A common event in cellular signal transduction pathways is the phosphorylation of proteins on tyrosine residues. Tyrosine phosphorylation is reversible. The forward reaction is mediated by protein tyrosine kinases. By contrast, the reverse reaction is performed by protein tyrosine phosphatases (PTP). The PTP family consists of 107 genes whose protein products are diverse in form and specificity (Alonso et al. 2004). PTPN3 and PTPN4 constitute two members of this family that were initially identified by PCR amplification using primers specific to conserved regions of the catalytic domain of canonical PTP. PTPN3 was initially cloned from a HeLa cell cDNA library, whereas PTPN4 was cloned from a megakaryoblastic cell line (Gu et al. 1991; Yang and Tonks 1991). PTPN3 and PTPN4 are 50% identical and 67% homologous at the amino acid level.
Structure and Expression Studies
PTPN3 and PTPN4 are cytosolic proteins that localize to the plasma membrane. Structurally, each PTP comprises of an amino-terminal FERM (band 4.1, ezrin, radixin, and moesin) domain, a central PDZ (PSD-95, Dlg, ZO-1) domain, and a carboxy-terminal PTP domain. FERM and PDZ domains are protein–protein interaction domains that commonly bind the cytosolic tail of transmembrane proteins and can also interact with the phospholipid, phosphatidylinositol 4,5 bisphosphate, PI(4,5)P2. PTPN3 and PTPN4 each require the FERM domain for plasma membrane association (Gjorloff-Wingren et al. 2000). The PDZ domain blocks catalytic activity of PTPN3 and PTPN4, and this autoinhibition is relieved upon PDZ domain ligand binding (Chen et al. 2014; Maisonneuve et al. 2016).
PTPN3 and PTPN4 are expressed ubiquitously (Pilecka et al. 2007; Bauler et al. 2008). Both PTP are expressed at high levels in the thalamus. In addition, PTPN4 is highly expressed in the testes. A recent comprehensive examination of the PTP transcriptome in the murine immune system reported expression of PTPN3 in all immune cell types examined, with elevated transcript levels in immature dendritic cells (DC), NKT cells, activated CD4 cells, and intestinal intraepithelial CD8 cells (Arimura and Yagi 2010). The same study reported elevated transcript levels for PTPN4 specifically in immature DC, NK cells, and B cells and also found that PTPN4 transcript levels in DC were increased following LPS stimulation. PTPN3 mutations and altered levels of PTPN3 expression have been associated with multiple different types of cancer. PTPN3 mutations described in colorectal cancer cell lines were not detected in a subsequent examination of primary colorectal cancer samples (Wang et al. 2004; Wood et al. 2007). Recently, miR-183 has been demonstrated to regulate PTPN4 expression (Zhu et al. 2016).
Enzymatic Substrates and Protein–Protein Interactions
PTPN3 has been shown to associate with and dephosphorylate several different target proteins. In COS-7 cells, PTPN3 was demonstrated to associate with and dephosphorylate cotransfected T-cell antigen receptor (TCR) ζ chain (Sozio et al. 2004). Furthermore, in a T cell line, overexpressed PTPN3 was shown to inhibit TCR-induced activation of the promoter of the gene for the cytokine interleukin-2 (Han et al. 2000). PTPN3 interacts with phosphorylated growth hormone receptor in vitro, and overexpression of PTPN3 in cell lines modulates signaling from this receptor (Pasquali et al. 2003; Pilecka et al. 2007). Expression of PTPN3 in fibroblasts can inhibit cellular growth via dephosphorylation of p97/valosin-containing protein, an established regulator of the cell cycle (Zhang et al. 1999). In contrast to this, PTPN3 binding to and dephosphorylation of p38γ mitogen-activated protein kinase promotes cellular proliferation (Hou et al. 2010). Oncogenic human papillomavirus E6 protein binds PTPN3 and targets it for degradation by the proteasome (Jing et al. 2007). PTPN3 has also been shown to dephosphorylate epidermal growth factor receptor and estrogen receptor and to interact with tumor necrosis factor alpha-convertase, vitamin D receptor, the cardiac sodium channel Na v1.5, and 14-3-3β protein (Li et al. 2015; Ma et al. 2015; Suresh et al. 2014; Zhang et al. 1997; Jespersen et al. 2006).
