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  • The second class of proteins

    2021-09-18

    The second class of proteins necessary for maintaining gstp1 dynamics is chromatin modifiers, which are considered ‘writers’ and ‘erasers’ and are responsible for modifications of histone N-terminal tails. These post-translational modifications can have direct or indirect effects on chromatin structure. One of the most widely studied modifications is lysine acetylation, which mitigates the positive charge on the lysine residue (Haberland et al., 2009, Shahbazian and Grunstein, 2007, Tapias and Wang, 2017). This modification decreases association of the positively charged histone tail with the negatively charged DNA backbone making removal or movement of the nucleosome more likely. Lysine acetylation is usually enriched at actively transcribed genes as it allows for unobstructed access of promotor or enhancer DNA sequences to transcription factors and enhancers (Haberland et al., 2009, Tapias and Wang, 2017, Chuang et al., 2009, Peserico and Simone, 2011). Histone acetyltransferases (HATs) are the ‘writers’ that catalyze the addition of the acetyl groups and histone deacetylases (HDACs) are the counteracting ‘erasers’ (Tapias and Wang, 2017, Peserico and Simone, 2011). HDACs, in particular, have been widely studied as potential therapeutic targets in the nervous system (Haberland et al., 2009, Kazantsev and Thompson, 2008, Machado-Vieira et al., 2011). Class II HDACs were shown to be essential for learning and memory formation and synaptic plasticity in mice (Kim et al., 2012, Sando et al., 2012). Additionally, pharmacological inhibition of HDACs has been shown to be effective in mouse models of genetic disorders including Rubinstein-Taybi syndrome, Rett syndrome, and Friedreich's ataxia (Herman et al., 2006, Vecsey et al., 2007, Kavalali et al., 2011). Furthermore, general HDAC inhibitors (HDACi) like Valproate were FDA approved for the treatment of complex psychiatric disorders such as major depressive disorder, bipolar disorder, and epilepsy (Vasudev et al., 2012, Vigo and Baldessarini, 2009). Despite the importance of these enzymes for nervous system function, little is known about their interactions with other classes of chromatin associated proteins such as chromatin remodelers. We sought to determine the interaction between one chromatin remodeler, the epigenetic ‘reader’ Kis, and a class of chromatin modifiers, HDACs, in Drosophila. Our lab previously showed that ubiquitous Kis knockdown produces morphological, postural, and motor-function defects (Ghosh et al., 2014, Melicharek et al., 2010). Using these phenotypes, we performed a small-scale screen with a group of drugs that target epigenetic proteins. From this screen, we identified a number of HDAC inhibitors capable of suppressing the phenotypes associated with Kis knockdown. We then selected the top general HDAC inhibitor (suberoylanilide hydroxamic acid, SAHA) and top specific inhibitor (suberohydroxamic acid, SBHA) from the HDAC inhibitors we tested to evaluate further. Using the well-defined Drosophila neuromuscular junction (NMJ) as a model of glutamatergic synaptic function, we show both SAHA and SBHA alleviate defects in synaptic morphology, motor behavior, and neurotransmission associated with decreased Kis levels. These results provide further understanding into the complex interactions between epigenetic reader and eraser functions in vivo and lead us to hypothesize that Kis and HDACs may converge on a similar set of target genes in the nervous system.
    Materials and methods
    Results
    Discussion The results reported here indicate an antagonistic crosstalk between HDACs and the chromatin reader Kis in the larval Drosophila nervous system (Fig. 5). This is evidenced by the capacity of pharmacological inhibition of HDACs to alleviate or suppress synaptic morphology, neurotransmission, and behavioral defects in Kis knockdown larvae. In contrast, HDAC inhibitors were not able to rescue adult morphological and behavioral defects associated with ubiquitously decreased Kis levels. This is intriguing. Why would HDAC inhibition have a more significant effect on larval phenotypes compared to adult phenotypes? This might be explained by the diminished exposure to drug treatments during the 4–5days of metamorphosis during our study. Metamorphosis is a critical developmental milestone, which requires precise control of gene expression (Fisk and Thummel, 1995, Sullivan and Thummel, 2003, Thummel, 1995). While both larvae and adults are raised on food containing treatment, larvae have an increased exposure to the drugs as they have not yet entered into metamorphosis, a stage that sequesters them from the food and treatment. This may result in a diminished drug exposure once they eclose as adult animals. Indeed, although we observed trends towards suppression in adult animals treated with SAHA, none of these trends were significant. While in the larval stage, however, SAHA showed the strongest effect in our animals.