CRHSP-28 (rat);  CSPP28 (rabbit);  hD52 (human);  mD52 (mouse);  N8 (human);  PC-1 (human);  PrLZ (human);  R10 (quail);  Tpd52 (mouse)

Mammalian TPD52 sequences were first described in the mid-1990s, through a number of independent reports and experimental approaches. Publications from 1995 to 1996 identified TPD52 sequences through the detection of increased TPD52 transcript levels in human cancer tissue or cell lines, relative to nonmalignant controls. Orthologous rat, rabbit, or quail transcripts were identified as either encoding proteins that are phosphorylated in response to raised intracellular calcium levels or as a retroviral target gene, respectively. Early reports also identified paralogous human and mouse transcript sequences, and these reports, combined with genome sequencing, demonstrated that TPD52 is one gene within a four-member gene family (reviewed by Boutros et al. 2004; Byrne et al. 2014). While TPD52-like protein sequences are conserved both within and between species, they show limited similarity with sequences of other proteins.

Human TPD52 isoforms are around 200 residues in length (Fig. 1) and represent largely hydrophilic peptides that lack demonstrated catalytic activity or unambiguous predictors of specific function. TPD52 isoforms feature a coiled-coil motif of approximately 50 residues and N- and C-terminal PEST sequences (Fig. 1). Numerous alternatively spliced TPD52 transcripts have been identified. Notably, the use of alternative gene promoters results in the inclusion of alternative first exons encoding either a shorter or a more extended N-terminus (Fig. 1). Isoforms carrying the longer N-terminus are referred to as PC-1 or PrLZ, and their expression is predicted to be largely restricted to the prostate. Two internal coding exons may also be alternatively spliced (Fig. 1), including an exon encoding a consensus 14-3-3 binding site. In addition to the different isoform identifiers used by investigators, the description of TPD52 isoforms in the literature is further complicated by differences between the NCBI and UniProt nomenclature systems. For example, the ubiquitously expressed TPD52 isoform of 184 residues (Fig. 1) is referred to as both isoform 2 according to UniProt (P55327–2) and as isoform 3 according to NCBI (NP_005070).

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Despite the many isoforms of TPD52 that have been predicted to arise through alternative splicing, most alternative splicing events may not substantially alter protein molecular weights or isoform detection by TPD52 antisera that target shared epitopes. TPD52 transcripts also feature long 3′-untranslated regions (UTR’s), and thus the insertion or removal of short alternatively spliced exons might not produce obvious changes to transcript length. Many published laboratory analyses may therefore have examined more than one TPD52 transcript or isoform at the same time. For these reasons, the identifier TPD52 will be used generically in this chapter, whereas isoform-specific identifiers will be employed when discussing functions that have been ascribed to a particular isoform. In view of restrictions on the number of cited references, this chapter will also focus upon literature published from 2014. References to earlier publications can be found in previous review articles (Boutros et al. 2004; Shehata et al. 2008; Byrne et al. 2014).

A number of papers have been published since 2014 that ascribe particular functions to TPD52 isoforms. Many of these concern the prostate-specific isoform PC-1/PrLZ, with this different nomenclature being used by different research groups to describe isoforms bearing a longer N-terminus from an alternative promoter and first exon (Fig. 1). It was proposed that PC-1 exerts some oncogenic functions in LnCaP cells through inducing the expression of EphA3 (Wu et al. 2014). Another study from the same group reported that PC-1 increased the resistance of prostate cancer cells to rapamycin, by increasing the stability of eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) (Yu et al. 2015). Interactions between PC-1 and 4E-BP1 led to the inhibition of proteosomal regulation of 4E-BP1 (Yu et al. 2015). PC-1 was also proposed to cooperate with the E3 ligase C-terminus of heat shock cognate 70 interacting protein (CHIP) to reduce the stability of the androgen receptor (AR) in prostate cancer cell lines by promoting AR ubiquitination (Wang et al. 2016). This result contrasted with PC-1’s role in stabilizing 4E-BP1 (Yu et al. 2015).

The finding that PC-1 expression destabilized the AR in prostate cancer cell lines (Wang et al. 2016) also contrasted with previous studies showing that increased PC-1/PrLZ expression in LnCaP cells was associated with increased AR levels (reviewed by Byrne et al. 2014). A more recent study also indicated that PC-1 expression combined with IL-6 treatment of LnCaP cells resulted in increased AR levels (Moritz et al. 2016). This study included some functional comparison of TPD52 isoforms, where it was found that IL-6 treatment of LnCaP cells increased PC-1 levels but not those of TPD52 isoform 2 (referred to as TPD52 isoform 3, using NCBI nomenclature) (Moritz et al. 2016). PC-1 expression and IL-6 treatment both induced the neuroendocrine differentiation of LnCaP cells, a frequent characteristic of late-stage, endocrine-resistant prostate cancer (Moritz et al. 2016). Neuroendocrine differentiation of LnCaP cells was also accompanied by increased AR levels (Moritz et al. 2016).

