Restricted Location of PSEN2/γ-Secretase Determines Substrate Specificity and Generates an Intracellular Aß Pool.

γ-Secretases are a family of intramembrane-cleaving proteases involved in various signaling pathways and diseases, including Alzheimer's disease (AD). Cells co-express differing γ-secretase complexes, including two homologous presenilins (PSENs). We examined the significance of this heterogeneity and identified a unique motif in PSEN2 that directs this γ-secretase to late endosomes/lysosomes via a phosphorylation-dependent interaction with the AP-1 adaptor complex. Accordingly, PSEN2 selectively cleaves late endosomal/lysosomal localized substrates and generates the prominent pool of intracellular Aß that contains longer Aß; familial AD (FAD)-associated mutations in PSEN2 increased the levels of longer Aß further. Moreover, a subset of FAD mutants in PSEN1, normally more broadly distributed in the cell, phenocopies PSEN2 and shifts its localization to late endosomes/lysosomes. Thus, localization of γ-secretases determines substrate specificity, while FAD-causing mutations strongly enhance accumulation of aggregation-prone Aß42 in intracellular acidic compartments. The findings reveal potentially important roles for specific intracellular, localized reactions contributing to AD pathogenesis.

CA 15-3 serum levels in patients with ductal breast carcinoma: relationship with clinicopathological parameters and tumor markers.

INTRODUCTION: Cancer antigen 15-3 (CA 15-3) is the most widely used serum marker in diagnosing and monitoring breast cancer. The aim of this work was to analyze preoperative CA 15-3 serum levels in patients with ductal breast carcinoma in relation to 1) clinicopathological parameters, 2) hormone receptors, and 3) tissue-based tumor markers. METHODS: A group of 340 women with infiltrating ductal carcinoma of the breast who had undergone no prior treatment was studied. These women ranged in age between 27 and 83 years (mean age 61.5±9.9 years). Preoperative CA 15-3 serum levels were determined by an immunoradiometric method. Hormone receptors (estrogen, progesterone and androgen receptors), p53, bcl-2 and Ki67 were determined by different immunohistochemical methods. Epidermal growth factor receptor (EGFR) was analyzed in the cell membranes by radioligand assay whereas cathepsin D and pS2 were determined by immunoradiometric analysis. Tumor ploidy and S-phase fraction were studied by flow cytometry. RESULTS: CA 15-3 serum levels were higher in postmenopausal women (p=0.032), in patients with tumors exceeding 2 cm (p=0.003), lymph node involvement (p=0.026), distant metastases (M1) (p<0.0001), S-phase fraction <7% (p=0.015), EGFR <6 fmol/mg protein (p=0.025), and cathepsin D <50 pmol/mg protein (p=0.023). CONCLUSIONS: Preoperative CA 15-3 serum levels were associated with cell proliferation determined by the S-phase fraction, the concentration of cathepsin D in the cytosol, and the EGFR concentration in the cell membrane.

Association between deposition of beta-amyloid and pathological prion protein in sporadic Creutzfeldt-Jakob disease.

BACKGROUND: Alzheimer's disease (AD) and prion diseases such as sporadic Creutzfeldt-Jakob disease (sCJD) share common features concerning their molecular pathogenesis and neuropathological presentation and the coexistence of AD and CJD in patients suggest an association between the deposition of the proteolytically processed form of the amyloid precursor protein, beta-amyloid (Abeta), which deposits in AD, and the abnormal form of the prion protein, PrP(Sc), which deposits in sCJD. METHODS: We have characterized sCJD patients (n = 14), AD patients (n = 5) and nondemented controls (n = 5) with respect to the deposition of PrP(Sc) and Abeta morphologically, biochemically and genetically and correlated these findings to clinical data. RESULTS: sCJD-diseased individuals with abundant deposits of Abeta present with a specific clinicopathological profile, defined by higher age at disease onset, long disease duration, a genetic profile and only minimal amounts of PrP(Sc) in the cerebellum. CONCLUSION: The co-occurrence of pathological changes typical for sCJD and AD in combination with the inverse association between accumulation of Abeta and PrP(Sc) in a subgroup of sCJD patients is indicative of common pathways involved in the generation or clearance of Abeta and PrP(Sc) in a subgroup of sCJD patients.

Biochemical properties of endogenous presenilin 1 and presenilin 2 in cultured human B-lymphocytes.

BACKGROUND: Presenilin 1 (PS1) and presenilin 2 (PS2) are membranous proteins involved in the pathology of Alzheimer's disease. The development of specific therapies targeted at PS1 or PS2 requires the determination of biochemical properties of presenilins. Hence, in this study we analyzed the hydrophobic and ionic properties of endogenous presenilins. METHODS: Lysates of immortalized human B-lymphocytes were used as a source of endogenous presenilins. The presence of 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonate (CHAPSO) detergent in lysates favored preservation of PS1 and PS2 native protein complexes. We compared Kyte-Doolittle hydropathicity profiles and hydrophobic interactions of PS1 and PS2 with phenyl-agarose. We also compared the ionic properties of presenilins using anion-exchange chromatography. RESULTS: The hydropathicity profiles of PS1 and PS2 revealed similarly located hydrophobic regions and more hydrophobic region in the C-terminal fragment of PS2. However, both PS1 and PS2 under physiological conditions showed no interactions with phenyl-agarose. Despite similar predicted isoelectric points, PS1 and PS2 exhibited different ionic behavior during anion-exchange chromatography. CONCLUSIONS: The different than expected hydrophobic and ionic behavior of PS1 and PS2 may be caused by interactions with other proteins present in complexes formed by endogenous presenilins. The observed difference in ionic properties of PS1 and PS2 can be further explained assuming that PS1 and PS2 form complexes with different sets of proteins. The composition of such variegated PS1 and PS2 complexes can be explored using a proteomic approach. The difference in PS1 and PS2 ionic behavior can be used for purification of endogenous PS1 from PS2, which has not yet been achieved by any other means.

Alzheimer-associated presenilins 1 and 2: neuronal expression in brain and localization to intracellular membranes in mammalian cells.

Mutations in two recently identified genes appear to cause the majority of early-onset familial Alzheimer's disease (FAD). These two novel genes, presenilin 1 (PS1) and presenilin 2 (PS2) are members of an evolutionarily conserved gene family. The normal biological role(s) of the presenilins and the mechanism(s) by which the FAD-associated mutations exert their effect remain unknown. Employing in situ hybridization, we demonstrate that the expression patterns of PS1 and PS2 in the brain are extremely similar to each other and that messages for both are primarily detectable in neuronal populations. Immunochemical analyses indicate that PS1 and PS2 are similar in size and localized to similar intracellular compartments (endoplasmic reticulum and Golgi complex). FAD-associated mutations in PS1 and PS2 do not significantly modify either their migration patterns on SDS-polyacrylamide gel electrophoresis or their overall subcellular localization, although subtle differences in perinuclear staining were noted for mutant PS1.