The Genom of Homo sapiens.pdf

1. The levels of α-synuclein protein may be critical to thedevelopment of PD; even small changes in α-synuα-SYNUCLEINEXPRESSION AND PD 413Thus, the effect of different alleles on promoter strengthin the luciferase assay was not linear with respect to repeatlength. Deletion analysis of the NACP-Rep1 locusand surrounding DNA suggested that two domains flankingthe repeat interact to enhance expression while the repeatitself modulates this interaction to a greater or lesserextent depending on which allele is present at the NACP-Rep1 locus (Chiba-Falek and Nussbaum 2001).We then examined the effect on SNCA promoter activityof sequence variation within a single size “1” allele.Two intra-allelic variants, defined by y = 10 z = 8 versusy = 11 and z = 7 in the segment (TC) y (TA)z within theNACP = Rep1 repeat were studied. We found only a verysmall, insignificant difference in luciferase expressionlevels (Chiba-Falek et al. 2003). These findings obtainedusing the luciferase reporter system in human neuroblastomacells imply that the overall length of the NACP-Rep1allele plays the main role in the transcriptional regulationby the NACP-Rep1 element, whereas the intra-size variationhas only a minor contribution to the function of theNACP-Rep1 element as a transcriptional regulator.THE MOUSE NACP-Rep1 REGIONExamination of the mouse sequence reveals a complexrepeat similar to the human NACP-Rep1, located ~6.1 kbupstream of the transcriptional start site of Snca (Touchmanet al. 2001). The human and mouse repeats are only40% identical but contain similar dinucleotide elements,although the human element contains a CA dinucleotidenot present in the mouse element. DNAs from 22 inbredmouse strains derived from Mus musculus musculus, twofrom M. musculus subspecies (CAST/Ei and MOLG/Dn),and one from the species Mus spretus (SPRET/Ei) wereexamined by PCR for polymorphisms at this complexdinucleotide repeat (Touchman et al. 2001). The M. musculus-derivedstrains were not polymorphic; all can bedenoted as (CT) 8 N 2 (AT) 9 N 5 (GT) 4 N 8 (GT) 3 . TheCAST/Ei, MOLG/Dn, and SPRET/Ei, however, do differin size. The CAST/Ei and MOLG/Dn sequences differfrom the other inbred musculus strains only in the size ofthe AT repeat, while showing the identical sequence andspacing for the other dinucleotide elements of the complexrepeat. The CAST/Ei product contains (AT) 29 insteadof (AT) 9 , whereas that from MOLG/Dn has (AT) 22 .SPRET/Ei shows a more complex polymorphism that canbe denoted as (CT) 13 (AT) 35 N 9 (GT) 5 N 8 (GT) 3 (Touchmanet al. 2001). It will be of great interest to study the levelsof α-synuclein mRNA in the different mouse strains thatwere shown to carry variant alleles at the NACP-Rep1site and to correlate the α-synuclein expression level tothe composition of the mouse NACP-Rep1 allele.OTHER GENES HAVING A FUNCTIONALPOLYMORPHIC MICROSATELLITEIN THEIR PROMOTER REGIONSSeveral studies in the past have implicated differentlengthdinucleotide repeats at promoter regions in regulatingvariable transcription activity. Using reporter geneexpression systems, the expression of several genes wasshown to be regulated by polymorphic dinucleotide repeatsin their 5´-flanking region, and some alleles of thesepolymorphic repeat sequences were shown to enhancetheir expression. Some of these studies are listed below.1. The dinucleotide repeat (CA) l N(CG) m (CA) n upstreamof the human COL1A2 has an enhancing activity on genetranscription, and that variation in the number of repetitionsmay be responsible for the difference in the transcriptionactivity of the gene (Akai et al. 1999).2. Variation in the length of a (CA) repeat element in thepromoter of the human MMP-9 gene was shown tomodulate its promoter activity (Peters et al. 1999; Shimajiriet al. 1999).3. Variants of the PAX-6 polymorphic dinucleotide repeat(AC) m (AG) n were shown to have different transcriptionalefficiencies and to drive variable mRNA expressionlevels in human brain (Okladnova et al. 1998).4. Four alleles in the promoter of the human NRAMP1gene consist of T(GT) x AC(GT) y AC(GT) z G and differin their ability to drive gene expression (Searle andBlackwell 1999).5. A (GT) n dinucleotide repeat upstream of the humanHO-1 gene shows length polymorphism, which modulatesthe level of transcription (Yamada et al. 2000).6. Certain alleles at the (CA) n polymorphic site located2.1 kb upstream of the human AR2 gene lead to significantlyhigher expression level (Ikegishi et al. 1999).7. A functional polymorphic dinucleotide repeat(TCTCT(TC) n ) upstream of the translational startcodon of human HMGA2 was found to regulatestrongly the human HMGA2 promoter with an activationpattern that correlates with its TC-repeat length(Borrmann et al. 2003).The mechanisms by which dinucleotide repeats mightaffect gene expression in cis are largely unknown. In onecase, however, a dinucleotide-binding protein, angiogenin,has been implicated in regulation of expression ofthe human rRNA gene. Angiogenin binds (CT) n repeatsspecifically, in a length-dependent manner, and the affinityof its binding increases for longer (CT) repeats. Moreover,the CT repeats exhibit angiogenin-dependent promoteractivity in a luciferase reporter system, and thelevel of the promoter activity depends on the numbers ofthe CTs. This study suggested that angiogenin may playa role in regulating expression of genes containing CT repeatsin their 5´-flanking regions, which are common inthe eukaryotic genome (Xu et al. 2003).It is important to note that in most of the genes studied,the combined dinucleotide repeat was at a distance of ≤2kb upstream of the transcriptional start site. The complexdinucleotide repeat, NACP-Rep1, is unusual in that it caninfluence gene expression from a much longer distance of~10 kb, such as might be seen with a long-range enhanceror a locus control region (LCR).CONCLUSIONSWe propose the following hypotheses: