Alternative polyadenylation of MeCP2: Influence of cis-acting elements and trans-acting factors.
The human MeCP2 gene encodes a ubiquitously expressed methyl CpG binding protein. Mutations in this gene cause a neurodevelopmental disorder called Rett Syndrome (RS). Mutations identified in the coding region of MeCP2 account for approximately 65% of all RS cases. However, 35% of all patients do not show mutations in the coding region of MeCP2, suggesting that mutations in non-coding regions likely exist that affect MeCP2 expression rather than protein function. The gene is unusual in that is has a >8.5 kb 3' untranslated region (3' UTR), and the size of the 3'UTR is differentially regulated in various tissues because of distinct polyadenylation signals. We have identified putative cis-acting auxiliary regulatory elements that play a role in alternative polyadenylation of MeCP2 using an in vivo polyadenylation reporter assay and in a luciferase assay. These cis-acting auxiliary elements are found both upstream and downstream of the core CPSF binding sites. Mutation of one of these cis-acting auxiliary elements, a G-rich element (GRS) significantly reduced MeCP2 polyadenylation efficiency in vivo. We further investigated what trans-acting factor(s) might be binding to this cis-acting element and found that hnRNP F protein binds specifically to the element. We next investigated the MeCP2 3' UTRs by performing quantitative real-time PCR; the data suggest that altered RNA stability is not a major factor in differential MeCP2 3' UTR usage. In sum, the mechanism(s) of regulated alternative 3'UTR usage of MeCP2 are complex, and insight into these mechanisms will aid our understanding of the factors that influence MeCP2 expression.
Newnham
Catherine M
CM
Department of Physiology and Experimental Medicine, University of Toronto, Toronto, ON, Canada.
Hall-Pogar
Tyra
T
Liang
Songchun
S
Wu
Jing
J
Tian
Bin
B
Hu
Jim
J
Lutz
Carol S
CS
D020413:3' Untranslated Regions / Q000235:genetics; D001483:Base Sequence; D002478:Cells, Cultured; D006367:HeLa Cells; D006801:Humans; D051783:Methyl-CpG-Binding Protein 2 / Q000235:genetics* / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D026723:Polyadenylation / Q000235:genetics; D020871:RNA Stability / Q000502:physiology; D012045:Regulatory Sequences, Nucleic Acid / Q000502:physiology*; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism / Q000502:physiology*; D014162:Transfection
D020413:3' Untranslated Regions; C497392:MECP2 protein, human; D051783:Methyl-CpG-Binding Protein 2; D015534:Trans-Activators
Hum Mutat. 2001 Mar;17(3):183-90
11241840
Nat Genet. 2001 Mar;27(3):322-6
11242117
Nucleic Acids Res. 1997 Jul 1;25(13):2547-61
9185563
Nucleic Acids Res. 1998 Dec 1;26(23):5343-50
9826757
ACS Chem Biol. 2008 Oct 17;3(10):609-17
18817380
Nucleic Acids Res. 2005 Jan 12;33(1):201-12
15647503
Nucleic Acids Res. 2010 May;38(9):2757-74
20044349
Hum Mol Genet. 2000 Oct;9(16):2365-75
11005791
Methods. 2001 Dec;25(4):402-8
11846609
Nucleic Acids Res. 1995 Jul 25;23(14):2614-20
7651822
Prog Nucleic Acid Res Mol Biol. 2002;71:285-389
12102557
Cell. 1992 Jun 12;69(6):905-14
1606614
Nucleic Acids Res. 1998 Jun 15;26(12):2891-8
9611233
Mol Cell Biol. 1999 Jan;19(1):69-77
9858532
Am J Med Genet B Neuropsychiatr Genet. 2004 Jul 1;128B(1):50-3
15211631
Hum Mol Genet. 1999 Jul;8(7):1253-62
10369871
Neurology. 2001 Mar 13;56(5):611-7
11245712
Genomics. 2005 Apr;85(4):441-52
15780747
Psychiatr Genet. 2007 Apr;17(2):113-6
17413451
Hum Genet. 2001 Jan;108(1):43-50
11214906
J Biol Chem. 2002 Nov 8;277(45):42733-40
12200454
Neurology. 2001 Jun 12;56(11):1486-95
11402105
Genes Dev. 1995 Nov 1;9(21):2659-71
7590243
Mol Cell Biol. 2001 Feb;21(4):1228-38
11158309
Nat Genet. 1999 Oct;23(2):185-8
10508514
Biochem J. 2006 Jan 1;393(Pt 1):361-71
16171461
Hum Mol Genet. 2000 May 22;9(9):1377-84
10814719
Genes Dev. 1994 Mar 1;8(5):576-86
7926751
Nat Genet. 2001 Mar;27(3):327-31
11242118
Am J Med Genet B Neuropsychiatr Genet. 2007 Jun 5;144B(4):475-83
17427193
Nucleic Acids Res. 2002 Apr 15;30(8):1842-50
11937639
Am J Hum Genet. 1999 Dec;65(6):1520-9
10577905
Nucleic Acids Res. 2005 May 04;33(8):2565-79
15872218
Methods Mol Biol. 2008;419:171-85
18369983