scholarly journals PKCγ-Mediated Phosphorylation of CRMP2 Regulates Dendritic Outgrowth in Cerebellar Purkinje Cells

2020 ◽  
Vol 57 (12) ◽  
pp. 5150-5166
Author(s):  
Sabine C. Winkler ◽  
Etsuko Shimobayashi ◽  
Josef P. Kapfhammer
Keyword(s):  
Purkinje Cell ◽  
Purkinje Cells ◽  
Synaptic Function ◽  
Mass Spectrometry Analysis ◽  
Spinocerebellar Ataxia Type ◽  
Proximity Ligation Assay ◽  
Wild Type ◽  
Dynamic Regulation ◽  
Dendritic Development ◽  
Cerebellar Purkinje Cells

Abstract The signalling protein PKCγ is a major regulator of Purkinje cell development and synaptic function. We have shown previously that increased PKCγ activity impairs dendritic development of cerebellar Purkinje cells. Mutations in the protein kinase Cγ gene (PRKCG) cause spinocerebellar ataxia type 14 (SCA14). In a transgenic mouse model of SCA14 expressing the human S361G mutation, Purkinje cell dendritic development is impaired in cerebellar slice cultures similar to pharmacological activation of PKC. The mechanisms of PKCγ-driven inhibition of dendritic growth are still unclear. Using immunoprecipitation-coupled mass spectrometry analysis, we have identified collapsin response mediator protein 2 (CRMP2) as a protein interacting with constitutive active PKCγ(S361G) and confirmed the interaction with the Duolink™ proximity ligation assay. We show that in cerebellar slice cultures from PKCγ(S361G)-mice, phosphorylation of CRMP2 at the known PKC target site Thr555 is increased in Purkinje cells confirming phosphorylation of CRMP2 by PKCγ. miRNA-mediated CRMP2 knockdown decreased Purkinje cell dendritic outgrowth in dissociated cerebellar cultures as did the transfection of CRMP2 mutants with a modified Thr555 site. In contrast, dendritic development was normal after wild-type CRMP2 overexpression. In a novel knock-in mouse expressing only the phospho-defective T555A-mutant CRMP2, Purkinje cell dendritic development was reduced in dissociated cultures. This reduction could be rescued by transfecting wild-type CRMP2 but only partially by the phospho-mimetic T555D-mutant. Our findings establish CRMP2 as an important target of PKCγ phosphorylation in Purkinje cells mediating its control of dendritic development. Dynamic regulation of CRMP2 phosphorylation via PKCγ is required for its correct function.

scholarly journals Progress in pathogenesis studies of spinocerebellar ataxia type 1

1999 ◽  
Vol 354 (1386) ◽  
pp. 1079-1081 ◽  
Author(s):  
Christopher J. Cummings ◽  
Harry T. Orr ◽  
Huda Y. Zoghbi
Keyword(s):  
Transgenic Mice ◽  
Purkinje Cell ◽  
Purkinje Cells ◽  
Nuclear Localization ◽  
Hela Cells ◽  
Protein Misfolding ◽  
Spinocerebellar Ataxia Type ◽  
Spinocerebellar Ataxia Type 1 ◽  
Cerebellar Purkinje Cells

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited disorder characterized by progressive loss of coordination, motor impairment and the degeneration of cerebellar Purkinje cells, spinocerebellar tracts and brainstem nuclei. Many dominantly inherited neurodegenerative diseases share the mutational basis of SCA1: the expansion of a translated CAG repeat coding for glutamine. Mice lacking ataxin-1 display learning deficits and altered hippocampal synaptic plasticity but none of the abnormalities seen in human SCA1; mice expressing ataxin-1 with an expanded CAG tract (82 glutamine residues), however, develop Purkinje cell pathology and ataxia. These results suggest that mutant ataxin-1 gains a novel function that leads to neuronal degeneration. This novel function might involve aberrant interaction(s) with cell-specific protein(s), which in turn might explain the selective neuronal pathology. Mutant ataxin-1 interacts preferentially with a leucine-rich acidic nuclear protein that is abundantly expressed in cerebellar Purkinje cells and other brain regions affected in SCA1. Immunolocalization studies in affected neurons of patients and SCA1 transgenic mice showed that mutant ataxin-1 localizes to a single, ubiquitin-positive nuclear inclusion (NI) that alters the distribution of the proteasome and certain chaperones. Further analysis of NIs in transfected HeLa cells established that the proteasome and chaperone proteins co-localize with ataxin-1 aggregates. Moreover, overexpression of the chaperone HDJ-2/HSDJ in HeLa cells decreased ataxin-1 aggregation, suggesting that protein misfolding might underlie NI formation. To assess the importance of the nuclear localization of ataxin-1 and its role in SCA1 pathogenesis, two lines of transgenic mice were generated. In the first line, the nuclear localization signal was mutated so that full-length mutant ataxin-1 would remain in the cytoplasm; mice from this line did not develop any ataxia or pathology. This suggests that mutant ataxin-1 is pathogenic only in the nucleus. To assess the role of the aggregates, transgenic mice were generated with mutant ataxin-1 without the self-association domain (SAD) essential for aggregate formation. These mice developed ataxia and Purkinje cell abnormalities similar to those seen in SCA1 transgenic mice carrying full-length mutant ataxin-1, but lacked NIs. The nuclear milieu is thus a critical factor in SCA1 pathogenesis, but large NIs are not needed to initiate pathogenesis. They might instead be downstream of the primary pathogenic steps. Given the accumulated evidence, we propose the following model for SCA1 pathogenesis: expansion of the polyglutamine tract alters the conformation of ataxin-1, causing it to misfold. This in turn leads to aberrant protein interactions. Cell specificity is determined by the cell-specific proteins interacting with ataxin-1. Submicroscopic protein aggregation might occur because of protein misfolding, and those aggregates become detectable as NIs as the disease advances. Proteasome redistribution to the NI might contribute to disease progression by disturbing proteolysis and subsequent vital cellular functions.

