16 research outputs found
TMPAP co-localize and interact with snapin in the cell lamellipodia.
<p>A, co-localization (yellow) of TMPAP (green) with snapin (red) was observed in the vesicles and lamellipodia of the TMPAP/LNCaP cells. Arrows mark the co-localization points in the upper panel (scale bar: 20 µm). Lower panel (scale bar: 3 µm) showing the lamellipodia region, amplification of the area marked with a box in the upper panel (left). B, intensification of donor (TMPAP-GFP) fluorescence in LNCaP cells was observed after acceptor (snapin-DsRed) photobleaching which confirms FRET between two molecules (Scale bar: 2 µm).</p
Significant ontological terms obtained with GoMiner software from two-color microarrays experiments.
<p><b>GO ID</b>: gene ontology ID accession number. <b><i>P</i></b><b>-value</b>: <i>P</i>-value for the number of changed genes in the input list, significant <i>P</i>-value <0.05. <b>Term</b>: associated ontological term. <b>Rows in bold</b>: relevant ontological groups for vesicular transport. Mice in microarray experiment per group, <i>n = </i>3.</p
Proliferation of DLP cells is increased in PAP<sup>−/−</sup> mice compared to WT mice.
<p>A, bar plot showing the ratio (as percentage) between proliferative cell counts and total amount of cells. (**, P value <0.01; ***, P value <0.001). Error bars indicate S.E.M. values. B, bar plot showing the ratio (as percentage) between apoptotic cell counts and total amount of cells. Error bars indicate S.E.M. values.</p
DLP lobeexhibits the primary changes in the PAP<sup>−/−</sup> mouse prostate.
<p>The panels show an overview of the 12-old mice prostate dissected lobes. The DLP, AP and VP lobes were dissected from WT and PAP<sup>−/−</sup> mouse. The monolayer epithelium (white arrows) is seen in all the lobes of the WT mouse, whereas in the PAP<sup>−/−</sup> mouse an increased amount of cells is present in the lumen of the DLP lobe (black arrows). The AP and VP of PAP<sup>−/−</sup> mouse show no significant changes. Scale bars: 100 µm.</p
TMPAP is involved in endo-/exocytosis (proposed mechanism).
<p>TMPAP synthesized in the endoplasmic reticulum is transported in vesicles to the plasma membrane through the trans-Golgi network (TGN). After the vesicle docking and fusion events leading to release of vesicle content, TMPAP inserted in plasma membrane exerts its phosphatase function over AMP. The resulting product adenosine (Ado) activates the adenosine receptors, which are GPCRs, A1 or A3with G<sub>αi</sub> (inhibitory G-protein β-subunit) specificity leading to the inhibition of adenylate cyclase (AC) activity, and A2 adenosine receptors with G<sub>αs</sub> (stimulatory G-protein α-subunit) producing the stimulation of AC activity. Activated AC produces cAMP, which activates PKA responsible for the phosphorylation of snapin. The turnover is completed by clathrin-mediated endocytosis of SNARE components and TMPAP for recycling and degradation in lysosomes vÃa the endosomal-lysosomal pathway. From early endosomes, the cargo can be sorted to late endosomes or to MVE, which can follow the route leading to exosome release. Additional dephosphorylation events by TMPAP can occur while trafficking between different compartments. From late endosomes, TMPAP can go to lysosomes or back to TGN via the retrograde pathway. ATP: adenosine triphosphate, ADP: adenosine diphosphate, AMP: adenosine monophosphate, Ado: adenosine, TGN: trans-Golgi network, P: phosphate group, AP-2: adaptor protein complex 2, ADORA: adenosine receptor A (types A1, A2 and A3), AC: adenylate cyclase, G<sub>αs</sub>, G<sub>αi</sub>, G<sub>β</sub>, G<sub>γ</sub>: G-protein subunits, VDCC: Voltage-gated calcium channel. Synaptobrevin, syntaxin and SNAP25 are SNARE proteins. PI (4,5) P<sub>2</sub>: phosphatidylinositol 4,5-bisphosphate.</p
The prostate adenocarcinoma in PAP<sup>−/−</sup> mice is also detected by immunohistochemistry and electronmicroscopy.
