CAG Repeats of KCNN3 Gene in the Patients with Schizophrenia / 신경정신의학

OBJECTS:We investigated a possible association between the polymorphic trinucleotide repeat(TNR) expansion in neuronal potassium channel gene KCNN3 and schizophrenia. METHODS: CAG/CTG repeat distribution in KCNN3, CTG18.1 and ERDA1 was examined and the copy number of ligation product in repeat expansion detection(RED) was measured in Korean patients with schizophrenia(n=245) and ethnically matched healthy controls(n=116). RESULTS: Longer alleles in the KCNN3 gene were over-represented in patients. The frequency of alleles with CAG repeats longer than 19 copy in the KCNN3 gene was higher in the patients with schizophrenia as compared to controls(73.3% vs. 65.1%;p=0.029, Fisher's exact test). And this difference was more prominent in schizophrenic patients with familial background(p=0.03, Fisher's exact test). We found no difference in the frequency of longer alleles between negative and positive subtypes of schizophrenia. Ligation product size in RED and alleles with CAG repeat number in the CTG18.1 gene was not increased in the patients. The copy number of ligation product in RED was highly correlated with CAG/CTG copies of ERDA1 in the patient group(r=0.45, p<0.001) as well as in the control group(r=0.44, p<0.001). However, CAG repeat length in the KCNN3 gene was not correlated with ERDA1 score. CONCLUSIONS: Our results support the hypothesis that the longer allele of KCNN3 may be considered as a candidate gene for schizophrenia, especially in the case with familial background. And the RED assay results was affected by the CAG copy number of ERDA1.

A Molecular Genetic Study with EcoRII Restriction Enzyme on the Steroidogenic Acute Regulatory Protein (StAR) Gene / 대한소아내분비학회지

The molecular defect of congenital lipoid adrenal hyperplasia has been discovered to be in the transport of cholesterol into mitochondria due to defective regulatory protein called "Steroidogenic Acute Regulatory Protein (StAR)", while the enzyme P450scc itself is normal. This study with EcoRII restriction enzyme aimed at elucidating more conveniently the molecular defect in the StAR gene. The genomic DNAs were extracted from their peripheral blood. We amplified the exon 7, hot spot, of the StAR gene with 1 set of primers by Polymerase Chain Reaction (PCR). Subsequently, a PCR product corresponding to target sequence (~437 bps) from the patient and her father have been sequenced by automatic sequence analyzer. The PCR-RFLP (Restriction Fragment Length Polymorphism) analysis after restriction digestion with EcoRII restriction enzyme was also performed on 12% polyacrylamide gel electrophoresis. The mutation was identified in the exon 7 of the StAR gene, substituting C for T at codon 258, consequently replacing glutamine by stop codon. This mutation alters EcoRII restriction site. In addition, we obtained the good result of PCR-RFLP (Restriction Fragment Length Polymorphism) analysis on 12% polyacrylamide gel electrophoresis. Therefore, the PCR-RFLP (Restriction Fragment Length Polymorphism) analysis with EcoRII restriction enzyme can be easily utilized to screen carrier, diagnose the patient prenatally or postnatally.

CAG Repeat Expansions in the Patients with Mood Disorder / 신경정신의학

OBJECTIVES: The genetic facotrs have been suggested for the etiology of mood disorders but the mode of inheritance is complex. Increased severity and an earlier onset of the bipolar and major depressive disorder over generations within families(Anticipation) were reported. In order to test the hypothesis that trinucleotide repeat expansions underlie the genetic basis of Bipolar and major depressive disorders, we have analyzed the extent of CAG reapeats in genomic DNA from mood disorder patients. METHODS: 55 bipolar disorder, 67 major depressive disorder patients were recruited according to the DSM-III-R criteria. 89 normal controls were recruited from the medical personnel, students and the visitors to the health services center who had no history of psychiatric illness and show normal profile of MMPI. The genomic DNA of patients and controls was analyzed by use of the(CTG) 17 oligonucleotide and the repeat expansion detection(RED) method. The Mann-Whitney U test was used to compare the distribution of the number of CAG repeats among the groups. RESULTS: when the bipolar disorder, major depressive disorder patients were compared with the control group, no significant differences were observed. CONCLUSION: Our results do not support the hypothesis that expanding CAG repeats are causing the observed genetic anticipation in bipolar disorders and major depressive disorders.

