Tumor suppressor PTEN affects tau phosphorylation: deficiency in the phosphatase activity of PTEN increases aggregation of an FTDP-17 mutant Tau

BACKGROUND: Aberrant hyperphosphorylation of tau protein has been implicated in a variety of neurodegenerative disorders. Although a number of protein kinases have been shown to phosphorylate tau in vitro and in vivo, the molecular mechanisms by which tau phosphorylation is regulated pathophysiologically are largely unknown. Recently, a growing body of evidence suggests a link between tau phosphorylation and PI3K signaling. In this study, phosphorylation, aggregation and binding to the microtubule of a mutant frontal temporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) tau in the presence of tumor suppressor PTEN, a major regulatory component in PI3K signaling, were investigated. RESULTS: Phosphorylation of the human mutant FTDP-17 tau, T40RW, was evaluated using different phospho-tau specific antibodies in the presence of human wild-type or phosphatase activity null mutant PTEN. Among the evaluated phosphorylation sites, the levels of Ser214 and Thr212 phospho-tau proteins were significantly decreased in the presence of wild-type PTEN, and significantly increased when the phosphatase activity null mutant PTEN was ectopically expressed. Fractionation of the mutant tau transfected cells revealed a significantly increased level of soluble tau in cytosol when wild-type PTEN was expressed, and an elevated level of SDS-soluble tau aggregates in the presence of the mutant PTEN. In addition, the filter/trap assays detected more SDS-insoluble mutant tau aggregates in the cells overexpressing the mutant PTEN compared to those in the cells overexpressing wild-type PTEN and control DNA. This notion was confirmed by the immunocytochemical experiment which demonstrated that the overexpression of the phosphatase activity null mutant PTEN caused the mutant tau to form aggregates in the COS-7 cells. CONCLUSION: Tumor suppressor PTEN can alleviate the phosporylation of the mutant FTDP-17 tau at specific sites, and the phosphatase activity null PTEN increases the mutant tau phosphorylation at these sites. The changes of the tau phosphorylation status by ectopic expression of PTEN correlate to the alteration of the mutant tau's cellular distribution. In addition, the overexpression of the mutant PTEN can increase the level of the mutant tau aggregates and lead to the formation of visible aggregates in the cells

Statins in unconventional secretion of insulin-degrading enzyme and degradation of the amyloid-β peptide.

Population-based studies demonstrated that statins might decrease the risk of developing Alzheimer's disease (AD). Statins inhibit the 3-hydroxy-3-methyl-glutaryl-coenzyme-A reductase and thereby de novo synthesis of cholesterol. Cell culture and animal studies indicated that cholesterol affects the proteolytic processing of the amyloid precursor protein and the generation of amyloid-β (Aβ). Recently, we have demonstrated that statins can also stimulate the degradation of Aβ. The statin-induced clearance of Aβ could be attributed to increased release of the insulin-degrading enzyme (IDE) via an exosome-related unconventional secretory pathway. Interestingly, this statin-induced secretion of exosome-associated IDE was independent of cellular cholesterol concentrations, but rather caused by impairment of isoprenoid biosynthesis and protein prenylation. We further identified a new hexapeptide sequence in the C-terminal region of IDE, named the SlyX motif that is critically involved in IDE secretion. Taken these findings together, the increased clearance of Aβ by stimulated secretion of IDE might contribute to the protective effects of statins against AD

Loss of Caveolin-1 Accelerates Neurodegeneration and Aging

The aged brain exhibits a loss in gray matter and a decrease in spines and synaptic densities that may represent a sequela for neurodegenerative diseases such as Alzheimer's. Membrane/lipid rafts (MLR), discrete regions of the plasmalemma enriched in cholesterol, glycosphingolipids, and sphingomyelin, are essential for the development and stabilization of synapses. Caveolin-1 (Cav-1), a cholesterol binding protein organizes synaptic signaling components within MLR. It is unknown whether loss of synapses is dependent on an age-related loss of Cav-1 expression and whether this has implications for neurodegenerative diseases such as Alzheimer's disease.We analyzed brains from young (Yg, 3-6 months), middle age (Md, 12 months), aged (Ag, >18 months), and young Cav-1 KO mice and show that localization of PSD-95, NR2A, NR2B, TrkBR, AMPAR, and Cav-1 to MLR is decreased in aged hippocampi. Young Cav-1 KO mice showed signs of premature neuronal aging and degeneration. Hippocampi synaptosomes from Cav-1 KO mice showed reduced PSD-95, NR2A, NR2B, and Cav-1, an inability to be protected against cerebral ischemia-reperfusion injury compared to young WT mice, increased Aβ, P-Tau, and astrogliosis, decreased cerebrovascular volume compared to young WT mice. As with aged hippocampi, Cav-1 KO brains showed significantly reduced synapses. Neuron-targeted re-expression of Cav-1 in Cav-1 KO neurons in vitro decreased Aβ expression.Therefore, Cav-1 represents a novel control point for healthy neuronal aging and loss of Cav-1 represents a non-mutational model for Alzheimer's disease