<p>A, smooth muscle actin (SMA) immunohistochemistry in 12 month-old mice. Monolayer epithelium (mL) and open lumen in PAP<sup>+/+</sup> DLP. White arrows show the broken fibromuscular sheath (SM, smooth muscle) and bulging of epithelial cells to the stroma. Prostate adenocarcinoma (black arrows) is present in AP and DLP, showing a multilayer epithelium (ML) and inflammatory cells (black arrowhead) spreading in neighboring areas. Scale bars: 100 µm. (<i>n</i> =  4, per group). B, ultrastructural changes in 3 month-old and 12 month-old PAP<sup>−/−</sup> mouse DLPs. Monolayer epithelium, regular basement membrane (BM) and apical secretion are clearly seen in PAP<sup>+/+</sup> mouse DLPs. 3 month-old PAP<sup>−/−</sup> DLPs show irregular BM and numerous apical vacuoles (red arrow head), as well the presence of basal lysosomes (Ly). In 12 month-old PAP−/− mouse DLPs, the epithelium has transformed to a multilayer epithelium containing hyperchromatic nuclei with multiple nucleoli. Pseudolumens (pL) have formed as a result of the growing and fusion of the epithelium. Invaginations of BM (red arrows) into the epithelium and numerous vesicles in the basal side of the cells (blue arrow heads) were additional signs of the transformation in the cells. Scale bars: 2000 nm (<i>n</i> = 4, per group).</p
The colocalization of PAP (green) and GAD65/67 (red) was seen in several areas of brain.
<p>In cerebral Purkinje cells (A–C), strong colocalization was seen especially in the axon hillock of the neuron (small picture in C; yellow color and white arrows depicting the colocalization). Similarly, PAP was present in GABAergic neurons in prefrontal cortex (PFC; infralimbic cortex) (D–F). Scale bars are 10 µm.</p
Immunofluorescent colocalization stainings of TMPAP and synaptic vesicle associated proteins.
<p>TMPAP (green) is colocalized with a presynaptic marker, synaptophysin (red) (A–C; yellow color and white arrows depicting the colocalization). PAP was seen in vesicle-like structures that had strong colocalization with Snapin (D–F). Small picture is a magnification from panel C, depicting the colocalization. Moreover, largest PAP-immunoreactive structures had a colocalization with multivesicular bodies (MVB, red; G–I). All pictures are from striatum. Scale bars are 10 µm in A–C and G–I, and 3 µm in D–F.</p
TMPAP is expressed in the mouse brain and colocalizes with GABAergic marker, GAD 65/67.
<p>Representative confocal images depict intense TMPAP (brown color) expression in molecular cell layer (M) and Purkinje cells (P) of cerebellum (Panel A), in substantia nigra pars reticulata (SNpr; Panel B), in red nucleus (RN; Panel C) and in oculomotor nucleus (O; Panel C). Small picture in Panel B depicts the TMPAP staining of the substantia nigra in PAP<i><sup>−/−</sup></i> mouse. TMPAP (green) was colocalized with GABAergic marker (red) in medium spiny neurons of striatum (Panels D-G, yellow color and white arrows indicating the colocalization) and in SNpr (Panels H–K). Colocalization was evident also in GABAergic neurons of hippocampus CA1 (Panels L–O). DAPI (blue color) was used as a nuclear marker. Scale bars are 500 µm in Panels A–C, and 10 µm in panels D–O.</p
Immunostaining of Snapin in various brain areas of WT and PAP<sup>−/−</sup> mice.
<p>In the PAP<sup>−/−</sup> mouse, Snapin (red) is localized more diffusely in the cell soma (B, D, F). Cellular localization of Snapin in WT mouse is more vesicular-like (A, C, E). Scale bar is 10 µm in all figures. M1 - primary motor cortex; SN - substantia nigra.</p