scholarly journals Activation of MAP3K DLK and LZK in Purkinje Cells Causes Rapid and Slow Degeneration Depending on Signaling Strength

2020 ◽  
Author(s):  
Yunbo Li ◽  
Erin M Ritchie ◽  
Christopher L. Steinke ◽  
Cai Qi ◽  
Lizhen Chen ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Purkinje Cell ◽  
Purkinje Cells ◽  
Leucine Zipper ◽  
Activity Levels ◽  
Cell Type ◽  
Cell Degeneration ◽  
Mechanistic Basis ◽  
Cerebellar Purkinje Cells ◽  
Jnk Activation

SummaryThe conserved MAP3K Dual leucine zipper kinases can activate JNK via MKK4 or MKK7. Vertebrate DLK and LZK share similar biochemical activities and undergo auto-activation upon increased expression. Depending on cell-type and nature of insults DLK and LZK can induce pro-regenerative, pro-apoptotic or pro-degenerative responses, although the mechanistic basis of their action is not well understood. Here, we investigated these two MAP3Ks in cerebellar Purkinje cells using loss- and gain-of function mouse models. While loss of each or both kinases does not cause discernible defects in Purkinje cells, activating DLK causes rapid death and activating LZK leads to slow degeneration. Each kinase induces JNK activation and caspase-mediated apoptosis independent of each other. Significantly, deleting CELF2, which regulates alternative splicing of Mkk7, strongly attenuates Purkinje cell degeneration induced by activation of LZK, but not DLK. Thus, controlling the activity levels of DLK and LZK is critical for neuronal survival and health.

Astrocytes-derived soluble factor promotes the survival and dendritic development of rat cerebellar Purkinje cells

1998 ◽  
Vol 31 ◽  
pp. S310
Author(s):  
Shigeki Furuya ◽  
Toshihide Tabata ◽  
Junya Mitoma ◽  
Asami Makino ◽  
Masanobu Kano ◽  
...  
Keyword(s):  
Purkinje Cells ◽  
Soluble Factor ◽  
Dendritic Development ◽  
Cerebellar Purkinje Cells

scholarly journals Expression of the yes proto-oncogene in cerebellar Purkinje cells.

1989 ◽  
Vol 9 (10) ◽  
pp. 4545-4549 ◽  
Author(s):  
M Sudol ◽  
C F Kuo ◽  
L Shigemitsu ◽  
A Alvarez-Buylla
Keyword(s):  
In Situ Hybridization ◽  
Purkinje Cell ◽  
Gene Product ◽  
Purkinje Cells ◽  
Immunofluorescent Staining ◽  
Cell Degeneration ◽  
Functional Studies ◽  
Cerebellar Purkinje Cells ◽  
The Brain

To identify the kinds of cells in the brain that express the yes proto-oncogene, we examined chicken brains by using immunofluorescent staining and in situ hybridization. Both approaches showed that the highest level of the yes gene product was in cerebellar Purkinje cells. In addition, we analyzed Purkinje cell degeneration (pcd) mutant mice. The level of yes mRNA in cerebella of pcd mutants was four times lower than that found in cerebella of normal littermates. Our studies point to Purkinje cells as an attractive model for functional studies of the yes protein.

scholarly journals Spike burst–pause dynamics of Purkinje cells regulate sensorimotor adaptation

2018 ◽  
Author(s):  
Niceto R. Luque ◽  
Francisco Naveros ◽  
Richard R. Carrillo ◽  
Eduardo Ros ◽  
Angelo Arleo
Keyword(s):  
Purkinje Cell ◽  
Purkinje Cells ◽  
Vestibular Nuclei ◽  
Response Patterns ◽  
Spike Burst ◽  
Vor Adaptation ◽  
Cell Firing ◽  
Cerebellar Purkinje Cells ◽  
Oculomotor Adaptation

