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The tau tubulin kinases TTBK1/2 promote accumulation of pathological TDP-43.

Liachko NF, McMillan PJ, Strovas TJ, Loomis E, Greenup L, Murrell JR, Ghetti B, Raskind MA, Montine TJ, Bird TD, Leverenz JB, Kraemer BC - PLoS Genet. (2014)

Bottom Line: TTBK1/2 overexpression drives phosphorylation and relocalization of TDP-43 from the nucleus to cytoplasmic inclusions reminiscent of neuropathologic changes in disease states.Furthermore, protein levels of TTBK1 and TTBK2 are increased in frontal cortex of FTLD-TDP patients, and TTBK1 and TTBK2 co-localize with TDP-43 inclusions in ALS spinal cord.These kinases may represent attractive targets for therapeutic intervention for TDP-43 proteinopathies such as ALS and FTLD-TDP.

View Article: PubMed Central - PubMed

Affiliation: Geriatric Research Education and Clinical Center, Veterans Affairs Puget Sound Health Care System, Seattle, Washington, United States of America; Department of Medicine, University of Washington, Seattle, Washington, United States of America.

ABSTRACT
Pathological aggregates of phosphorylated TDP-43 characterize amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD-TDP), two devastating groups of neurodegenerative disease. Kinase hyperactivity may be a consistent feature of ALS and FTLD-TDP, as phosphorylated TDP-43 is not observed in the absence of neurodegeneration. By examining changes in TDP-43 phosphorylation state, we have identified kinases controlling TDP-43 phosphorylation in a C. elegans model of ALS. In this kinome-wide survey, we identified homologs of the tau tubulin kinases 1 and 2 (TTBK1 and TTBK2), which were also identified in a prior screen for kinase modifiers of TDP-43 behavioral phenotypes. Using refined methodology, we demonstrate TTBK1 and TTBK2 directly phosphorylate TDP-43 in vitro and promote TDP-43 phosphorylation in mammalian cultured cells. TTBK1/2 overexpression drives phosphorylation and relocalization of TDP-43 from the nucleus to cytoplasmic inclusions reminiscent of neuropathologic changes in disease states. Furthermore, protein levels of TTBK1 and TTBK2 are increased in frontal cortex of FTLD-TDP patients, and TTBK1 and TTBK2 co-localize with TDP-43 inclusions in ALS spinal cord. These kinases may represent attractive targets for therapeutic intervention for TDP-43 proteinopathies such as ALS and FTLD-TDP.

No MeSH data available.


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Upregulated Tau tubulin kinases are also co-expressed with phospho-TDP-43 pathology.Representative photomicrographs depicting TTBK1 (A, C) and TTBK2 (B, D) immunoreactivity in cortical neurons in normal (A, B) and FTLD-TDP Type B (C, D) cases. The cellular distribution is both cytoplasmic and nuclear (insets), and immunoreactivity appears to be more widespread in FTLD cases relative to normal controls. Cortical layers I-VI are indicated (C). Quantification of immunostaining demonstrated a statistically significant increase in both TTBK1 (E) and TTBK2 (F) in FTLD cases compared to normal controls (**P = 0.003; ***P<0.0001). The distribution of phospho-TDP-43 immunoreactivity in the cortex of an FTLD case (G) overlaps with TTBK1 (C) and TTBK2 (D). Double label immunohistochemical experiments suggest co-localization of phospho-TDP-43 with TTBK1 (H) and TTBK2 (I) in an FTLD case. Scale bars: 100 µm A–D,G; 50 µm insets A–D; 25 µm H,I. See S5 Figure for controls for antibody specificity.
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pgen-1004803-g004: Upregulated Tau tubulin kinases are also co-expressed with phospho-TDP-43 pathology.Representative photomicrographs depicting TTBK1 (A, C) and TTBK2 (B, D) immunoreactivity in cortical neurons in normal (A, B) and FTLD-TDP Type B (C, D) cases. The cellular distribution is both cytoplasmic and nuclear (insets), and immunoreactivity appears to be more widespread in FTLD cases relative to normal controls. Cortical layers I-VI are indicated (C). Quantification of immunostaining demonstrated a statistically significant increase in both TTBK1 (E) and TTBK2 (F) in FTLD cases compared to normal controls (**P = 0.003; ***P<0.0001). The distribution of phospho-TDP-43 immunoreactivity in the cortex of an FTLD case (G) overlaps with TTBK1 (C) and TTBK2 (D). Double label immunohistochemical experiments suggest co-localization of phospho-TDP-43 with TTBK1 (H) and TTBK2 (I) in an FTLD case. Scale bars: 100 µm A–D,G; 50 µm insets A–D; 25 µm H,I. See S5 Figure for controls for antibody specificity.

