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We have investigated the molecular lesions of T-protein deficiency causing typical or atypical nonketotic hyperglycinemia (NKH) in two unrelated pedigrees. A patient with typical NKH was identified as being homozygous for a missense mutation in the T-protein gene, a G-to-A transition leading to a Gly-to-Asp substitution at amino acid 269 (G269D). Sibling patients of a second family with atypical NKH had two different missense mutations in the T-protein gene (compound heterozygote), a G-to-A transition leading to a Gly-to-Arg substitution at amino acid 47 (G47R) in one allele, and a G-to-A transition leading to an Arg-to-His substitution at amino acid 320 (R320H) in the other allele. Gly 269 is conserved in T-proteins of various species, even in E. coli, whereas Gly 47 and Arg 320 are replaced by Ala and Leu, respectively, in E. coli. The mutation occurring in more conservative amino acid residues thus results in more deleterious damage to the T-protein, and gives the severe clinical phenotype, viz., typical NKH.

Direct sequencing of the coding region of the cystathionine β-synthase (CBS) gene in two homocystinuric patients revealed the presence of two novel missense mutations. The first mutation, a 1111G → A transition, resulted in the substitution of the evolutionary conserved valine-371 by a methionine residue (V371M) and created a new NlaIII restriction site. The second mutation, a G→A transition at base-pair 494, resulted in an amino acid change from cysteine to tyrosine (C165Y) and abolished a BsoFI restriction site. Both mutations were found in a compound heterozygous state with the previously described 833T → C transition.

Southern blot analysis of the iduronate sulfatase (IDS) gene in 52 unrelated Japanese patients with mucopolysaccharidosis type II was carried out using a cDNA probe, and mutations in 13 patients (25%) were identified. Of these, 3 had partial gene deletions (in 2 the normal 9.4-kb fragment was absent and in 1 the normal 7.4-kb fragment was absent, as determined by Southern blot analysis using EcoRI-digested DNA, respectively), 2 had gene insertions (in 1 there was a unique 11.2kb fragment and in the other there was a unique 5kb fragment, determined by Southern blot analysis using EcoRI digested DNA), and 8 had rearrangements (in 6 the normal 9.4kb and 7.0kb fragments were absent and a unique 11.2kb fragment was present; in the remaining 2 patients there were different rearrangements). In these 13 patients, the similar Southern blot patterns were indicative of structural alterations of the IDS gene, as revealed when their DNA was digested with HindIII or PstI and probed with IDS cDNA. All patients with these structural alterations were in a clinically severe state, except for 1 with an intermediate clinical phenotype. Our analyses of four families among those of the 13 patients revealed that all four mothers were carriers. The detection of structural abnormalities led to a precise identification of Hunter heterozygotes and revealed one de novo rearrangement in a germ cell of one of the maternal grandparents.

The distribution of Gc subtypes in a sample of the Chinese population of Hong Kong was studied using isoelectric focusing followed by immunofixation. A sensitive modification of this technique is described. Nine distinct phenotypes were observed which appear to result from the three common alleles Gc IF, Gc IS, and Gc 2, which are found in most populations. The respective gene frequencies were 0.494, 0.258, and 0.247. In addition, two rare phenotypes were observed which appear to be due to a rare allele tentatively identified as Gc 2G2.

Human claudin-1 is an integral protein component of tight junctions, a structure controlling cell-to-cell adhesion and, consequently, regulating paracellular and transcellular transport of solutes across human epithelia and endothelia. Recently, a claudin-1 (CLDN1) cDNA has been isolated from human mammary epithelial cells (HMECs). CLDN1 expression in HMECs, in contrast to low or undetectable levels of expression in a number of breast tumors and breast cancer cell lines, points to CLDN1 as a possible tumor-suppressor gene. In order to evaluate the CLDN-1 gene in sporadic and hereditary breast cancer, we have characterized its genomic organization and have screened the four coding exons for somatic mutations in 96 sporadic breast carcinomas and for germline mutations in 93 breast cancer patients with a strong family history of breast cancer. In addition, we have compared the 5'-upstream sequences of the human and murine CLDN1 genes to identify putative promoter sequences and have examined both the promoter and coding regions of the human gene in the breast cancer cell lines showing decreased CLDN1 expression. In the sporadic tumors and hereditary breast cancer patients, we have found no evidence to support the involvement of aberrant CLDN1 in breast tumorigenesis. Likewise, in the breast cancer cell lines, no genetic alterations in the promoter or coding sequences have been identified that would explain the loss of CLDN1 expression. Other regulatory or epigenetic factors may be involved in the down-regulation of this gene during breast cancer development.

A syndrome due to 3:1 meiotic segregation of balanced 11/22 translocation is defined from nine personally observed patients and 22 cases from the literature with apparently the same aberration. Frequent findings include a characteristic face with deepset eyes, flat nose, prominent upper lip, receding mandible and preauricular pits or tags, male genital hypoplasia, anal atresia or other anomalies of the anus, cleft palate, and congenital heart defect. Less frequent are severe reduction of the auricles, an additional pair of ribs, and hypoplasia of the diaphragm. Perinatal mortality is high. Growth is usually and psychomotor development is invariably and severely delayed. Balanced 11/22 translocations are apparently disproportionally frequent; as the balanced rearrangement is not easy to detect, it is important to be aware of it at the family investigation of cases with extra chromosomes similar to a No.22 or 22q-.The unbalanced products are most probably trisomic for both a segment of 22 (22q-) and a distal segment of 11q; the exact determination of the breakpoints is not possible at present due to the similar banding characteristics of the two segments involved in the translocation.

Utilizing the 5-bromo-deoxyuridine (BrdU) incorporation technique, we have recently studied the DNA replication kinetics in a dicentric X chromosome, formed by long arm-to-long arm fusion at band q23, from a 16-year-old black female with primary amenorrhea. The patient has a karyotype 45,X/46,X,dic(X)(q23). In the buccal smear the presence of X chromatin was found in 33% of the cells examined. The Barr bodies are large and 21% of them are bipartite. DNA replication studies were performed on the patient's lymphocytes by the thymidine pulse (T-pulse) method and confirmed comparatively by the BrdU pulse (B-pulse) method. The results indicate that the dicentric X chromosome is always late-replicating. The replication pattern is symmetric on both sides of the breakpoint and the replication sequence is, in order, p11, p22, q1(1–3), q22, q23, p21, and q21. This finding is comparable to those of other investigators and supports the theory that there exist two inactivation centers in the dicentric X chromosome, located on or near the q21 band.

Chromosome counts and a search for structural abnormalities were performed in cultured leukocytes from 60 elderly subjects (ages 62–96) and 60 controls (ages 10–13). A total of 3900 cells were examined, 3600 in 3-day cultures and 300 in cultivations maintained for 2 days. Our results and those obtained by other investigators indicate clearly that there is an increase in the level of aneuploidy and of structural abnormalities in the blood of aged persons. There is an excess of missing C-group chromosomes in elderly females, but no consistent preferential loss was observed among the males. In contrast with two previous studies, we found a significant decrease of aneuploid cells in 3-day cultures, as compared to those cultivated for 2 days only.

Isoelectric focusing (IEF) in polyacrylamide gels has been used to study the isozymes of human galactose-1-phosphate uridylyltransferase (GALT) in erythrocytes and fibroblasts. In addition to the usefulness of IEF in differentiating normal, Duarte variant, and galactosemic homozygotes and heterozygotes, the ability of IEF to distinguish the residual GALT activity in two different galactosemic fibroblast lines and in revertants from them is demonstrated.