AbstractCerebellar Purkinje cells mediate accurate eye movement coordination. However, it remains unclear how oculomotor adaptation depends on the interplay between the characteristic Purkinje cell response patterns, namely tonic, bursting, and spike pauses. Here, a spiking cerebellar model assesses the role of Purkinje cell firing patterns in vestibular ocular reflex (VOR) adaptation. The model captures the cerebellar microcircuit properties and it incorporates spike-based synaptic plasticity at multiple cerebellar sites. A detailed Purkinje cell model reproduces the three spike-firing patterns that are shown to regulate the cerebellar output. Our results suggest that pauses following Purkinje complex spikes (bursts) encode transient disinhibition of targeted medial vestibular nuclei, critically gating the vestibular signals conveyed by mossy fibres. This gating mechanism accounts for early and coarse VOR acquisition, prior to the late reflex consolidation. In addition, properly timed and sized Purkinje cell bursts allow the ratio between long-term depression and potentiation (LTD/LTP) to be finely shaped at mossy fibre-medial vestibular nuclei synapses, which optimises VOR consolidation. Tonic Purkinje cell firing maintains the consolidated VOR through time. Importantly, pauses are crucial to facilitate VOR phase-reversal learning, by reshaping previously learnt synaptic weight distributions. Altogether, these results predict that Purkinje spike burst-pause dynamics are instrumental to VOR learning and reversal adaptation.Author SummaryCerebellar Purkinje cells regulate accurate eye movement coordination. However, it remains unclear how cerebellar-dependent oculomotor adaptation depends on the interplay between Purkinje cell characteristic response patterns: tonic, high-frequency bursting, and post-complex spike pauses. We explore the role of Purkinje spike burst-pause dynamics in VOR adaptation. A biophysical model of Purkinje cell is at the core of a spiking network model, which captures the cerebellar microcircuit properties and incorporates spike-based synaptic plasticity mechanisms at different cerebellar sites. We show that Purkinje spike burst-pause dynamics are critical for (1) gating the vestibular-motor response association during VOR acquisition; (2) mediating the LTD/LTP balance for VOR consolidation; (3) reshaping synaptic efficacy distributions for VOR phase-reversal adaptation; (4) explaining the reversal VOR gain discontinuities during sleeping.

scholarly journals Cerebellar neuronal dysfunction accompanies early motor symptoms in Spinocerebellar Ataxia Type 3 and is partially alleviated upon chronic citalopram treatment

2021 ◽  
Author(s):  
KJ Palarz ◽  
A Neves-Carvalho ◽  
S Duarte-Silva ◽  
P Maciel ◽  
K Khodakhah
Keyword(s):  
Mouse Model ◽  
Purkinje Cell ◽  
Purkinje Cells ◽  
Spinocerebellar Ataxia ◽  
Cell Activity ◽  
Spinocerebellar Ataxia Type ◽  
Neuronal Dysfunction ◽  
Spinocerebellar Ataxia Type 3 ◽  
Cell Dysfunction ◽  
Type 3

ABSTRACTSpinocerebellar ataxia type 3 (SCA3) is an adult-onset, progressive ataxia with no current disease modifying treatments. SCA3 patients have mild degeneration of the cerebellum, a brain area involved in motor coordination and maintenance of balance, as well as of the brainstem, of the spinal cord and of other movement-related subcortical areas. However, both SCA3 patients and SCA3 mouse models present clinical symptoms before any gross pathology is detectable, which suggests neuronal dysfunction precedes neurodegeneration, and opens an opportunity for therapeutic intervention. Such observations also raise the question of what triggers these abnormal motor phenotypes. Purkinje cells are the major computational unit within the cerebellum and are responsible for facilitating coordinated movements. Abnormal Purkinje cell activity is sufficient to cause ataxia. In this study, we show that the CMVMJD135 mouse model of SCA3 has dysfunctional deep cerebellar nuclei and Purkinje cells. Both cell types have increased irregularity as measured by inter-spike interval coefficient of variation. Purkinje cell dysfunction is likely a combination of intrinsic and extrinsic (synaptic) dysfunction. Interestingly, Citalopram, a selective serotonin reuptake inhibitor previously shown to alleviate disease in CMVMJD135 mice, also improved cerebellar neuron function in the CMVMJD135 mouse model. Specifically, we found that Purkinje cell dysfunction when synaptic transmission is intact was alleviated with citalopram treatment, however, intrinsic Purkinje cell dysfunction was not alleviated. Altogether, our findings suggest that cerebellar neuronal dysfunction contributes to the onset of SCA3 motor dysfunction and that citalopram, while effective at alleviating the motor phenotype, does not restore Purkinje cell intrinsic activity in SCA3. A novel therapeutic approach that combines citalopram with another therapeutic that targets this intrinsic dysfunction in a complementary manner might further reduce disease burden in SCA3.

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