Cruciform DNA, a non-double helix form of DNA, can be generated as an intermediate in genetic recombination as well as from palindromic sequences under the effect of supercoiling. Eukaryotic cells are equipped with a DNA-binding protein that selectively recognizes cruciform DNA. Biochemical and immunological data showed that this protein is HMG1, an evolutionarily conserved, essential, and abundant component of the nucleus. The interaction with a ubiquitous protein points to a critical role for cruciform DNA conformations.

NF1‐like proteins play a role in transcription of liver‐specific genes. A DNA‐binding protein, recognizing half of the canonical NF1 binding site (TGGCA) present on the human albumin and retinol‐binding protein genes, has been purified from rat liver. Several peptides deriving from a tryptic digest of the purified protein were sequenced and the sequence was used to synthesize specific oligonucleotides. Two overlapping cDNA clones were obtained from a rat‐liver cDNA library; their sequence reveals an open reading frame coding for 505 amino acids, including all the peptides sequenced from the purified protein. The DNA‐binding domain, most likely located within the first 250 amino acids, is highly homologous to the sequence of CTF/NF1 purified from HeLa cells. Northern analysis reveals several mRNA species present in different combinations in various rat tissues.

The helical cytokine interleukin (IL) 6 and its specific binding subunit IL‐6R alpha form a 1:1 complex which, by promoting homodimerization of the signalling subunit gp130 on the surface of target cells, triggers intracellular responses. We expressed differently tagged forms of gp130 and used them in solution‐phase binding assays to show that the soluble extracellular domains of gp130 undergo dimerization in the absence of membranes. In vitro receptor assembly reactions were also performed in the presence of two sets of IL‐6 variants carrying amino acid substitutions in two distinct areas of the cytokine surface (site 2, comprising exposed residues in the A and C helices, and site 3, in the terminal part of the CD loop). The binding affinity to IL‐6R alpha of these variants is normal but their biological activity is poor or absent. We demonstrate here that both the site 2 and site 3 IL‐6 variants complexed with IL‐6R alpha bind a single gp130 molecule but are unable to dimerize it, whereas the combined site 2/3 variants lose the ability to interact with gp130. The binding properties of these variants in vitro, and the result of using a neutralizing monoclonal antibody directed against site 3, lead to the conclusion that gp130 dimer is formed through direct binding at two independent and differently oriented sites on IL‐6. Immunoprecipitation experiments further reveal that the fully assembled receptor complex is composed of two IL‐6, two IL‐6R alpha and two gp130 molecules. We propose here a model representing the IL‐6 receptor complex as hexameric, which might be common to other helical cytokines.

Receptor subunits for the neurocytokine ciliary neurotrophic factor (CNTF) share sequence similarity with the receptor for leptin, an adipocyte-derived cytokine involved in body weight homeostasis. We report here that CNTF and leptin activate a similar pattern of STAT factors in neuronal cells, and that mRNAs for CNTF receptor subunits, similarly to the mRNA of leptin receptor, are localized in mouse hypothalamic nuclei involved in the regulation of energy balance. Systemic administration of CNTF or leptin led to rapid induction of the tis-11 primary response gene in the arcuate nucleus, suggesting that both cytokines can signal to hypothalamic satiety centers. Consistent with this idea, CNTF treatment of ob͞ob mice, which lack functional leptin, was found to reduce the adiposity, hyperphagia, and hyperinsulinemia associated with leptin deficiency. Unlike leptin, CNTF also reduced obesity-related phenotypes in db͞db mice, which lack functional leptin receptor, and in mice with diet-induced obesity, which are partially resistant to the actions of leptin. The identification of a cytokine-mediated anti-obesity mechanism that acts independently of the leptin system may help to develop strategies for the treatment of obesity associated with leptin resistance.

The human scavenger class B type 1 receptor (SR-B1/Cla1) was identified as a putative receptor for hepatitis C virus (HCV) because it binds to soluble recombinant HCV envelope glycoprotein E2 (sE2). High-density lipoprotein (HDL), a natural SR-B1 ligand, was shown to increase the in vitro infectivity of retroviral pseudoparticles bearing HCV envelope glycoproteins and of cell culture-derived HCV (HCVcc), suggesting that SR-B1 promotes viral entry in an HDL-dependent manner. To determine whether SR-B1 participates directly in HCV infection or facilitates HCV entry through lipoprotein uptake, we generated a panel of monoclonal antibodies (MAbs) against native human SR-B1. Two of them, 3D5 and C167, bound to conformationdependent SR-B1 determinants and inhibited the interaction of sE2 with SR-B1. These antibodies efficiently blocked HCVcc infection of Huh-7.5 hepatoma cells in a dose-dependent manner. To examine the role of HDL in SR-B1-mediated HCVcc infection, we set up conditions for HCVcc production and infection in serum-free medium. HCVcc efficiently infected Huh-7.5 cells in the absence of serum lipoproteins, and addition of HDL led to a twofold increase in infectivity. However, the HDL-induced enhancement of infection had no impact on the neutralization potency of MAb C167, despite its ability to inhibit both HDL binding to cells and SR-B1-mediated lipid transfer. Of note, MAb C167 also potently blocked Huh-7.5 infection by an HCV strain recovered from HCVcc-infected chimpanzees. These results demonstrate that SR-B1 is essential for infection with HCV produced in vitro and in vivo and suggest the possible use of anti-SR-B1 antibodies as therapeutic agents.Hepatitis C virus (HCV) is the major etiological agent of both community-acquired and posttransfusion non-A, non-B viral hepatitis. Approximately 80% of infected patients develop chronic hepatitis, among which 20% to 30% progress to liver cirrhosis and end-stage liver disease. Chronic infection correlates with an increased risk of hepatocellular carcinoma. Currently available therapies are limited to administration of pegylated alpha interferon in combination with ribavirin (27). Such treatment is expensive, is often unsuccessful, and carries the risk of significant side effects. Consequently, the development of novel therapeutic approaches against HCV remains a high-priority goal.HCV is an enveloped virus of the family Flaviviridae whose viral genome is a single-stranded, positive-sense RNA of approximately 9.6 kb that encodes a single polyprotein of 3,010 to 3,033 amino acids that is cleaved into nine mature proteins by a combination of host and viral peptidases (24). The predicted structural components comprise the core (C) (ϳ21 kDa) and two heavily N-glycosylated envelope glycoproteins, E1 (ϳ31 kDa) and E2 (ϳ70 kDa). Both E1 and E2 are believed to be type I transmembrane proteins, with N-terminal ectodomains and C-terminal hydrophobic anchors. HCV entry into target cells occurs after attachment to specific cellular receptors via its surface glycoprotei...