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S Gandy

Publications and source records attributed to S Gandy.

At least 55 records · Page 3Linked to original sources
PubMed1994

Regulated cleavage of the Alzheimer amyloid precursor protein: molecular and cellular basis.

The relative utilization of alternative processing pathways for APP can be regulated by the activation state of certain protein phosphorylation signal transduction pathways. For example, activation of protein kinase C (PKC), or inactivation of protein phosphatases 1 and 2A, leads to a relative increase in utilization of the nonamyloidogenic, 'alpha-secretase' cleavage pathway for APP processing at the expense of other pathways. The molecular and cellular basis for this regulatory event is unknown. The possible mechanisms of regulated APP cleavage include (either singly or in combination): 1) substrate (ie APP) activation; 2) substrate redistribution; 3) enzyme (ie alpha-secretase) activation; or 4) enzyme redistribution. APP is a phosphoprotein; however, recent evidence from studies of the metabolism of mutant APP molecules suggests that changes in the APP cytoplasmic tail phosphorylation state may not be necessary for the phosphorylation-dependent activation of 'alpha-secretase' cleavage. Further, indirect immunofluorescent studies of the subcellular distribution of APP in the absence or presence of phorbol esters (PKC activators) fail to disclose obvious phorbol-induced redistribution of APP immunoreactivity. Taken together, current data suggest that major candidate phosphorylation-state sensitive targets relevant to the molecular basis of PKC-activated processing (or 'regulated cleavage') of APP include the APP ectodomain as well as secretase enzymes and/or other components of the APP trafficking/processing apparatus. Progress in distinguishing among these possibilities is discussed.

Alzheimer Disease↗
PubMed1993

Serine phosphorylation of the secreted extracellular domain of APP.

The phosphorylation status of full-length APP (FL-APP) and secreted APP (s-APP) was investigated in stably transfected cells. 32P incorporation was detected in the mature full-length APP both in the absence and presence of phorbol ester. Surprisingly, 32P-phosphate was incorporated in the secreted ectodomain, and this was stable to treatment of the [32P]-phospho-s-APP with a large excess of PNGase F, suggesting that N-linked oligosaccharide sites do not account for phosphate incorporation. Phosphoamino acid analysis of the [32P]-phospho-s-APP resulted in the recovery of [32P]-phosphoserine as the preponderant species. Brefeldin A completely inhibited the release of [32P]-phospho s-APP, but did not inhibit the incorporation of 32P into the FL-APP, suggesting that phosphorylation occurs early in the central vacuolar pathway. It is possible that ectodomain phosphorylation by a novel luminal or extracellular protein kinase may play a role in regulating the metabolic fate of APP.

Amino Acids↗
PubMed1993

Regulated cleavage of Alzheimer beta-amyloid precursor protein in the absence of the cytoplasmic tail.

Alzheimer beta-amyloid precursor protein can be phosphorylated on residues Thr654, Ser655 and Thr668 on its cytoplasmic domain. Proteolytic cleavage of the amyloid precursor protein and release of the amyloid precursor protein ectodomain into the medium of cultured cells can be activated by phorbol esters which stimulate protein kinase C. In the present study, using mutated amyloid precursor protein, we show that phosphorylation of cytoplasmic residues is not required for the phorbol ester-activated cleavage and release of the amyloid precursor protein ectodomain. Remarkably, deletion of the entire amyloid precursor protein cytoplasmic tail had no effect on the phorbol ester-activated cleavage/release. The results indicate that activation of amyloid precursor protein cleavage/release by protein kinase C involves phosphorylation of some component of the processing pathway, instead of or in addition to the cytoplasmic tail of the amyloid precursor protein.

Amino Acid Sequence↗
PubMed1992

Amyloidogenesis in Alzheimer's disease: some possible therapeutic opportunities.