Tumor-suppressor PTEN affects tau phosphorylation, aggregation, and binding to microtubules

Neurofibrillary tangles (NFTs), consisting of abnormally hyperphosphorylated tau, are implicated in the pathogenesis of several neurodegenerative diseases including Alzheimer's disease (AD). The molecular mechanisms underlying the regulation of tau phosphorylation are largely unknown. While the PI3K/ Akt pathway has been shown to regulate multiple cellular events pertinent to AD pathogenesis, potential functions of tumor suppressor phosphatase and tensin homologue deleted on chromosome 10 (PTEN) in AD pathogenesis have not been explored. Here, we examine the effects of PTEN on tau phosphorylation, its microtubule association and formation of aggregates, and consequentially neuronal morphology. In cultured cells, overexpression of wild-type (WT) PTEN alters tau phosphorylation at several sites, increases tau-microtubule association and decreases formation of tau aggregates. In addition, the phosphatase-null PTEN increases tau aggregation and impairs tau binding to microtubule and neurite outgrowth of neurons expressing the mutant PTEN. We also found a significant loss of PTEN in AD patient brains correlated with a dramatically increased concentration of phospho-tau at Ser-214 in NFTs. Together, our results demonstrate that PTEN regulates tau phosphorylation, binding to micro-tubules and formation of aggregates and neurite outgrowth. These findings suggest a link between malfunction of PTEN and tauopathy, and imply PTEN as a therapeutic target for tauopathy.-Zhang, X., Li, F., Bulloj, A., Zhang, Y.-w., Tong, G., Zhang, Z., Liao, F.-F., Xu, H. Tumor-suppressor PTEN affects tau phosphorylation, aggregation, and binding to microtubules

Insulin-degrading enzyme is not secreted from cultured cells

Abstract Insulin-degrading enzyme (IDE) functions in the catabolism of bioactive peptides. Established roles include degrading insulin and the amyloid beta peptide (Aβ), linking it to diabetes and Alzheimer’s disease. IDE is primarily located in the cytosol, and a longstanding question is how it gains access to its peptide substrates. Reports suggest that IDE secreted by an unconventional pathway participates in extracellular hydrolysis of insulin and Aβ. We find that IDE release from cultured HEK-293 or BV-2 cells represents only ~1% of total cellular IDE, far less than has been reported previously. Importantly, lactate dehydrogenase (LDH) and other cytosolic enzymes are released at the same relative level, indicating that extracellular IDE results from a loss of cell integrity, not secretion. Lovastatin increases IDE release from BV-2 cells as reported, but this release is mirrored by LDH release. Cell viability assays indicate lovastatin causes a loss of cell integrity, explaining its effect on IDE release. IDE is present in an exosome-enriched fraction from BV-2 cell conditioned media, however it represents only ~0.01% of the total cellular enzyme and is unlikely to be a significant source of IDE. These results call into question the secretion of IDE and its importance in extracellular peptide degradation

Nitric Oxide Decreases the Enzymatic Activity of Insulin Degrading Enzyme in APP/PS1 Mice

peer reviewedNitric oxide has been implicated in the regulation of enzyme activity, particularly the activity of metalloproteinases. Since the inducible form of the nitric oxide synthase (NOS2), is upregulated in Alzheimer's disease, we investigated the activity of two amyloid β degrading enzymes, IDE and neprilysin. In vitro we demonstrated that the activity of IDE was inhibited by *NO donor Sin-1, whereas activity of neprilysin remained unaffected. In vivo the activity of insulin-degrading enzyme was lowered in APP/PS1 mice, but not in APP/PS1/NOS2(-/-) mice. These data suggest that NOS2 upregulation impairs amyloid β degradation through negative regulation of IDE activity and thus loss of NOS2 activity will positively influence amyloid β clearance