CAPITULO III EVOLUCIÓN NORMATIVA DE LA
3.2 LA LEY 44/1978, DEL IMPUESTO SOBRE LA RENTA DE LAS PERSONAS FÍSICAS
The aggregation of proteins in neurodegenerative disease contrasts with the functional aggregation of Sup35 in yeast, but also TIA1 in mammalian stress granules. These cytoplasmic foci are heavily involved in the processing and turn-over of mRNA. RNA binding proteins such as those we see aggregating in FTLD-FUS have both nuclear and cytoplasmic functions related to mRNA. Nuclear functions encompass regulating mRNA maturation including; RNA helicase activity, RNA polymerase elongation, splicing, and nuclear export (Heyd and Lynch, 2011). In the cytoplasm these proteins can regulate RNA transport, translation or silencing, as well as degradation (Liu-Yesucevitz et al., 2011). Much of this cytoplasmic regulative activity occurs at distinct macromolecular sites assembled by protein-protein interactions through glycine rich domains and Q/N rich regions, and protein-mRNA interactions through RNA recognition motifs (RRM) (Krichevsky and Kosik, 2001). Importantly, different macromolecular granules are
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assembled to undertake different functions. Visualizing proteins associated with these different functions allows microscopic delineation of these separate granules.Stress granules can be visualized with antibodies directed to TIA-1, whose RRM will recognize uracil rich 30-37 nucleotide bipartite motifs (Lopez de Silanes et al., 2005), or TIA1 cytotoxic granule-associated RNA binding protein-like 1 (TIAR) that will recognize mRNA with 28-32 long stem loops (Kim et al., 2007). The HuR proteins will bind mRNA uracil rich 17-20 nucleotide long sequences (Lopez de Silanes et al., 2004), and Ras-GAP associated endoribonucelase (G3BP), which is important in the formation of stress granules, cleaves mRNA between CA dinucleotides (Tourriere et al., 2001). The reaction cascade that leads to stress granule formation is complex but can be best interpreted through the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α).
Cellular stress prompts the phosphorylation of eIF2α at serine 51 by stress kinases (PKR, HRI, PERK or GCN2), which inhibits the translation complex containing
tRNAimet.(Kedersha et al., 1999). Capped mRNA remains bound to the pre-initiation complex and forms a nucleus for aggregation through TIA-1 and other protein-protein interactions. Stress granules are initially small and punctate but will increase in size as the RNA binding proteins listed above begin to coalesce and aggregate through their glycine-rich domains and Q/N rich regions. The vital nature of stress granules in the survival response is highlighted by increase apoptosis upon stress after knockdown of TIA-1 or inhibition of eIF2α phosphorylation (Phillips et al., 2004, Jiang et al., 2003).
However, this response has evolved to cope with acute transient stress, and the consequence of long term persistence of stress granules has not been explored. Stress granules dissociate relatively rapidly (1-3 hours) once the stress has been removed (Kedersha et al., 1999), once again through the phosphorylation status of eIF2α (Moreno et al., 2012). Pharmacological agents allow the manipulation of stress granules by altering the processes outlined here. Cycloheximide and emetine inhibit their formation by interrupting protein elongation whilst maintaining polysomes, thus preventing free
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mRNA accumulating in the cytoplasm and therefore stress granule nucleation (Kedersha and Anderson, 2007). Conversely, puromycin causes premature chain termination within the ribosome, and therefore disassembly of the polysome.Whilst sequestration of basal mRNA is the remit of stress granules, their subsequent degradation is mediated by a separate but related granule called P-bodies. These can be visualized by a separate group of markers including; mRNA-decapping enzyme 1a (DCP1a), the Sm and SM-like (LSm 1-7) proteins (Ingelfinger et al., 2002). Given their highly related function it is not unexpected that some components of one can be found within the other and moreover they are often found adjacent to one another (Parker and Sheth, 2007). The vast majority of P-body constituents are geared towards mRNA repression, interference and non-sense mediated decay. Examples include, mRNA
decapping machinery, activators of decapping and the 5’ to 3’ exonuclease Xrn1p (Cougot et al., 2004, Ingelfinger et al., 2002).
Both stress granules and P-bodies are not free floating entities but rather they are closely associated to the cytoskeleton and require microtubule alteration to function properly (Kwon et al., 2007). Histone deacetylase 6 (HDAC6) deacetylates tubulin to reduce cellular motility and is vital to the formation of stress granules. This enzyme also provides an interesting link with tau, because low activity is associated with tau accumulation, a hallmark of neurodegeneration (Cook et al., 2012). Furthermore, the dynein motor has been shown to tether stress granules to the cytoskeleton and aid the coalescence of small granules (Tsai et al., 2009).
The discovery that the cytoplasmic inclusions of FTLD-FUS contain stress granule markers (Dormann et al., 2010) has led to a flurry of research around this theme (Bosco et al., 2010, Gal et al., 2011). Some authors hypothesized that stress granule formation may be a precursor to the more sinister cytoplasmic aggregates (Dormann et al., 2010).
Yet these proteins are known to easily disperse and return to solution (Kedersha et al.,
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2000). Despite this, stress granule markers such as TIA-1 co-localize with thecharacteristic neuronal aggregates of Alzheimers disease, ALS, FTLDP-17, FTLD-TDP, and FTLD-FUS (Liu-Yesucevitz et al., 2010) suggesting there may be a link between the two phenomena. However, when considering human pathological tissue it is worth
remembering that any observation is of the absolute final time point of death. It is possible that as the aggregate grew it absorbed other non-membranous structures like stress granules as it dominated the cytoplasm. Evidence to the contrary comes from animal models which aim to emulate decades of disease in a matter of weeks. These animals also show stress granule markers within their induced pathology (Vanderweyde et al., 2012).
All of the widely accepted pathological markers of FTLD-FUS have been found to re-localise to stress granules or P-bodies when subjected to cellular stress (Liu-Yesucevitz et al., 2010, Blechingberg et al., 2012b, Chang and Tarn, 2009). However, re-localisation to the cytoplasm and arrangement into foci does not cause these proteins to become highly insoluble. This is crucial since highly insoluble protein is a core feature of aggregated disease protein.