Cerebral deposition of fibrils formed from the beta/A4 amyloid protein is an invariable feature of Alzheimer's disease. Evidence suggests that generation of such fibrils may be involved in the etiology of this disease, since mutations in the coding region of the beta/A4 amyloid precursor protein (APP) gene segregate with familial cerebral amyloidoses, including familial Alzheimer's disease. Transgenic models of cerebral amyloidosis have been produced, and some progress has been made in elucidating the cell biology of amyloidogenesis. For example, agents that alter protein phosphorylation are potent modulators of the expression and proteolytic processing of APP. Sam Gandy and Paul Greengard review these recent studies, and discuss those that may provide rational therapeutic opportunities.

Alzheimer Disease↗
PubMed1990

A "G" to "A" mutation at position -1 of a 5' splice site in a late infantile form of Tay-Sachs disease.

Tay-Sachs disease is an autosomal recessive genetic disease caused by a deficiency in beta-hexosaminidase A. We have characterized a new mutation in a Tunisian patient displaying a late infantile form of Tay-Sachs disease. Northern blot analysis of patient's fibroblast total RNAs shows a broad, fast migrating band in the region of the normal beta-hexosaminidase alpha transcripts. The mRNA coding for beta-hexosaminidase alpha subunit was first reverse transcribed and then amplified in four overlapping segments spanning the entire coding sequence by polymerase chain reaction. We found in the products of polymerase chain reaction (PCR) that amplify the segment spanning exons 2-7, in addition to a normal fragment, two smaller size fragments, one of which is also seen in normal control fibroblasts. The analysis of the patient's specific abnormal fragment by hybridization with exon-specific oligonucleotides and then sequencing allowed us to conclude that this fragment lacked exon 5. The other smaller species lacked exons 4 and 5 in both patient and normal control. The sequence of a genomic fragment containing exon 5 and of the patient's normal cDNA fragment spanning exons 2-7, revealed a point mutation G to A at the last nucleotide of exon 5. This mutation doesn't change the sense of the affected codon. Northern blot of patient's fibroblast poly(A+) RNAs allowed us to quantify two of the forms of transcripts seen by PCR. In the patient, the normal size transcript and the exon 5-deleted transcript represent, respectively, 3 and 7% of the normal control beta-hexosaminidase alpha mRNA. We propose that this point mutation is responsible for an inefficient and abnormal processing of the mutant transcript resulting in the appearance of two low abundance spliced mRNAs. One is lacking exon 5 and most likely codes for an inactive protein; the other is similar to normal beta-hexosaminidase alpha mRNA, except for the presence of the silent G to A mutation and codes therefore for a normal enzyme accounting for the 2.5% residual beta-hexosaminidase A activity measured in patient's fibroblasts by a fluorometric assay. The third form, without exons 4 and 5, is also evidenced in normal fibroblasts by PCR so that we think that it is not related to Tay-Sachs disease.

Adenine↗
PubMed1989

Comparison of the physical and recombination maps of the mouse X chromosome.

The locations of five random mouse genomic DNA markers and five cloned genes, including the genes for clotting factors VIII and IX (Cf-8 and Cf-9), Duchenne muscular dystrophy (Dmd), phosphoglycerate kinase-1 (Pgk-1), and alpha-galactosidase (Ags), on the mouse X chromosome were determined by in situ hybridization. The five random DNA markers provide new genetic loci with useful restriction fragment length polymorphisms between mouse strains and species, including one locus close to the centromeric region of the mouse X chromosome. The physical map and the recombination map of these loci on the X chromosome were compared. There was good agreement in the order of loci. Relative distances between loci were consistent along the X chromosome, with the exception of the telomeric end of the long arm, where the recombination fraction observed between loci closely associated on the physical map was higher than that between similarly spaced markers located in the proximal region of the X chromosome. These results are discussed in comparison to the human X-chromosome map.

Animals↗
PubMed1988

Phosphorylation of Alzheimer disease amyloid precursor peptide by protein kinase C and Ca2+/calmodulin-dependent protein kinase II.

