Tumor Protein D52 (TPD52)
Synonyms
Historical Background
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.
Diagrammatic representations of (a) TPD52 gene structure and (b) domain organization of two major TPD52 isoforms. (a) Exon-intron structure of the TPD52 gene at human chromosome 8q21.13, not drawn to scale. Coding sequences are shown in color (rust, orange, green, red), whereas 5′- and 3′-untranslated regions are shown in gray. Lengths of the coding regions of each exon (in bp) are indicated above the diagram, whereas intron lengths (in kb) are shown below the diagram. The TPD52 gene includes two alternative first exons 1α and 1β. Exon 1α (gray and rust) is included in TPD52 transcripts in a tissue-ubiquitous fashion, whereas the inclusion of exon 1β (gray and orange) is tissue-restricted. Exons 5 and 6 (red) are alternatively spliced such that either or both can be included, although both exons 5 and 6 are predicted to be absent from ubiquitously expressed TPD52 transcripts. Exon 6 encodes a putative 14-3-3 binding site. (b) Sequence features of the ubiquitously expressed TPD52 isoform and the prostate-specific isoform PC-1 or PrLZ, with their associated UniProt identifiers shown in brackets. Numbers above each representation denote amino acid (aa) coordinates of the different N-terminal domains, the shared coiled-coil motif (in blue), the shared D52 motif (in pink), and isoform lengths (at right). The shorter TPD52 N-terminus encoded by exon 1α is shown in rust, whereas the longer PC-1/PrLZ N-terminus encoded by exon 1β is shown in orange. Positions of the shared N- and C-terminally located PEST sequences are shown as solid lines below each representation. The scale bar indicates 10 amino acid (aa) residues
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).
TPD52 Associations with Molecular Subtypes of Breast and Prostate Cancer
TPD52 amplification and overexpression occur in significant proportions of breast cancers and have been associated with poorer patient outcomes in a number of studies, and TPD52 overexpression occurs in the majority of prostate cancers (reviewed by Byrne et al. 2014). Recent studies have provided additional information about the breast and prostate cancer molecular subtypes that are associated with TPD52 expression. Integration of genomic and transcriptomic data from a large number of breast cancer cases has produced the so-called METABRIC molecular classification system with ten integrative breast cancer clusters (reviewed by Dawson et al. 2013). A subsequent large-scale analysis of the functional genomics landscape of breast cancer identified TPD52 as being preferentially essential for the survival of breast cancer cell lines corresponding to integrative cluster 1 (Marcotte et al. 2016). Integrative cluster 1 tumors are luminal B-like and frequently estrogen receptor positive and also tend to feature chromosome 17q23 amplification and GATA3 mutations (Dawson et al. 2013). The finding that TPD52 may be an essential gene for integrative cluster 1 breast cancers is broadly consistent with previous studies identifying TPD52 amplification and/or elevated transcript levels in luminal B breast cancer (reviewed in Byrne et al. 2014). Integration of genomic and transcriptomic data in prostate cancer has also identified five distinct prostate cancer clusters (Ross-Adams et al. 2015). Here, TPD52 was included in the gene signatures for 3/5 integrative cluster (iCluster) subtypes, including iCluster3 tumors which are characterized by chromosome 8q gains (Ross-Adams et al. 2015). The inclusion of TPD52 within these iCluster signatures reflected TPD52 being included in the 100 mRNA transcript probes with the highest levels of inter-tumor variability in primary prostate cancer tissues (Ross-Adams et al. 2015).
Regulation of TPD52 Expression by microRNAs
A new mechanism for the regulation of TPD52 transcript levels has emerged through studies identifying microRNA (miR) functions (reviewed in Byrne et al. 2014). A number of miRs have been found to target TPD52 transcript levels, and thus deregulated expression of tumor suppressive miRs may represent an additional mechanism that gives rise to increased TPD52 levels in cancer (Byrne et al. 2014).
microRNA (miR) regulation of TPD52 in human cancer cell lines
Study | microRNA (miR) | Cancer type | Rationale for analysis of TPD52 | Target sequence(s) in TPD52 described? | Confirmation studies performed |
---|---|---|---|---|---|
Li et al. (2016) | miR-34a a | Breast | Predicted target using TargetScan, Sanger miRNA database | 1 site in TPD52 3′-UTR | Reporter gene assays, miR-34a mimic transfection reduced TPD52 levels in MCF-7, MDA-MB-231 cells |
Han et al. (2015) | miR-218 | Prostate | Predicted target using TargetScan | 2 sites in TPD52 3′-UTR | Reporter gene assays, miR-218 transfection reduced TPD52 levels in PC3 cells |
Goto et al. (2014) | miR-224 | Predicted target using TargetScan, downregulated in miR-224-transfected PC3 and DU145 cells | 1 site in TPD52 3′-UTR | Reporter gene assays, miR-224 transfection reduced TPD52 levels in PC3, DU145 cells | |
Okato et al. (2016) | miR-320a | Predicted target using TargetScan, downregulated in miR-320a-transfected PC3 cells | Not shown | Not done | |
Donzelli et al. (2015) | miR-145-5p, miR-145-3p | Non-small cell lung | Predicted target using miRWalk | 2 sites in TPD52 3′-UTR | miR-145-5p transfection but not miR-145-3p transfection reduced TPD52 levels in H1299 cells |
Mataki et al. (2016) | Lung squamous cell | Predicted target using TargetScan, downregulated by miR-145-5p and miR-145-3p transfection into EBC-1 cells | Not shown | miR-145-3p transfection but not miR-145-5p transfection reduced TPD52 levels in EBC-1 cells | |
Kumamoto et al. (2016) | miR-218 | Predicted target using TargetScan, downregulated by miR-218 transfection into EBC-1 cells | 3 sites in TPD52 3′-UTR | Reporter gene assays, miR-218 transfection reduced TPD52, TPD52 levels in EBC-1, SK-MES-1 cells |
Summary of effects of reduced TPD52 expression reported in human cancer cell lines
Study | Experiment | Cancer type | Cell line(s) | Cell migration | Cell invasion | Cell proliferation |
---|---|---|---|---|---|---|
Kumamoto et al. (2016) | ↓TPD52 | Lung squamous cell | EBC-1, SK-MES-1 | ↓ | ↓ | ↔ |
Goto et al. (2014) | Prostate | PC3, DU145 | ||||
Li et al. (2016) | Breast | MCF7, MDA-MB-231 | NDa | |||
Donzelli et al. (2015) | Non-small cell lung | H1299 | ND | |||
Han et al. (2015) | Prostate | PC3 | ND | ↓ | ||
Shang et al. (2016) | ↓PC-1/PrLZ | C4–2 |
Cellular and Signaling Functions
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.
Proposed roles for TPD52 in regulating lipid metabolism in cancer cells. Triglycerides are synthesized from either endogenously synthesized fatty acids or from exogenous fatty acids obtained through the diet. Increased cellular triglyceride leads to increased lipid storage requirements. If these storage requirements are met, excess free fatty acids can be safely stored as triglyceride and then mobilized when required to enhance cancer cell survival, proliferation, and invasive capacity. However, if lipid storage requirements are not met, lipotoxicity may result. TPD52 has been shown to increase de novo free fatty acid synthesis, triglyceride synthesis, and lipid storage in 3T3 fibroblasts (Kamili et al. 2015). Upregulation of TPD52 levels in cancer cells (through increased gene copy number, reduced expression of tumor-suppressive miRs, and/or other mechanisms) may therefore allow cancer cells to benefit from increased lipogenesis and to avoid the harmful consequences of excess free fatty acids
Summary
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.
References
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