HNF4 and HNF1 as well as a panel of hepatic functions are extinguished and reexpressed in parallel in chromosomally reduced rat hepatoma-human fibroblast hybrids.

Rat hepatoma-human fibroblast hybrids of two independent lineages containing only 8-11 human chromosomes show pleiotropic extinction of thirteen out of fifteen hepatic functions examined. Reexpression of the entire group of functions most often occurs in a block, and except for one discordant subclone, correlates with loss of human chromosome 2. The extinguished cells and their reexpressing derivatives have been examined for the expression of seven liver-enriched transcription factors. C/EBP, LAP, DBP, HNF3, and vHNF1 expression are not systematically extinguished in parallel with the hepatic functions. However, HNF1 and HNF4 show a perfect correlation with phenotype: these factors are expressed only in the cells showing pleiotropic reexpression. Since recent evidence indicates that HNF4 controls HNF1 expression, it can be proposed that the HNF4 gene is the primary target of the pleiotropic extinguisher.

Repression of the lambda pcin promoter by integrative host factor.

The cin-1 mutation creates a new promoter (pcin) in the tR1 region of bacteriophage lambda. The pcin promoter transcribes the cI repressor gene constitutively. lambda cin-1 does not propagate on Escherichia coli mutants lacking the integrative host factor (IHF). lambda cI- cin-1 grows normally in IHF- mutants, indicating that repressor overproduction from pcin blocks lytic growth. The presence of an IHF binding site which overlaps the pcin promoter led us to the hypothesis that IHF functions as a repressor of pcin transcription. We find that the pcin promoter is fivefold more active in a host lacking IHF than in wild-type cells. In vitro studies show that IHF directly inhibits transcription initiation at pcin. Abortive initiation and gel retardation assays demonstrate that IHF interferes with the binding of RNA polymerase to the pcin promoter. RNA polymerase bound in an open promoter complex is resistant to IHF. We propose that IHF binding to the pcin promoter region blocks the binding of RNA polymerase to the promoter, either by covering specific nucleotides or by distorting DNA structure.

Characterization and chromosomal mapping of a genomic clone encoding human alanine:glyoxylate aminotransferase.

We have previously reported the isolation of a genomic clone encoding human liver-specific peroxisomal alanine:glyoxylate aminotransferase (AGT, EC 2.6.1.44), the deficient enzyme in primary hyperoxaluria type 1 (PH1) (P. E. Purdue, Y. Takada, and C. J. Danpure, J. Cell Biol. 111: 2341-2351, 1990). This clone has now been characterized, revealing that the coding sequence is distributed among 11 exons covering 10 kb. The nucleotide sequences of each exon have been determined, confirming that this clone corresponds to previously characterized AGT cDNA (Y. Takada, N. Kaneko, H. Esumi, P. E. Purdue, and C. J. Danpure, Biochem. J. 268: 517-520, 1990). In addition, to provide sequence data for the design of exon-specific PCR primers, the intron sequences immediately flanking each exon have been determined. Furthermore, in an attempt to identify putative transcriptional control sequences we have determined the sequence of 1.25 kb directly upstream of the cDNA 5' end. The results of genomic Southern blotting indicate that human AGT is probably encoded by a single copy gene, and a combination of in situ hybridization and PCR analysis of rodent/human somatic cell hybrids suggests that this gene is located on chromosome 2q36-q37. The gene symbol AGXT has been assigned for this locus.

Prevalence, risk factors, and genotype distribution of hepatitis C virus infection in the general population: a community-based survey in southern Italy.

In 1996 the prevalence, risk factors, and genotype distribution of hepatitis C virus (HCV) infection were assessed in the general population of a town in southern Italy. The sample was selected from the census by a systematic 1:4 sampling procedure. The participation rate was 96.6%. Among the 1,352 subjects enrolled, 195 (14.4%) tested reactive to antibody to HCV (anti-HCV) with enzyme immunoassay (EIA 3). When further tested with recombinant immunoblot assay (RIBA 3), 170 subjects (87.2%) tested positive, 23 subjects (11.8%) had indeterminate results, and 2 subjects (1%) tested negative. Thus, the overall anti-HCV EIA-positive RIBA-confirmed prevalence was 12.6% (170 of 1,352 subjects) and increased from 1.3% in subjects younger than 30 years to 33.1% in those > or =60 years of age. This latter age group accounted for 72.3% of all anti-HCV-positive subjects. Females tested positive more frequently than males (14.1% vs. 10.5%; P < .05). Alanine transaminase (ALT) concentrations were abnormal in only 4.1% (7/170) of anti-HCV EIA-positive RIBA-confirmed subjects. This suggests that ALT screening is not useful in the detection of anti-HCV-positive subjects in a general population. The results of multiple logistic regression analysis showed that an age of less than 45 years, the use of glass syringes, and dental therapy were all independent predictors of anti-HCV positivity. HCV RNA was detected by polymerase chain reaction in 75.9% of the 195 anti-HCV EIA-positive subjects: in 84.7% (144/170) of the RIBA-confirmed subjects; in 17.4% (4/23) tested as RIBA indeterminate; and in neither of the two subjects who tested RIBA negative. HCV type 1b was detected in 75 subjects (50.7%), type 2b in 1 subject (0.7%), type 2c in 66 subjects (44.6%), type 3a in 4 subjects (2.7%), and type 4 in two subjects (1.3%). These figures differ from those of Italian patients with chronic liver disease in whom genotype 2 is more rare. None of the individuals was infected with more than one genotype. The distribution of the two most common HCV viral types (1b and 2c) was not statistically different in terms of mean age, sex, or risk factors and suggests that they may have had a parallel spread in this community. These findings provide one of the highest overall anti-HCV prevalence rates in a general population with a likely cohort effect, i.e., decreased risk of infection along generations. These observations may indicate an epidemic or focus of hepatitis C that occurred several years earlier. The majority of anti-HCV-positive subjects in the oldest age group and with no clinical evidence suggests that HCV infection is a very prolonged and indolent disease.