Intron-3 Retention/Splicing Controls Neuronal Expression of Apolipoprotein E in the CNS

Abstract

Neuronal expression of apolipoprotein (apo) E4 may contribute to the pathogenesis of Alzheimer's disease (AD). In studying how apoE expression is regulated in neurons, we identified a splicing variant of apoE mRNA with intron-3 retention (apoE-I3). ApoE-I3 mRNA was detected in neuronal cell lines and primary neurons, but not in astrocytic cell lines or primary astrocytes, from humans and mice by reverse transcription (RT)-PCR. In both wild-type and human apoE knock-in mice, apoE-I3 was found predominantly in cortical and hippocampal neurons by in situ hybridization. Cell fractionation and quantitative RT-PCR revealed that over 98% of the apoE-I3 mRNA was retained in the nucleus without protein translation. In transfected primary neurons, apoE expression increased dramatically when intron-3 was deleted from a genomic DNA construct and decreased markedly when intron-3 was inserted into a cDNA construct, suggesting that intron-3 retention/splicing controls apoE expression in neurons. In response to excitotoxic challenge, the apoE-I3 mRNA was markedly increased in morphologically normal hippocampal neurons but reduced in degenerating hippocampal neurons in mice; apoE mRNA showed the opposite pattern. This apparent precursor-product relationship between apoE-I3 and apoE mRNA was supported by a transcriptional inhibition study. Thus, neuronal expression of apoE is controlled by transcription of apoE-I3 under normal conditions and by processing of apoE-I3 into mature apoE mRNA in response to injury.

Apolipoprotein E ε4 is associated with disease-specific effects on brain atrophy in Alzheimer's disease and frontotemporal dementia

“…Through A␤-independent pathways (1), apoE4 may contribute to neurodegenerative diseases other than AD. One such mechanism is the ''two-hit'' hypothesis (13,29). In response to stressors or injury (''first hit''), neurons begin to synthesize apoE (1,29).…”

“…One such mechanism is the ''two-hit'' hypothesis (13,29). In response to stressors or injury (''first hit''), neurons begin to synthesize apoE (1,29). Although apoE may promote neuronal repair (2,29), it can also undergo proteolytic cleavage.…”

“…In response to stressors or injury (''first hit''), neurons begin to synthesize apoE (1,29). Although apoE may promote neuronal repair (2,29), it can also undergo proteolytic cleavage. Carboxyl-terminal-truncated apoE fragments may disrupt the cytoskeleton, stimulate tau phosphorylation, impair mitochondrial function, and ultimately cause cell death (''second hit'') (29).…”

“…Although apoE may promote neuronal repair (2,29), it can also undergo proteolytic cleavage. Carboxyl-terminal-truncated apoE fragments may disrupt the cytoskeleton, stimulate tau phosphorylation, impair mitochondrial function, and ultimately cause cell death (''second hit'') (29). The apoE4 isoform may have the most detrimental effect because it is more susceptible to proteolytic cleavage than E3 or E2 (1).…”

Cellular Source of Apolipoprotein E4 Determines Neuronal Susceptibility to Excitotoxic Injury in Transgenic Mice

“…30 Our study suggests that astroglial production of apoE may be excitoprotective in both APOE 3 carriers and APOE 4 carriers and that APOE 4 carriers, but not APOE 3 carriers, may benefit from suppression of apoE expression specifically in neurons. Recent studies have identified unique mechanisms by which neuronal apoE expression is regulated 11 that might be exploited to selectively suppress expression of apoE4 by neurons.…”