-depleted cells were left untreated, treated with pantethine, with TSA or with TSA and pantethine.Quantification of the relative levels of tubulin acetylation under the conditions presented in A compared to control cells.Cell extracts of control cells and dPANK/Fbl-depleted cells were analysed using Western blot to determine acetylation levels of specific histones. Specific antibodies were used to detect levels of acetylated histone 3 and acetylated histone 4. Control and dPANK/Fbl-depleted cells were left untreated or were treated with pantethine, with TSA or with TSA and pantethine. The efficiency of the RNAi treatment was investigated by the use of an antibody against dPANK/Fbl. H2A was used as a loading control.Quantification of the relative levels of histone acetylation under the conditions presented in C compared to control cells.
Mentions: Next we identified the indicated proteins starting with the protein migrating at 55 kD. Previously it has been shown that tubulin is a protein that can be acetylated (Akella et al, 2010; LeDizet and Piperno, 1987) and its molecular weight matches with the protein indicated by the upper asterisk in Fig 1B. Western blot analysis using antibodies that recognize acetylated-tubulin confirmed that indeed in dPANK/Fbl depleted cells; levels of acetylated-tubulin, but not the total levels of tubulin are decreased (Fig 2A compare lane 1 with lane 3, see Fig 2B for quantification). Acetylation levels of tubulin in dPANK/Fbl depleted cells are restored by addition of pantethine and by addition of the HDAC inhibitor TSA (Fig 2A and B) confirming further the results of Fig 1B and C. Altogether these data demonstrate that a decrease in CoA levels coincides with decreased levels of acetylated-tubulin.

Bottom Line: Previously, we observed reduced Coenzyme A levels in a Drosophila model for PKAN.Here we investigate whether decreased levels of the central metabolite Coenzyme A induce alterations in protein acetylation and whether this correlates with specific phenotypes of PKAN models.We show that in various organisms proper Coenzyme A metabolism is required for maintenance of histone- and tubulin acetylation, and decreased acetylation of these proteins is associated with an impaired DNA damage response, decreased locomotor function and decreased survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Radiation and Stress Cell Biology, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.

Show MeSH
Related in: MedlinePlus