The amino acid sequence of the Alzheimer disease amyloid precursor (ADAP) has been deduced from the corresponding cDNA, and hydropathy analysis of the sequence suggests a receptor-like structure with a single transmembrane domain. The putative cytoplasmic domain of ADAP contains potential sites for serine and threonine phosphorylation. In the present study, synthetic peptides derived from this domain were used as model substrates for various purified protein kinases. Protein kinase C rapidly catalyzed the phosphorylation of a peptide corresponding to amino acid residues 645-661 of ADAP [ADAP peptide(645-661)] on Ser-655. Ca2+/calmodulin-dependent protein kinase II phosphorylated ADAP peptide (645-661) on Thr-654 and Ser-655. This peptide was virtually ineffective as a substrate for cAMP-dependent protein kinase, cGMP-dependent protein kinase, casein kinase II, or insulin receptor protein-tyrosine kinase. When a homogenate of rat cerebral cortex was used as the source of protein kinase, phosphorylation of ADAP peptide(645-661) was stimulated by calcium/phosphatidylserine/diolein to a level 4.6-fold above the basal level of phosphorylation, consistent with a prominent stimulation by protein kinase C. Using rat cerebral cortex synaptosomes prelabeled with 32Pi, a 32P-labeled phosphoprotein of approximately equal to 135 kDa was immunoprecipitated by using antisera prepared against ADAP peptide(597-624), consistent with the possibility that the holoform of ADAP in rat brain is a phosphoprotein. Based on analogy with the effect of phosphorylation by protein kinase C of juxtamembrane residues in the cytoplasmic domain of the epidermal growth factor receptor and the interleukin 2 receptor, phosphorylation of ADAP may target it for internalization.

Alzheimer Disease↗
PubMed1986

Back pain in the elderly: updated diagnosis and management.

CT of compression fractures is a useful adjunct to the plain x-ray in excluding signs of metastasis. However, increased bone density on CT may not distinguish osteoporotic fractures from neoplastic disease, in which case radioiodine scan, bone and marrow biopsies, or myelography may be necessary. Surgery for painful osteoarthritic spinal disease is controversial. The potential advantages of surgery must be weighted against the risk of anesthesia, the length and tolerance of postoperative immobility, and the effect of laminectomy and fusion on the biomechanics of the spine. Furthermore, the elderly are at increased risk for postoperative complications.

Aged↗
PubMed1985

Isolation and characterization of two repetitive DNA fragments located near the centromere of the mouse X chromosome.

Two repetitive DNA fragments located on the mouse X chromosome are described. The fragments were isolated from a lambda phage library enriched in X-chromosomal sequences by flow sorting. Both fragments, which are repeated 20 to 50 times in the genome, were mapped to the mouse X chromosome by Southern blot hybridization to DNA from hybrid cells retaining the mouse X chromosome, by dosage analysis, and by in situ hybridization to mouse chromosomes. In mouse strain C57BL/10BK, one fragment appeared to be located only on the X chromosome, while the other fragment had homologous sequences on chromosome 11 in addition to the X chromosome. The latter fragment showed DNA variants between mouse strains, which are potentially useful for mapping. Both fragments cross-hybridized to another mouse species: Mus caroli. In this species, each fragment appeared to be located on the X chromosome, indicating that some X-chromosome repetitive sequences are partially conserved. In addition, one fragment cross-hybridized to human DNA.

Animals↗
PubMed1984

Localization of copper-zinc superoxide dismutase in the endocrine pancreas.

Immunoelectron microscopy and cellular fractionation on sucrose density gradients have been used to examine the intracellular distribution of copper-zinc superoxide dismutase (SOD) in the canine and rat endocrine pancreas. Using rabbit anti-canine copper-zinc SOD as the primary antiserum, immunostaining in canine beta and non-beta islet cells was significantly greater than that in serial sections of the same islet incubated with preimmune serum from the same rabbit. Within the cells, immunostaining was associated with spherical and crystalloid granules as well as with the cytoplasm. Radioimmune assays of cellular fractions or rat islet-cell preparations showed that the granule-rich fraction containing large amounts of insulin was also rich in SOD. This interesting observation of association of SOD with hormone-containing granules suggests that SOD may play a role in protecting these oxidation-sensitive proteins.

Animals↗