Mentions: Fig. 4A–D demonstrates that TTBK1 and TTBK2 immunoreactivity is present in a subset of pyramidal neurons in the frontal cortex of both normal and FTLD cases. Immunoreactivity is more prominent in cortical layers II–VI compared to cortical layer I, where immunoreactivity is relatively sparse, and the cellular localization appears both nuclear and cytoplasmic (Fig. 4, insets). Furthermore, the distribution of TTBK1 and TTBK2 immunoreactivity appears to be more widespread in FTLD cases compared to normal controls. Optical density measurements relative to the proportional area for TTBK1 and TTBK2 immunostaining in frontal cortex confirmed a statistically significant increase in both TTBK1 (Fig. 4E) and TTBK2 (Fig. 4F) immunoreactive distribution in disease-affected subjects. This increase was observed in all FTLD cases surveyed relative to controls. Importantly, the distribution of TTBK1 and TTBK2 in the frontal cortex is consistent with the distribution of phosphorylated TDP-43 pathology in FTLD cases, where aggregates are sparse in cortical layer I, and more abundant in cortical layers II–VI, depending on the FTLD classification (Fig. 4G). To further demonstrate this relationship, we performed double label immunohistochemistry to determine if the tau tubulin kinases and phosphorylated TDP-43 co-expressed within the same neurons. Most neurons immunoreactive for phospho-TDP-43 were also immunoreactive for TTBK1 and TTBK2 (Fig. 4H, I).


The tau tubulin kinases TTBK1/2 promote accumulation of pathological TDP-43.

Liachko NF, McMillan PJ, Strovas TJ, Loomis E, Greenup L, Murrell JR, Ghetti B, Raskind MA, Montine TJ, Bird TD, Leverenz JB, Kraemer BC - PLoS Genet. (2014)

