人類遺傳疾病 第一部份：第八型脊髓小腦共濟失調症之分子遺傳及外遺傳研究 第二部份：台灣兩個Netherton徵候群病患家族之分子遺傳研究

Human Genetic Diseases: PartⅠ： Molecular genetic and epigenetic studies of spinocerebellar ataxia type 8 PartⅡ： Netherton syndrome: molecular analysis of two Taiwanese families

黃淑宜(Shu-Yi Huang)

指導教授 ： 李桂楨 博士

國立臺灣師範大學/理學院/生命科學研究所/碩士(2005年)

摘要

無資料

關鍵字

第八型脊髓小腦共濟失調症 ； 氧化壓力 ； 甲基化 ； Netherton徵候群

並列摘要

PartⅠ:

Spinocerebellar ataxia type 8 (SCA8) is a neurodegenerative disorder characterized by cerebellar dysfunction alone or in combination with other neurological abnormalities. The expansion of 3' CTG trinucleotide repeat on chromosome 13q21 was shown to cause dominantly inherited SCA8. Since first described in 1999, the CTG expansions of the SCA8 gene were found in various familial and sporadic ataxia patients, as well as in patients with psychiatric disorder, Friedreich's ataxia, Parkinson's disease (PD), Alzheimer's disease (AD), and in rare instances in the general population. The SCA8 transcripts are found ubiquitously expressed in various brain tissues and no extended open reading frames are present. Thus the SCA8 transcript was suggested to act as an antisense regulator of the KLHL1, a gene encoding the actin-binding protein. This antisense/sense transcriptional organization is evolutionary conserved in both human and mouse. Previously we assessed repeat sizes at the SCA8 locus and detected abnormal expansions in SCA and PD patients (Wu et al., 2004). In this study, SCA8 repeat size ranges in control subjects and in patients with ataxia, dementia, PD, and other neurological disorders were set up by polymerase chain reaction (PCR)-genotyping and DNA sequencing. A total of 8 subjects with expanded allele were found, including one normal, two ataxia, one dystonia, one parkinsonism(DLBD), and three PD. RT-PCR analysis revealed that both SCA8 and KLHL1 were expressed in lymphoblastoid cells with normal or expanded CTG repeats. Analysis of aberrant methylation by methylation specific PCR assay and restriction enzyme based-methylation assay further revealed differential methylation of the SCA8 and KLHL1 gene exon 1 region. However, CTG repeat length-dependent methylation was not observed. Finally, oxidative stress tolerance of lymphoblastoid cells carrying normal or expanded SCA8 CTG repeats was assessed by quantifying the cell viability and the amount of SOD upon t-butylhydroperoxide (TBH) treatment. The results of no significant difference suggest that cells expressed expanded SCA8 CTG repeats were not more vulnerable to TBH treatment.

Part Ⅱ:

Netherton syndrome (NS) is a severe autosomal recessive skin disorder characterized by congenital ichthyosis, hair shaft abnormalities, and atopic manifestations. NS is caused by deficiency of serine protease inhibitor, resulting in high serine protease activity. So far there is no effective treatment for NS. In year 2000, the gene for NS was mapped to 5q31-q32 and was subsequently identified as serine protease inhibitor Kazal-type 5 (SPINK5). The SPINK5 gene spans a region of 61 kb and is composed of 33 exons. It encodes LEKTI (lympho-epithelial Kazal-type related inhibitor), a predicted serine protease inhibitor highly expressed in thymus and mucous epithelia. The LEKTI protein consists of 1064 amino acids organized into 15 potential inhibitory Kazal-type domains (D1-D15). The pathogenic mutations identified in the SPINK5 gene are predominantly nonsense, predicting marked instability of mutated SPINK5 transcripts and loss of expression of LEKTI. In the study, the molecular lesions of two Taiwanese patients with NS were examined. The entire coding sequence of the SPINK5 gene, flanking intron boundaries, and the proximal promoter region from the two patients were amplified for direct sequencing. Linkage analysis using six flanking microsatellite markers and five single nucleotide polymorphisms (SNPs) in the SPINK5 gene were also performed in patient 270's family to generate haplotypes. Patient 2567 has heterozygous mutations; the maternal allele has T808I (C to T transition in codon 808) and the paternal allele has R790X (C to T transition in codon 790). Patient 270 is homozygous for a novel polymorphism R267Q (G to A transition in codon 267). The R267Q was seen frequently upon screening 200 control chromosomes, with Q allele frequency 0.31. However, the change was not detected in the patient's father. Haplotype analysis revealed that the patient was homozygous for the 5 SNP in the genomic sequence of SPINK5 as well as the flanking (GT)17 and D5S413, in addition to the discrepancy of R267Q. Quantitative real-time PCR analysis further excludes the possibility of small deletion. Thus a gene conversion event may have resulted in the homozygosity for R267Q.

並列關鍵字

SCA8 ； KLHL1 ； epigenetic ； oxidative stress ； methylation ； Netherton Syndrome

參考文獻

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