Transmembrane prostatic acid phosphatase (TMPAP) delays cells in G1 phase of the cell cycle.

BACKGROUND: Prostate adenocarcinoma is the most common form of prostate cancer. We have previously shown in a murine model that prostatic acid phosphatase (PAP) deficiency leads to increased cell proliferation and development of prostate adenocarcinoma. The association between PAP and prostate cancer has been reported. Indeed, high PAP enzymatic activity is detected in the serum of patients with metastatic disease while its expression is reduced in prostate cancer tissue. However, the molecular mechanisms behind the onset of the disease remains poorly understood. We previously identified a novel transmembrane prostatic acid phosphatase (TMPAP) isoform, which interacts with snapin. TMPAP is expressed on the plasma membrane, as well as endosomal/lysosomal and exosomal membrane vesicles by means of a tyrosine-based lysosomal targeting motif (Yxxϕ). METHODS: We used stable overexpression of the secreted isoform (SPAP) and TMPAP in LNCaP cells, live cell imaging, microarray and qRT-PCR analyses, and fluid phase uptake of HRP and transferrin. RESULTS: Our results indicate that the stable overexpression of TMPAP, but not SPAP in LNCaP cells reduces cell growth while increasing endo/exocytosis and cell size. Specifically, cells overexpressing TMPAP accumulate in the G1 phase of the cell cycle, and show altered gene expression profile. CONCLUSIONS: Our data suggests that TMPAP may function as a non-canonical tumor suppressor by delaying cell growth in G1 phase of the cell cycle.

Prostatic acid phosphatase is the main acid phosphatase with 5'-ectonucleotidase activity in the male mouse saliva and regulates salivation.

We have previously shown that in addition to the well-known secreted isoform of prostatic acid phosphatase (sPAP), a transmembrane isoform exists (TMPAP) that interacts with snapin (a SNARE-associated protein) and regulates the endo-/exocytic pathways. We have also shown that PAP has 5'-ectonucleotidase and thiamine monophosphatase activity and elicits antinociceptive effects in mouse models of chronic inflammatory and neuropathic pain. Therefore, to determine the physiological role of PAP in a typical exocrine organ, we studied the submandibular salivary gland (SMG) of PAP(-/-) and wild-type C57BL/6J mice by microarray analyses, microRNA sequencing, activity tests, immunohistochemistry, and biochemical and physiological analyses of saliva. We show that PAP is the main acid phosphatase in the wild-type male mouse saliva, accounting for 50% of the total acid phosphatase activity, and that it is expressed only in the granular convoluted tubules of the SMGs, where it is the only 5'-ectonucleotidase. The lack of PAP in male PAP(-/-) mice was associated with a significant increase in the salivation volume under secretagogue stimulation, overexpression of genes related to cell proliferation (Mki67, Aurkb, Birc5) and immune response (Irf7, Cxcl9, Ccl3, Fpr2), and upregulation of miR-146a in SMGs. An increased and sustained acinar cell proliferation was detected without signs of glandular hyperplasia. Our results indicate that in PAP(-/-) mice, SMG homeostasis is maintained by an innate immune response. Additionally, we suggest that in male mice, PAP via its 5'-ectonucleotidase activity and production of adenosine can elicit analgesic effects when animals lick their wounds.

Prostatic acid phosphatase is not a prostate specific target.

Prostatic acid phosphatase (PAP) is currently evaluated as a target for vaccine immunotherapy of prostate cancer. This is based on the previous knowledge about secretory PAP and its high prostatic expression. We describe a novel PAP spliced variant mRNA encoding a type I transmembrane (TM) protein with the extracellular NH(2)-terminal phosphatase activity and the COOH-terminal lysosomal targeting signal (YxxPhi). TM-PAP is widely expressed in nonprostatic tissues like brain, kidney, liver, lung, muscle, placenta, salivary gland, spleen, thyroid, and thymus. TM-PAP is also expressed in fibroblast, Schwann, and LNCaP cells, but not in PC-3 cells. In well-differentiated human prostate cancer tissue specimens, the expression of secretory PAP, but not TM-PAP, is significantly decreased. TM-PAP is localized in the plasma membrane-endosomal-lysosomal pathway and is colocalized with the lipid raft marker flotillin-1. No cytosolic PAP is detected. We conclude that the wide expression of TM-PAP in, for instance, neuronal and muscle tissues must be taken into account in the design of PAP-based immunotherapy approaches.

