We have recently reported that the Arg1152-->Gln insulin receptor mutation (QK single mutant) alters a conserved motif (RK motif) immediately next to the key tyrosine phosphorylation sites (Tyr1146, Tyr1150, Tyr1151) of the receptor and constitutively activates its kinase and metabolic signaling. To investigate further the function of the RK motif, we have expressed two additional mutant insulin receptors: a single mutant, in which the second basic residue in the RK motif (Lys1153) was substituted (RA mutant); and a double mutant, in which both the Arg and the Lys residues were replaced with noncharged amino acids (QA mutant). As compared with the transfected wild-type receptors (WT), both the single and the double mutant receptors were normally synthetized and transported to the plasma membrane and bound insulin normally. Whereas the double mutant receptor exhibited preserved insulin-dependent autophosphorylation, kinase activity, and 2-deoxyglucose uptake, all of these functions were grossly impaired in the two single mutant receptors. Two-dimensional analysis of tryptic phosphopeptides from receptor beta-subunits revealed that decreased autophosphorylation of the single mutant receptors mainly involved regulatory Tyr1150,1151 and carboxyl-terminal Tyr1316,1322. At variance with the insulin-stimulated, insulin-independent tyrosine kinase activity toward poly(Glu-Tyr) 4:1 was increased 3-fold in both the double and the single mutants. All mutant receptors induced a 2-fold increase in basal 2-deoxyglucose uptake in NIH-3T3 cells. Treatment of WT transfected cells with 12-O-tetradecanoyl-phorbol-13-acetate or 8-bromo-cAMP increased insulin receptor phosphorylation by 3-fold. No phosphorylation was observed in cells expressing the two single or the double mutant receptor. Consistently, purified preparations of PKC and PKA phosphorylated the WT but not the mutant receptors in vitro. A 17-amino acid synthetic peptide encoding the receptor sequence surrounding the RK motif inhibited phosphorylation of WT insulin receptors by both protein kinases A and C. A mutant peptide in which the RK sequence was replaced by QK (to mimic the mutation in the QK receptor) exhibited no inhibitory effect. Thus, the RK insulin receptor motif is required for insulin receptor phosphorylation by protein kinases C and A and may modulate insulin-independent receptor activity. The RK motif may also have an important structural role in allowing normal insulin regulation of the kinase.

In a prospective study of 33 infants born to hepatitis C virus (HCV)-positive human immunodeficiency virus-negative mothers the vertical transmission of HCV occurred in 6.8%. The evolution of HCV infection in two babies was studied from birth up to 5 or 6 years of age, and the sequencing of the hypervariable region (HVR) of the putative envelope-encoding E2 region of the HCV genome was performed. The HVR1 sequence variability and the different serological profiles during follow-up could reflect the differences in HCV transmission routes, HCV genotypes, and clinical evolution of infection.While the estimated rate of vertical transmission of hepatitis B virus from Hbe antigen-positive mothers is nearly 100% in the absence of immunoprophylactic measures, the rate of hepatitis C virus (HCV) vertical transmission is still very widely debated in the literature (estimates range from 0 to 100%), and to date no specific study has investigated differences among prenatal, perinatal, and postnatal infections (12, 13). Many studies have confirmed that the risk of transmission may be enhanced by coinfection with the human immunodeficiency virus (HIV) (10,15,17).The follow-up of infection in newborns may elucidate some aspects of virus evolution; the HCV variant detected at birth may be considered the starting viral sequence for a particular host, allowing a more reliable analysis of sequence variability over time.This study evaluates the rate of HCV transmission from HCV RNA-positive anti-human immunodeficiency virus (HIV) antibody-negative mothers to their offspring and the clinical evolution of acquired infection. We carried out a longitudinal study of 33 anti-HCV-positive infants for 24 months, and two infected children belonging to this cohort were followed up until 5 or 6 years of age by evaluating several clinical and virological parameters and sequencing hypervariable regions (HVR) of the putative envelope-encoding E2 region of the HCV genome (5,7,8).Longitudinal study. In our study 2,263 anti-HIV antibodynegative pregnant women were screened for anti-HCV antibodies between June 1992 and June 1995; anti-HCV antibody positivity was found in 56 cases (2.4%), and 33 women were enrolled in a prospective longitudinal study of HCV transmission. Their ages ranged between 19 and 42 years (mean age, 28 years); 20 women (60%) had an acknowledged history of intravenous drug use, 1 (3%) was a health care worker with professional exposure (3%), 4 (12%) had an acknowledged history of anti-HCV antibody-positive sexual partners, and 8 (24%) had no risk factor. No woman was in interferon therapy. Only 2 pregnant women were diagnosed as chronic hepatitis sufferers based on histological findings by liver biopsy, and 26 were asymptomatic. Four women breast fed up to 10 to 12 months, and two breast fed up to 30 days. All but 3 of 33 babies were delivered vaginally.Serum samples were stored at Ϫ80°C within 3 h of collection in a day hospital. Testing for anti-HCV antibodies was done by a commercially available third-generation enzyme immuno...

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