A gene expression profile of Alzheimer's disease.

Postmortem analysis of brains of patients with Alzheimer's disease (AD) has led to diverse theories about the causes of the pathology, suggesting that this complex disease involves multiple physiological changes. In an effort to better understand the variety and integration of these changes, we generated a gene expression profile for AD brain. Comparing affected and unaffected brain regions in nine controls and six AD cases, we showed that 118 of the 7050 sequences on a broadly representative cDNA microarray were differentially expressed in the amygdala and cingulate cortex, two regions affected early in the disease. The identity of these genes suggests the most prominent upregulated physiological correlates of pathology involve chronic inflammation, cell adhesion, cell proliferation, and protein synthesis (31 upregulated genes). Conversely, downregulated correlates of pathology involve signal transduction, energy metabolism, stress response, synaptic vesicle synthesis and function, calcium binding, and cytoskeleton (87 downregulated genes). The results support several separate theories of the causes of AD pathology, as well as add to the list of genes associated with AD. In addition, approximately 10 genes of unknown function were found to correlate with the pathology.

Similar organization of the lipopolysaccharide-binding protein (LBP) and phospholipid transfer protein (PLTP) genes suggests a common gene family of lipid-binding proteins.

The transfer of lipids in aqueous environments such as serum has been attributed to a recently characterized class of proteins. Abnormal regulation of serum lipids by these proteins is thought to be a key event in the pathophysiology of cardiovascular diseases. Lipopolysaccharide (endotoxin) binding protein (LBP) was identified by virtue of its ability to bind bacterial lipid A. We have analyzed the exon-intron organization of the LBP gene and the nucleotide sequence of its approximately 20 kb spanning 5'- and 3'-untranslated regions. When comparing the genomic organization of LBP with that of two other genes coding for lipid transfer proteins, significant homologies were found. The LBP gene includes 15 exons, and the 2-kb promoter contains recognition elements of acute phase-typical reactants and a repetitive 12-mer motif with an as yet unknown protein-binding property. Detailed sequence comparison revealed a closer relatedness of LBP with PLTP than with CETP as demonstrated by an almost identical intron positioning. This high degree of similarity supports functional studies by others suggesting that like LBP, PLTP may also be able to bind and transport bacterial lipopolysaccharide.

Cloning and characterization of novel rat and mouse low molecular weight Ca(2+)-dependent phospholipase A2s containing 16 cysteines.

A novel rat 4.4-kilobase (kb) cDNA encoding a low molecular weight Ca(2+)-dependent phospholipase A2 (PLA2) and its murine homologue were cloned. The rat and mouse cDNA predict a mature protein of 130 amino acids (M(r) = 14, 763) preceded by a 28-amino acid prepro-peptide. The deduced amino acid sequences encode a protein containing 16 cysteines which distinguishes them from both mammalian group I and II PLA2s and the recently described group of mammalian PLA2s containing 12 cysteines. A rat RNA blot hybridized with the rat cDNA exhibited an abundant 2.3-kb and a less abundant 5-kb transcript in testis. When the rat cDNA was expressed using an Epstein-Barr virus-based vector in human 293s cells, PLA2 activity accumulated in the culture medium. Conditioned medium optimally hydrolyzed the phospholipids of [1-14C]oleate-labeled Escherichia coli at neutral to alkaline pH with 1-7 mM Ca2+. In assays with individual substrates, L-alpha-1-stearoyl-2-arachidonyl phosphatidylinositol was hydrolyzed more efficiently than L-alpha-1-palmitoyl-2-oleoyl phosphatidylcholine, L-alpha-1-palmitoyl-2-arachidonyl phosphatidylcholine, or L-alpha-1-palmitoyl-2-arachidonyl phosphatidylethanolamine.

Cloning and recombinant expression of a novel human low molecular weight Ca(2+)-dependent phospholipase A2.

Extensive biochemical studies of phospholipase A2s (PLA2s) over the last two decades indicate that there are likely to be several distinct PLA2 genes in mammals. Here we report the cloning of a 1-kilobase pair cDNA encoding a novel human low molecular weight PLA2. The cDNA appears to encode a 118-amino acid mature peptide (M(r) = 13,592) preceded by a 20-residue prepeptide. The deduced amino acid sequence encodes a protein that lacks one of the seven disulfide bridges found in similar PLA2s and, therefore, represents a class of enzymes distinct from the mammalian group I and group II enzymes. An RNA blot hybridized with the cDNA exhibited a putative 1.2-kilobase pair transcript in heart and, less abundantly, in lung, as well as multiple putative transcripts in placenta. When the cDNA was expressed using an Epstein-Barr virus-based vector in human 293s cells, PLA2 activity accumulated in the culture medium. Conditioned medium optimally hydrolyzed the phospholipids of [1-14C]oleate-labeled Escherichia coli at neutral to alkaline pH with 10 mM or greater Ca2+. In assays done with individual substrates, L-alpha-1-palmitoyl-2-oleoyl phosphatidylcholine was more efficiently hydrolyzed than L-alpha-1-palmitoyl-2-arachidonyl phosphatidylcholine, L-alpha-1-palmitoyl-2-arachidonyl phosphatidylethanolamine, or L-alpha-1-stearoyl-2-arachidonyl phosphatidylinositol.