Structure of Acid phosphatases.

Acid phosphatases are enzymes that have been studied extensively due to the fact that their dysregulation is associated with pathophysiological conditions. This characteristic has been exploited for the development of diagnostic and therapeutic methods. As an example, prostatic acid phosphatase was the first marker for metastatic prostate cancer diagnosis and the dysregulation of tartrate resistant acid phosphatase is associated with abnormal bone resorption linked to osteoporosis. The pioneering crystallization studies on prostatic acid phosphatase and mammalian tartrate-resistant acid phosphatase conformed significant milestones towards the elucidation of the mechanisms followed by these enzymes (Schneider et al., EMBO J 12:2609-2615, 1993). Acid phosphatases are also found in nonmammalian species such as bacteria, fungi, parasites, and plants, and most of them share structural similarities with mammalian acid phosphatase enzymes. Acid phosphatase (EC 3.1.3.2) enzymes catalyze the hydrolysis of phosphate monoesters following the general equation. Phosphate monoester + H2O -->/<-- alcohol + phosphate. The general classification "acid phosphatase" relies only on the optimum acidic pH for the enzymatic activity in assay conditions using non-physiological substrates. These enzymes accept a wide range of substrates in vitro, ranging from small organic molecules to phosphoproteins, constituting a heterogeneous group of enzymes from the structural point of view. These structural differences account for the divergence in cofactor dependences and behavior against substrates, inhibitors, and activators. In this group only the tartrate-resistant acid phosphatase is a metallo-enzyme whereas the other members do not require metal-ion binding for their catalytic activity. In addition, tartrate-resistant acid phosphatase and erythrocytic acid phosphatase are not inhibited by L-(+)-tartrate ion while the prostatic acid phosphatase is tartrate-sensitive. This is an important difference that can be exploited in in vitro assays to differentiate between different kinds of phosphatase activity. The search for more sensitive and specific methods of detection in clinical laboratory applications led to the development of radioimmunoassays (RIA) for determination of prostatic acid phosphatase in serum. These methods permit the direct quantification of the enzyme regardless of its activity status. Therefore, an independent structural classification exists that helps to group these enzymes according to their structural features and mechanisms. Based on this we can distinguish the histidine acid phosphatases (Van Etten, Ann N Y Acad Sci 390:27-51, 1982), the low molecular weight protein tyrosine acid phosphatases and the metal-ion dependent phosphatases. A note of caution is worthwhile mentioning here. The nomenclature of acid phosphatases has not been particularly easy for those new to the subject. Unfortunately, the acronym PAP is very common in the literature about purple acid phosphatases and prostatic acid phosphatase. In addition, LPAP is the acronym chosen to refer to the lysophosphatidic acid phosphatase which is a different enzyme. It is important to bear in mind this distinction while reviewing the literature to avoid confusion.

Transmembrane prostatic acid phosphatase (TMPAP) interacts with snapin and deficient mice develop prostate adenocarcinoma.

The molecular mechanisms underlying prostate carcinogenesis are poorly understood. Prostatic acid phosphatase (PAP), a prostatic epithelial secretion marker, has been linked to prostate cancer since the 1930's. However, the contribution of PAP to the disease remains controversial. We have previously cloned and described two isoforms of this protein, a secretory (sPAP) and a transmembrane type-I (TMPAP). The goal in this work was to understand the physiological function of TMPAP in the prostate. We conducted histological, ultra-structural and genome-wide analyses of the prostate of our PAP-deficient mouse model (PAP(-/-)) with C57BL/6J background. The PAP(-/-) mouse prostate showed the development of slow-growing non-metastatic prostate adenocarcinoma. In order to find out the mechanism behind, we identified PAP-interacting proteins byyeast two-hybrid assays and a clear result was obtained for the interaction of PAP with snapin, a SNARE-associated protein which binds Snap25 facilitating the vesicular membrane fusion process. We confirmed this interaction by co-localization studies in TMPAP-transfected LNCaP cells (TMPAP/LNCaP cells) and in vivo FRET analyses in transient transfected LNCaP cells. The differential gene expression analyses revealed the dysregulation of the same genes known to be related to synaptic vesicular traffic. Both TMPAP and snapin were detected in isolated exosomes. Our results suggest that TMPAP is involved in endo-/exocytosis and disturbed vesicular traffic is a hallmark of prostate adenocarcinoma.