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1

Activity-dependent rearrangement of the neuronal cell nucleus

100%
Szczepankiewicz A. A., Walczak A., Parobczak K., Wilczynski G. M.
Acta Neurobiologiae Experimentalis
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2013
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tom 73
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nr Suppl.1
It is now firmly established that long-lasting synaptic plasticity involves dramatic changes in gene expression occurring under the influence of specific signaling pathways and transcription factors. Numerous studies have shown that DNA and histone epigenetic modifications play key roles in neuronal plasticity. Recent studies in non-neuronal cells, indicated the existence of epigenetic mechanism of yet another class, related to the nuclei structural remodeling and very poorly understood in neurons. Therefore, we decided to study the ultrastructure of the cell nuclei in the hippocampal dentate gyrus granule neurons upon seizures induced by kainic acid, an analog of glutamate. Under these conditions the granular neurons instead of degradation, undergo an intensive plasticity phenomena. We found that seizures led to rapid and dramatic enlargement and striking reorganization of internal component-structures of interchromatin granule clusters (IGCs) in granular cell’s nucleus. Moreover, unlike IGCs of control animals, the reorganized IGCs contained activated RNA polymerase II CTD phosphoepitopes. These observations may suggest involvement of IGC in activity-dependent transcription events in neurons.
2

RNA-binding proteins in dendritogenesis

100%
Perycz M., Parobczak K., Jaworski J.
Acta Neurobiologiae Experimentalis
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2009
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tom 69
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nr 3
Proper development of dendritic arbor is a precisely orchestrated process in which translation is shown to play pivotal role. RNA binding proteins such as ZBP1 (beta-actin zipcode binding protein 1), Staufen1 and Staufen2 bind to and transport their target mRNAs, enabling their local translation in dendrites and taking part in regulation of this process. ZBP1 knockdown with short interfering RNAs in developing rat hippocampal neurons in vitro, resulted in decrease in total number of dendrites and dendritic tree complexity, which was reversed both with siRNA-resistant ZBP1 rescue mutant and jasplakinolide treatment that prevents depolymerisation of actin cytoskeleton. In contrast, mature neurons with stable dendritic arbors, did not lose dendrites upon ZBP1 depletion, which suggests ZBP1 plays role in formation of new dendrites as opposed to maintenance of mature ones. We also show that both Staufen1 and Staufen2 knockdown in developing neurons results in impoverishment in dendritic arbors complexity and decrease in total number of dendrites. RNA-binding proteins can be subject to phosphorylation, which modulate their activity, and one possibility for a common regulator of their role in dendritogenesis would be Src kinase, which we show to phosphorylate ZBP1 and Staufen1. Taken together, presented data demonstrate that mRNA transport and possibly local protein synthesis play a prominent role in development of dendritic arbor. Ministry of Science and Higher Education Grant NN301314733.
3

Arc functional neighbourhood in the neuronal cell nucleus

76%
Parobczak K., Rutkowska-Wlodarczyk L., Szczepankiewicz A. A., Khanema-Jakobsen T., Schubert M., Bramham C. R., Wilczynski G. M.
Acta Neurobiologiae Experimentalis
|
2013
|
tom 73
|
nr Suppl.1
Arc protein is a versatile factor connecting memory formation, plasticity changes and neuronal activity. Through regulation of actin polymerization, Arc contributes to synapse expansion and may furthermore influence synapse strength via management of AMPA receptor turnover. The function of Arc in the neuronal cell nucleus is poorly understood. In this work we performed structural, functional and biochemical analysis to identify Arc`s nuclear interactome. Using confocal microscopy we investigated Arc`s functional neighborhood and found that it occupied internal parts of the nucleus, closely to hnRNPs. This observation were confirmed with electron microscopy, which demonstrated that Arc localizes mainly at the peripheral areas of chromatin. Furthermore, pull-down-based biochemical experiments suggested that Arc interacts with splicing machinery. Collectively, our data suggest that nuclear Arc is involved in the gene expression phenomena.
4

How neurons become giant-identification of proteins regulated by mTOR kinase in etiopathology of tuberous sclerosis

51%
Malik A., Swiech L., Perycz M., Urbanska M., Blazejczyk M., Wulf P., Parobczak K., Pietruszka P., Zarebska M., Rutowicz K., Jaworski J.
Acta Neurobiologiae Experimentalis
|
2009
|
tom 69
|
nr 3
mTOR is a serine-threonine protein kinase for several aspects of neuronal function. Increased mTOR signaling has been implicated in tuberous sclerosis (TS), a multiorgan disease affecting brain. TS is caused by mutations in genes encoding for hamartin and tuberin that lead to increase of the mTOR activity and subsequent abnormal cell growth and proliferation, leading to brain lesions containing giant cells. It is unclear how mTOR contributes to observed changes. Our aim was to identify downstream mTOR effectors important for the disease related abnormal cell growth. To model TS giant cells, rat cortical neurons cultured in vitro were transfected with short interfering RNA (siRNA) targeting tuberin that caused a Rapamycin-sensitive increase of neuron soma size. Next, we designed a siRNA library directed against 140 mRNAs encoding potential mTOR targets, selected based on published data. To select siRNAs decreasing soma size of enlarged cells, cortical neurons were contransfected with tuberin siRNA together with siRNAs from the library. Our screen revealed over 20 genes, whose expression downregulation reversed the giant-cell like phenotype and 6 siRNAs that further increased size of cells with tuberin knockdown. Among proteins contributing to abnormal neuron growth upon mTOR overactivation, we identifi ed those involved in actin cytoskeleton dynamics, vesicular transport and cellular signaling. This work has been fi nanced by PBZ-MNiI-2/1/2005 and PNRF-96- AI-1/07 grants.
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