TPD52 expression has been previously associated with increased radiosensitivity, as reviewed by Byrne et al. (2014). A number of studies have shown that increased TPD52 transcript levels are associated with compromised DNA repair capacity, and/or radiosensitivity, and that TPD52 knockdown improves DNA repair capacity. For example, in both mouse 3T3 fibroblasts and SK-BR-3 breast cancer cells, TPD52 functioned as a radiosensitizer, where TPD52 expression was associated with reduced ATM levels (Chen et al. 2013). TPD52 was found to bind ATM, with ATM-binding mapping to a TPD52 region shared by PC-1/PrLZ (Chen et al. 2013). In contrast to these results, Shang et al. (2016) reported that PC-1 depletion increased prostate cancer cell radiosensitivity and that reduced PC-1 levels were associated with reduced repair of double-strand breaks, reduced function of the ionizing radiation-induced G2/M checkpoint, and with increased autophagy. Future studies where the radiosensitizing functions of TPD52 (TPD52 isoform 2, P55327–2) and PC-1/PrLZ are directly compared may help to resolve these contrasting reports.

An additional function has been recently ascribed to TPD52, this being the regulation of cellular lipid storage (Kamili et al. 2015). Expression of TPD52 isoform 2 (P55327–2) in 3 T3 cells was associated with increased numbers of lipid storage organelles, or lipid droplets, per cell, and with increased lipid storage capacity in response to exogenous oleic acid treatment (Kamili et al. 2015). TPD52 detection also increased at lipid droplets in response to oleic acid supplementation (Kamili et al. 2015). TPD52 isoform 2 (P55327–2) was shown to bind to members of the perilipin protein family, a function that was not shared by the related protein TPD52L1 (Kamili et al. 2015). Interestingly, TPD52L1 expression also did not increase lipid droplet numbers or sizes in three T3 cells (Kamili et al. 2015). As lipid metabolism is deregulated in cancers, notably in breast and prostate cancers where TPD52 expression may be highest (http://www.cbioportal.org/), increased TPD52 expression may increase lipid storage capacity in order to match increased lipogenesis, to prevent lipotoxicity (Fig. 2).

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Whereas descriptions of TPD52 expression in cancer continue to emerge, typically through the application of unbiased or high-throughput techniques, fewer targeted studies examine TPD52 functions or indeed the functions of related family members (Byrne et al. 2014). The signaling functions of TPD52 therefore remain poorly understood. There have been some recent causes for optimism that this long-standing situation will improve in coming years. For example, TPD52 has been attracting research attention as a miR target, and a number of studies since 2014 have analyzed TPD52 functions as a result of TPD52 being targeted by different miRs (Table 1). While these studies have not yet provided a great deal of insight into TPD52’s signaling functions, the identities of genes that are deregulated in response to reduced TPD52 levels may provide some clues (Kumamoto et al. 2016). The discovery that TPD52 increases lipid storage in cells represents an unanticipated finding (Kamili et al. 2015), but again the cascades of events up- and downstream of TPD52 at the lipid droplet remain unknown. However the fact that TPD52 functions at the lipid droplet are likely to be related to perilipin protein function provides a molecular foundation from which to explore.

There are a number of challenges to be overcome if a more integrated understanding of TPD52 function is to emerge. Research on TPD52 remains limited and dispersed, and there tends to be a lack of integration between the results of some related published studies. Individual studies tend to be published as stand-alone analyses, with at times limited reference to or comparison with previous relevant results. Suggestions that PC-1/PrLZ represents an independent gene, as opposed to an alternatively spliced transcript from the TPD52 locus (Fig. 1), may have added to the difficulty of understanding an already fragmented body of literature. An example of this lack of integration is that in the field of prostate cancer, where TPD52 and PC-1/PrLZ have been best studied, very few studies have directly compared TPD52 and PC-1/PrLZ (Moritz et al. 2016). Common functions are likely to exist, given that PC-1/PrLZ and TPD52 are identical over the majority of their protein sequences (Fig. 1). While there are likely some isoform-specific roles for these proteins, it would seem more helpful to explore any isoform-specific functions from an understood platform of common functions.

In summary, TPD52 remains recognized as a frequently overexpressed protein in cancer, yet TPD52 signaling functions largely continue to elude the research community.