Xu Q, Walker D, Bernardo A, Brodbeck J, Balestra ME, Huang Y.
Intron-3 retention/splicing controls neuronal expression of apolipoprotein E in the CNS.
J Neurosci. 2008 Feb 6;28(6):1452-9.
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Comment by Chunjiang Yu and Mary Jo LaDu
In this paper, Yadong Huang and his colleagues identify a regulatory mechanism, relatively common for other genes, that is unique in its application to ApoE. In so doing, this work also addresses the long-standing controversy of whether neurons express ApoE. The authors demonstrate that ApoE is constitutively expressed in hippocampal neurons but retained in their nuclei because of alternative splicing of intron 3. Conditioned media from rat C6 cells (a glial-derived cell line) in vitro, or kainic acid treatment in vivo, results in the splicing of intron 3 and export of mature ApoE mRNA from the nucleus to the cytosol for translation. These results provide a nice molecular interpretation of the early work by this group (2), which showed that conditioned media from C6 cells or ApoE-KO mouse primary glia regulate neuronal ApoE expression (RT-PCR, WB) in stable N2a lines, NT2 cells, and mouse primary neurons.
However, to these readers, at least, these results are in apparent contrast to the group’s earlier work using ApoE-EGFP knock-in mice (3). If ApoE is constitutively expressed in hippocampal neurons, then GFP would be constitutively expressed in these mice, as well, because the EGFP cDNA contains a polyA site in the KI construct. This would free the mRNA from ApoE intron 3 retention and result in release from the nucleus, and translation. This was not the observation. In the knock-in model, hippocampal neurons only express GFP with kainic acid treatment.
In addition, we have these minor technical questions:
1. While TR-PCR and 5’- and 3’-RACE results strongly suggest that ApoE-I3 is expressed in neurons, a direct demonstration of this transcript by Northern blot would be very convincing.
2. We wonder how consistent the expression of ApoE-I3 is. In situ results show robust control expression of ApoE-I3 in the entire hippocampus (Fig. 2E), but weak expression in CA3 and the dentate gyrus (Fig. 6A) when compared to increased expression induced by kainic acid.
References:1. Xu Q, Walker D, Bernardo A, Brodbeck J, Balestra ME, Huang Y. Intron-3 retention/splicing controls neuronal expression of apolipoprotein E in the CNS. J Neurosci. 2008 Feb 6;28(6):1452-9. Abstract
2. Harris FM, Tesseur I, Brecht WJ, Xu Q, Mullendorff K, Chang S, Wyss-Coray T, Mahley RW, Huang Y. Astroglial regulation of apolipoprotein E expression in neuronal cells. Implications for Alzheimer's disease. J Biol Chem. 2004 Jan 30;279(5):3862-8. Abstract
3. Xu Q, Bernardo A, Walker D, Kanegawa T, Mahley RW, Huang Y. Profile and regulation of apolipoprotein E (ApoE) expression in the CNS in mice with targeting of green fluorescent protein gene to the ApoE locus. J Neurosci. 2006 May 10;26(19):4985-94. Abstract