Upregulated Tau tubulin kinases are also co-expressed with phospho-TDP-43 pathology.Representative photomicrographs depicting TTBK1 (A, C) and TTBK2 (B, D) immunoreactivity in cortical neurons in normal (A, B) and FTLD-TDP Type B (C, D) cases. The cellular distribution is both cytoplasmic and nuclear (insets), and immunoreactivity appears to be more widespread in FTLD cases relative to normal controls. Cortical layers I-VI are indicated (C). Quantification of immunostaining demonstrated a statistically significant increase in both TTBK1 (E) and TTBK2 (F) in FTLD cases compared to normal controls (**P = 0.003; ***P<0.0001). The distribution of phospho-TDP-43 immunoreactivity in the cortex of an FTLD case (G) overlaps with TTBK1 (C) and TTBK2 (D). Double label immunohistochemical experiments suggest co-localization of phospho-TDP-43 with TTBK1 (H) and TTBK2 (I) in an FTLD case. Scale bars: 100 µm A–D,G; 50 µm insets A–D; 25 µm H,I. See S5 Figure for controls for antibody specificity.
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pgen-1004803-g004: Upregulated Tau tubulin kinases are also co-expressed with phospho-TDP-43 pathology.Representative photomicrographs depicting TTBK1 (A, C) and TTBK2 (B, D) immunoreactivity in cortical neurons in normal (A, B) and FTLD-TDP Type B (C, D) cases. The cellular distribution is both cytoplasmic and nuclear (insets), and immunoreactivity appears to be more widespread in FTLD cases relative to normal controls. Cortical layers I-VI are indicated (C). Quantification of immunostaining demonstrated a statistically significant increase in both TTBK1 (E) and TTBK2 (F) in FTLD cases compared to normal controls (**P = 0.003; ***P<0.0001). The distribution of phospho-TDP-43 immunoreactivity in the cortex of an FTLD case (G) overlaps with TTBK1 (C) and TTBK2 (D). Double label immunohistochemical experiments suggest co-localization of phospho-TDP-43 with TTBK1 (H) and TTBK2 (I) in an FTLD case. Scale bars: 100 µm A–D,G; 50 µm insets A–D; 25 µm H,I. See S5 Figure for controls for antibody specificity.
Mentions: Fig. 4A–D demonstrates that TTBK1 and TTBK2 immunoreactivity is present in a subset of pyramidal neurons in the frontal cortex of both normal and FTLD cases. Immunoreactivity is more prominent in cortical layers II–VI compared to cortical layer I, where immunoreactivity is relatively sparse, and the cellular localization appears both nuclear and cytoplasmic (Fig. 4, insets). Furthermore, the distribution of TTBK1 and TTBK2 immunoreactivity appears to be more widespread in FTLD cases compared to normal controls. Optical density measurements relative to the proportional area for TTBK1 and TTBK2 immunostaining in frontal cortex confirmed a statistically significant increase in both TTBK1 (Fig. 4E) and TTBK2 (Fig. 4F) immunoreactive distribution in disease-affected subjects. This increase was observed in all FTLD cases surveyed relative to controls. Importantly, the distribution of TTBK1 and TTBK2 in the frontal cortex is consistent with the distribution of phosphorylated TDP-43 pathology in FTLD cases, where aggregates are sparse in cortical layer I, and more abundant in cortical layers II–VI, depending on the FTLD classification (Fig. 4G). To further demonstrate this relationship, we performed double label immunohistochemistry to determine if the tau tubulin kinases and phosphorylated TDP-43 co-expressed within the same neurons. Most neurons immunoreactive for phospho-TDP-43 were also immunoreactive for TTBK1 and TTBK2 (Fig. 4H, I).

Bottom Line: TTBK1/2 overexpression drives phosphorylation and relocalization of TDP-43 from the nucleus to cytoplasmic inclusions reminiscent of neuropathologic changes in disease states.Furthermore, protein levels of TTBK1 and TTBK2 are increased in frontal cortex of FTLD-TDP patients, and TTBK1 and TTBK2 co-localize with TDP-43 inclusions in ALS spinal cord.These kinases may represent attractive targets for therapeutic intervention for TDP-43 proteinopathies such as ALS and FTLD-TDP.

View Article: PubMed Central - PubMed

Affiliation: Geriatric Research Education and Clinical Center, Veterans Affairs Puget Sound Health Care System, Seattle, Washington, United States of America; Department of Medicine, University of Washington, Seattle, Washington, United States of America.

ABSTRACT
Pathological aggregates of phosphorylated TDP-43 characterize amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD-TDP), two devastating groups of neurodegenerative disease. Kinase hyperactivity may be a consistent feature of ALS and FTLD-TDP, as phosphorylated TDP-43 is not observed in the absence of neurodegeneration. By examining changes in TDP-43 phosphorylation state, we have identified kinases controlling TDP-43 phosphorylation in a C. elegans model of ALS. In this kinome-wide survey, we identified homologs of the tau tubulin kinases 1 and 2 (TTBK1 and TTBK2), which were also identified in a prior screen for kinase modifiers of TDP-43 behavioral phenotypes. Using refined methodology, we demonstrate TTBK1 and TTBK2 directly phosphorylate TDP-43 in vitro and promote TDP-43 phosphorylation in mammalian cultured cells. TTBK1/2 overexpression drives phosphorylation and relocalization of TDP-43 from the nucleus to cytoplasmic inclusions reminiscent of neuropathologic changes in disease states. Furthermore, protein levels of TTBK1 and TTBK2 are increased in frontal cortex of FTLD-TDP patients, and TTBK1 and TTBK2 co-localize with TDP-43 inclusions in ALS spinal cord. These kinases may represent attractive targets for therapeutic intervention for TDP-43 proteinopathies such as ALS and FTLD-TDP.

No MeSH data available.


Related in: MedlinePlus