**3. The Tau gene and its alternative splicing**

a trimer that bind to U2 along with U5). U1 is released, U5 shifts from exon to intron, and U6 binds to the 5′ splicing site). Complex C (U4 is released, and U6/U2 catalyze a transesterification to induce the binding of the 5′-end intron to the complex A, forming an intron lariat. U5 bind to the exon 3′ splicing site, which is cleaved). Afterward, U2, U5, and U6 remain bound to the lariat forms and the 3′ site is cleaved, whereas exons are ligated by means of ATP hydrolysis.

Alternative splicing generally is a mean by which a gene may generate a variety of messen‐ ger RNAs (mRNAs) with biological significance, that is coding for a protein. It has been estimated that at least 90% of all expressed genes are subjected to alternative splicing.

It has been identified at least six ways to generate alternative splicing: (a) exon exclusion or inclusion, (b) selecting one or more exons, (c) and (d) competition for the splicing site at a defined exon either in the 5′ or 3′ region, (e) retaining an intron, (f) multiple promoters, (g) multiple poly-A sites [12] (**Figure 2**). Exon and intron sequences may regulate the splicing site

**Figure 1.** RNA splicing. Exon 1 flanked on its 3′ end by the GU sequence and exon 2 on its 5′ end by AG, with both target sites for the ribonucleoproteins and the assembled spliceosome complex. The spliceosome will cut the intron in the consensus sequences and will enable the joining of the exons, generating a mature RNA. Scheme taken from [49].

**Figure 2.** Forms of alternative splicing. (**A**) Exclusion or inclusion of exons, (**B**) selection of one or more exons, (**C**) in‐ tron retention, (**D**) competencies by the site of splicing in a particular exon in the region 5′ or 3′, (**E**) multiple promot‐

Lariat forms are degraded, and the snRNP are recycled (**Figure 1**).

through enhancer or silencer sequences.

6 Update on Dementia

ers, (**F**) multiple poly-A sites.

Tau is a cytoskeleton protein involved in neuron morphology and polarity. It posses the ability to bind to microtubules in order to provide stability, and it maintains the neuron phenotype at the axon level [13].

It has been determined that Tau is located at the axon hillock, the axon and the growth cone, as its mRNA is transported to its translation site by a protein complex involving kinesin-3 as transporter and the HuD protein as mRNA stabilizer [14–16]. This is possible because Tau mRNA posses in its 3′-UTR region a uracil-rich axon localization sequence [14, 17].

Tau protein is mainly constituted by two domains: the N-terminal whose function is to interact with the plasma membrane [18] and the C-terminal domain, in which the microtubule-binding region is coded [19].

The human Tau gene is located at chromosome 17 [20], it is formed by 16 exons, and it has a promoter region that confers it with neuron specificity [21].

This gene is transcribed into three RNAs of 2, 6, and 9 kb, which are differentially expressed in the central nervous system, depending on their maturity state and the neuron type [18]. Six Tau mRNA isoforms have been identified as consequence of alternative splicing, five of them in the adult central nervous system and one fetal isoform. These messenger RNAs code six proteins ranging from 352 to 441 amino acids (aa). The fetal isoform (352 aa) does not contain the exons 2, 3, and 10. The adult form of 383 aa lacks exons 2 and 3; however, it includes exon 10. The 381 aa isoform include exon 2 but not 10. The 412 aa isoform includes both exons 2 and 10; the 410 aa isoform includes exons 2 and 3, but not 10. The 441 aa isoform includes exons 2, 3, and 10 [22] (**Figure 3**).

**Figure 3.** Tau isoforms, showing the different Tau proteins from the alternative splicing of exons 2, 3 and 10. Scheme taken from [49].

Tau alternative splicing occurs in exons 2, 3, and 10 and its form is of the (a) type that corre‐ sponds to exon exclusion or inclusion.

The studies conducted on Tau alternative splicing have been comprehensive, and most of them have been focused on exon 10.

Exon 10 displays a splicing pattern of inclusion and it is not present on fetal neurons. It is influenced by exon 9, which promotes its inclusion [18]. Exon 10 codes the second region of the (R) (KXGS) repeats in Tau. Alternative splicing generates Tau isoforms with 3 or 4 repeats that bind to microtubules. In mature brains, the level of 3R and 4R is similar. Exon 10 disruption is able and enough to cause neuron degeneration or taupathies [23].

Exon 10 is flanked by a long 13.6-kb intron and a short 3.8-kb intron, possessing a weak 5′ splicing site, which is similar to that in 3′. This would allow the inclusion or not of exon 10 in order to generate proteins with or without it [24, 25].

Exon 2 alternative splicing has been less studied. However, the studies conducted in our laboratory show that when PC12 cells (rat pheochromocytoma) cultures are exposed to the β1 → 42 amyloid peptide; alternative splicing of exons 2 and 3 is affected, as immature forms of Tau mRNA are transcribed in mature PC12 cells (phenotype differentiated into neuron). We observed that processes in these cells begin to retract. In spite the mechanism is still unknown, the effect produced in these cells indicate that immature Tau forms cease to stabilize micro‐ tubules in these cells processes [26] (**Figure 4**). The inclusion of exons 2 and 3 promotes the shift from immature Tau forms to their mature counterparts, stabilizing microtubules.

**Figure 4.** Tau exons 1–9 (modified from [50]), the primers Ex1 and Ex5A, amplify from exons 1 to 5. The exons 2 and 3 are amplified by (Ex1/Ex5A), the exon 6 is amplified by (Ex5/Ex9, Ex6S/Ex6AS) and exon 8 by (Ex7/Ex9). (**A**) Untreated undifferentiated cells and (**B**) NGF-induced differentiated PC12 cells; and from PC12 cells exposed to Aβ(1–42) peptide in (**C**) undifferentiated cells and (**D**) NGF-induced differentiated cells. Differentiation inhibits fetal tau expression. Aβ exposure promotes exclusion of exons 2/3 in undifferentiated and differentiated cells, and exclusion of exon 6 in undif‐ ferentiated cells. Scheme taken from [26].

It has been demonstrated that immature Tau forms are similar to those found on PHF [27], suggesting that exclusion of exons 2 and 3 induced by amyloid peptide in AD may destabilize neurites, and thus, the cells would lose their polarity.

Currently, splicing regulation has been studied with microRNAs (miRNAs), which are regulators of genetic expression [28]. miRNAs are short RNA molecules that bind to transcripts in order to repress and regulate expression. A miRNA deregulation was found on hippocam‐ pus and the prefrontal cortex. It was ascertained that miR-132-3-p was the most affected in this disease. miR-132-3-p downregulation in neurons was inversely proportional to the occurrence of hyperphosphorylated Tau [29], and it is linked to the splicing of exon 10 [30].

The inclusion of exon 10 is inhibited by the constitutive factors ASF/SF2, SRp55, SRp75, and SWAP [31].

Exon 2 regulation has been determined by inclusion and exclusion of exons 2 and 3, and it has been determined that exon 3 never appears without exon 2 [32].