The Molecular Genetic Study Using Automatic Sequence Analyzer on the Steroidogenic Acute Relulatory Protein(StAR) Gene / 대한소아내분비학회지

PURPOSE:The lesion of Congenital Lipoid Adrenal Hyperplasia has been suggested to be in the 1st step of steroidogenesis of conversion of cholesterol to pregnenolone by P450scc. In 1995, however, the molecular defect of this disease has been located in the transport of cholesterol into mitochondria due to defective regulatory protein called Steroidogenic Acute Regulatory Protein(StAR), while the enzyme P450scc itself is normal. This genetic study using automatic sequence analyzer aimed at elucidating the molecular defect in the StAR gene of the two patients. METHODS:This study was performed on the two patients of Congenital Lipoid Adrenal Hyperplasia. Both children were phenotypically females. However, one turned out to have a karyotype of 46, XY, the other 46, XX. Genomic DNAs were extracted from their peripheral blood. We amplified the last exon, hot spot, of the StAR gene using 1 set of primer, S4, 5'-CCT GGC AGC CTG TTT GTG ATA G-3' AS4, 5'-CCT CAT GTC ATA GCT AAT CAG TG-3'. Subsequently, one PCR product have been directly sequenced by dideoxy termination method, and also the other products(patient's and her father's) have been sequenced by automatic sequence analyzer. RESULTS:The mutation was identified in the last exon of the StAR gene, substituting T for A at codon 258, replacing glutamine by stop codon in the two unrelated Korean patients with congenital lipoid adrenal hyperplasia. One patient were found to be homozygote, but the other to be heterozygote for the mutation. CONCLUSIONS:These results indicate that Korean children with congenital lipoid adrenal hyperplasia may be genetically identical as in Japanese. But, we discovered that the hot spot, codon 258, are not always homozygote. We want to emphasize the different point, and to say that we did experiece the automatic sequence analyzer successfully.

A study of trinucleotide repeat expansions in myotonic dystrophy

PURPOSE: The trinucleotide repeat expansion in the 3' untanslated resion of the gene is known to be the cause of myotonic dystrophy which is one of most common neurodegenerative disorder manifested by myotonia, cataract, mental retardation and even respiratory distress in neonates. The hereditary pattern of myotonic dystrophy shows more severe symptoms and shows earlier onset with successive generations and congenital cases, the most severe form of myotonic dystrophy,. Occurs by maternal transmission. This genetic transmission mode does not follow Mendelian genetic trait. To find the molecular genetic abnormalities of Korean myotonic dystrophy patients, we investigated the general distribution of myotonic dystrophy alleles and compared the results with referred patients. METHODS: During an 8 month study, from June 1995 to February 1996, 5 patients were referred with presumed diagnosis of myotonicdystrophy. Among these patients, four cases were confirmed to have the disease by clinical and electrophysiological findings. We included family members of the studied probands and 50 normal blood donor DNAs were included as controls. The DNAs of the enrolled cases were evaluated by Southern blot. Subsequently, copy numbers of the repeats were determined using PCR amplification. RESULTS: (1) Two peaks were found in the distribution of trinucleotide repeats in the normal Korean population. One peak had 5 copies and the other had 11 to 13 copies. The highest number of copies was 27. (2) Of the referred cases, 4 pedigrees showed typical expanded repeats. (3) The minimum expanded copy number was 55 and we were able to detect the expanded band only by PCR in 2 cases. In other cases, expaded bands were visible by Southern blotting. (4) There were trend of earlier onset of the disease, progressive worsening symptoms and larger expanded bands with successive generations. CONCLUSION: We established the methodology for myotonic dystrophy DNA diagnosis using Southern blot and PCR amplification based on the normal Korean allele distribution. These methods might be useful in genetic counselling and detection of minimally affected myotonic dystrophy patients.