Mutant CAG repeats of Huntingtin transcript fold into hairpins, form nuclear foci and are targets for RNA interference.
The CAG repeat expansions that occur in translated regions of specific genes can cause human genetic disorders known as polyglutamine (poly-Q)-triggered diseases. Huntington's disease and spinobulbar muscular atrophy (SBMA) are examples of these diseases in which underlying mutations are localized near other trinucleotide repeats in the huntingtin (HTT) and androgen receptor (AR) genes, respectively. Mutant proteins that contain expanded polyglutamine tracts are well-known triggers of pathogenesis in poly-Q diseases, but a toxic role for mutant transcripts has also been proposed. To gain insight into the structural features of complex triplet repeats of HTT and AR transcripts, we determined their structures in vitro and showed the contribution of neighboring repeats to CAG repeat hairpin formation. We also demonstrated that the expanded transcript is retained in the nucleus of human HD fibroblasts and is colocalized with the MBNL1 protein. This suggests that the CAG repeats in the HTT mRNA adopt ds-like RNA conformations in vivo. The intracellular structure of the CAG repeat region of mutant HTT transcripts was not sufficiently stable to be protected from cleavage by an siRNA targeting the repeats and the silencing efficiency was higher for the mutant transcript than for its normal counterpart.
Laboratory of Cancer Genetics, Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, 61-704 Poznan, Poland.
Mateusz
M
de Mezer
Marzena
M
Wojciechowska
Marek
M
Napierala
Krzysztof
K
Sobczak
Wlodzimierz J
WJ
Krzyzosiak
D001483:Base Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D015536:Down-Regulation; D006801:Humans; D000071058:Huntingtin Protein; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics*; D009687:Nuclear Proteins / Q000235:genetics*; D009690:Nucleic Acid Conformation; D034622:RNA Interference; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000032:analysis; D011944:Receptors, Androgen / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid; D019680:Trinucleotide Repeat Expansion
C086055:HTT protein, human; D000071058:Huntingtin Protein; C493663:MBNL1 protein, human; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011944:Receptors, Androgen
J Mol Biol. 2005 May 13;348(4):883-93
15843020
Cell. 1998 Oct 2;95(1):55-66
9778247
J Biol Chem. 2010 Apr 23;285(17):12755-64
20159983
EMBO J. 2010 Apr 7;29(7):1248-61
20186122
Cell. 1992 Apr 17;69(2):385
1568252
Nat Rev Genet. 2005 Oct;6(10):743-55
16205714
Nat Biotechnol. 2008 May;26(5):578-83
18438400
N Engl J Med. 1986 Nov 13;315(20):1267-76
2877396
Hum Mutat. 2007 May;28(5):451-8
17226796
J Biol Chem. 2005 Feb 4;280(5):3898-910
15533937
Proc Natl Acad Sci U S A. 2003 Jun 10;100(12):7195-200
12782788
J Biol Chem. 2004 Oct 1;279(40):41563-72
15292212
Nucleic Acids Res. 1987 Nov 25;15(22):9109-28
2446263
Pathol Biol (Paris). 2004 Jul;52(6):338-42
15261377
Nucleic Acids Res. 2003 Oct 1;31(19):5469-82
14500809
Methods Enzymol. 1989;180:192-212
2482414
Nature. 1991 Jul 4;352(6330):77-9
2062380
Anal Biochem. 2010 Jul 1;402(1):40-6
20302838
Nucleic Acids Res. 2008 Feb;36(2):640-7
18073195
Nature. 2008 Jun 19;453(7198):1107-11
18449188
Nat Genet. 2007 Oct;39(10):1278-84
17893677
J Biol Chem. 2004 Oct 1;279(40):42230-9
15292246
RNA. 2007 Oct;13(10):1631-40
17684233
Hum Mol Genet. 2001 Sep 1;10(18):1879-87
11555624
J Mol Biol. 2004 Jul 16;340(4):665-79
15223312
Cell. 2007 Jul 13;130(1):101-12
17632058
Nucleic Acids Res. 2005 Jan 19;33(2):451-63
15659577
PLoS One. 2008;3(12):e3968
19092997
J Mol Biol. 1999 May 21;288(5):911-40
10329189
FEBS Lett. 1989 Jun 5;249(2):279-84
2737289
Nucleic Acids Res. 2010 Dec;38(22):8370-6
20702420
Nucleic Acids Res. 2003 Jul 1;31(13):3406-15
12824337
Hum Mutat. 2004 Sep;24(3):236-47
15300851
J Cell Sci. 2005 Jul 1;118(Pt 13):2923-33
15961406
Nat Rev Mol Cell Biol. 2007 Jan;8(1):23-36
17183358
Science. 2000 Sep 8;289(5485):1769-73
10976074
J Biol Chem. 1997 Dec 5;272(49):31079-85
9388259
Science. 2001 Aug 3;293(5531):864-7
11486088
Cell. 1993 Mar 26;72(6):971-83
8458085
Nat Rev Genet. 2005 Oct;6(10):766-73
16136077
Nat Biotechnol. 2009 May;27(5):478-84
19412185
Science. 1997 Sep 26;277(5334):1990-3
9302293
Trends Neurosci. 2004 May;27(5):233-4
15111000