PTPN4 has also been characterized as having multiple binding partners. Like PTPN3, PTPN4 has been demonstrated to bind and dephosphorylate TCRζ in vitro, and when overexpressed in a T cell line, PTPN4 inhibits TCR signaling (Young et al. 2008). The PDZ domain of PTPN4 also binds p38γ, which relieves the catalytic autoinhibition of PTPN4 to prevent cell death. In addition, dephosphorylation of TRAM by PTPN4 inhibits TRIF-dependent signaling in response to TLR4 stimulation (Huai et al. 2015). PTPN4 has been shown to bind the δ- and ε-subunits of the glutamate receptor, signaling from which is required for learning and coordination (Hironaka et al. 2000). Interaction of PTPN4 with attenuated rabies virus glycoprotein and CrkI are associated with apoptotic death of infected cells and negative regulation of cell proliferation, respectively (Prehaud et al. 2010; Zhou et al. 2013).
Characterization of PTPN3- and PTPN4-Deficient Organisms
So as to understand the importance of PTPN3 and PTPN4 in normal physiological processes, organisms deficient in expression of these PTP were generated. Drosophila has a single homologue of PTPN3 and PTPN4, termed PTPMEG. Flies that lack expression of full-length PTPMEG were observed to become trapped alive in their food, suggesting a neuronal defect. Subsequent examination of neural connectivity patterns in the brain revealed roles for PTPMEG in the establishment and maintenance of axon projections in Drosophila (Whited et al. 2007).
PTPN3-deficient mice have been shown to be grossly normal, although subtle phenotypes have been observed. One group reported male PTPN3-deficient mice have a higher body mass and reduced working memory compared to wild-type littermates, whereas female PTPN3-deficient mice exhibit motor learning deficiencies (Pilecka et al. 2007; Patrignani et al. 2008). These mild phenotypes have not been replicated in PTPN3-deficient mice generated by an independent group (Bauler et al. 2008). A function for PTPN3 in TCR signal transduction has not been observed in primary murine T cells, contrary to the aforementioned evidence generated in cell lines (Bauler et al. 2007). Potential subtle roles for PTPN3 in spontaneous pain perception and the positive regulation of LPS-induced cytokine release in mice have been recently reported (Patrignani et al. 2010).
PTPN4-deficient mice have also been shown to be largely normal. Two independent groups demonstrated that T-cell function remained intact in PTPN4-deficient mice (Bauler et al. 2008; Young et al. 2008). One other group reported that motor learning and cerebellar synaptic plasticity were impaired in PTPN4-deficient mice, although altered motor learning was not confirmed by an independent group (Kina et al. 2007; Bauler et al. 2008). Recently, human disease has been reported due to haploinsufficiency of PTPN4 in a pair of identical twins, resulting in a neurodevelopmental disorder that resembles Rett syndrome (Williamson et al. 2015).
Owing to the high degree of homology between PTPN3 and PTPN4 and broad expression patterns of both PTP, it remained possible that loss of either protein was functionally compensated for by the other, thus accounting for the lack of discernible phenotypes in single PTP-deficient mice. However, double PTPN3-PTPN4-deficient mice are also indistinguishable from wild-type mice when considering T-cell function, body mass, and motor learning (Bauler et al. 2008).
PTPN3 and PTPN4 are homologous, ubiquitously expressed PTP that have been implicated as regulators of diverse cellular signaling cascades through interactions with a variety of protein substrates. However, definitive physiological roles for PTPN3 and PTPN4 have not yet been described in higher organisms. Further study of these PTP may reveal nonredundant functions that can be readily demonstrated in gene-targeted mice. Given the high degree of similarity and between PTPN3 and PTPN4, both in terms of structure and expression, it is likely that such functions will be revealed only in double PTP-deficient animals.
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