The other side of immunoglobulin G: suppressor of inflammation.

Immunoglobulin G (IgG) molecules can have two completely opposite functions. On one hand, they induce proinflammatory responses and recruit innate immune effector cells during infection with pathogenic microorganisms or autoimmune disease. On the other hand, intravenous infusion of high doses of pooled IgG molecules from thousands of donors [intravenous IG (IVIG) therapy] represents an efficient anti-inflammatory treatment for many autoimmune diseases. Whereas our understanding of the mechanism of the proinflammatory activity of IgG is quite advanced, we are only at the very beginning to comprehend how the anti-inflammatory activity comes about and what cellular and molecular players are involved in this activity. This review will summarize our current knowledge and focus upon the two major models of either IVIG-mediated competition for IgG-triggered effector functions or IVIG-mediated adjustment of cellular activation thresholds used to explain the mechanism of the anti-inflammatory activity.

The pro and anti-inflammatory activities of immunoglobulin G.

Immunoglobulin G (IgG) molecules are a family of glycoproteins essential for defending the body against invading pathogens. The antibody constant domain is very potent in initiating proinflammatory pathways such as the activation of innate immune effector cells via cellular receptors specific for the antibody constant region (Fc receptors) and the activation of the complement pathway. During autoimmune disease the normally protective antimicrobial function of these molecules is targeted to healthy tissues often with disastrous consequences. Interestingly, one successful anti-inflammatory therapy for many autoimmune diseases is the infusion of high doses of IgG molecules, the so-called intravenous IgG therapy. How one class of molecules can have such opposing functions will be the major focus of this review.

Construction and characterization of bispecific costimulatory molecules containing a minimized CD86 (B7-2) domain and single-chain antibody fragments for tumor targeting.

Efficient T-cell activation requires two signals. The first signal, which confers specificity, is provided by interaction of the T-cell receptor with peptides presented by MHC molecules. One of the second costimulatory signals is induced by binding of B7 proteins on the surface of antigen-presenting cells to CD28 on the T-cell surface. Expression of B7 molecules on tumor cells can result in the activation of tumor specific T lymphocytes and induce protective antitumor immunity. However, at present such gene-therapeutic approaches are limited by the inability to selectively target B7 gene expression to cancer cells. As an alternative approach we exploited recombinant antibody fragments to localize a costimulatory B7 molecule to the surface of tumor cells. We constructed chimeric proteins that contain in a single polypeptide chain a portion of human B7-2 (CD86) genetically fused to single-chain (sc) Fv antibody domains specific for the tumor-associated antigens epidermal growth factor receptor and the closely related ErbB2 receptor tyrosine kinase. A small recombinant fragment of human CD86 was characterized that corresponds to amino acid residues 1-111 (CD86(111)) of the mature protein. CD86(111) produced in the yeast Pichia pastoris and CD86(111) expressed in bacteria was functionally active and displayed specific binding to B7 counter receptors. Bacterially expressed CD86(111)-scFv fusion proteins specifically localized to the respective target antigens on the surface of tumor cells and markedly enhanced the proliferation of primary T cells when bound to immobilized tumor antigen.

DNA structure and flexibility in the sequence-specific binding of papillomavirus E2 proteins.

The papillomavirus E2 proteins are transcriptional regulators that bind to a consensus DNA sequence ACCG NNNN CGGT. Multiple copies of this binding site are found in the viral genomes. The affinities of the naturally occurring binding sites for the E2 proteins are predominantly dependent upon the sequence of the NNNN spacer. The hierarchies of binding site affinities among the sites present in the viral genomes result in differential occupancy during the viral life-cycle. In turn, this differential binding regulates transcription from viral promoters, including those for the oncogenes E6 and E7. Structural and biochemical studies have shown that E2 proteins bend the DNA to which they specifically bind. Atomic resolution structures of complexes of the bovine papillomavirus strain 1 (BPV-1) E2 protein and DNA show that the protein does not contact the spacer DNA. A direct comparison of the binding of the DNA-binding domains of the E2 proteins from BPV-1 and human papillomavirus strain 16 (HPV-16) to a series of binding sites as a function of the sequence of their central spacer and/or the presence of a nick or gap in one strand of the spacer DNA is presented in this paper. The BPV-1 E2 DNA-binding domain is only moderately sensitive to the nature of the central spacer; less than several fold differences in affinity were observed when the DNA sequence of the spacer was varied and/or a nick or gap was introduced. In contrast, the HPV-16 E2 DNA-binding domain binds to sites containing A:T-rich central spacers with significantly increased affinity. The introduction of a nick or gap into the spacer of these high affinity sequences is very detrimental to HPV-16 E2 binding while comparable nicks or gaps have only small effects in the low affinity sequences. These results suggest that the HPV-16 E2 protein recognizes the structure of the DNA spacer and that the mechanism of DNA-sequence specific binding of the homologous HPV-16 E2 and BPV-1 E2 proteins is significantly different.

Intragenic suppressors of induction-deficient TetR mutants: localization and potential mechanism of action.

Eight Tn10 Tet repressor mutants with an induction-deficient phenotype and with primary mutations located at or close to the dimer interface were mutagenized and screened for inducibility in the presence of tetracycline. The second-site suppressors with wild-type-like operator binding activity that were obtained act, except for one, at a distance, suggesting that they contribute to conformational changes in the Tet repressor. Many of these long-range suppressors occur along the dimer interface, indicating that interactions between the monomers play an important role in Tet repressor induction.

Oligo[d(C).(G)] runs exhibit a helical repeat of 11.1 bp in solution and cause slight DNA curvature when properly phased.

We have inserted d(C)10 in a set of DNA fragments with bent segments on both ends, which are rotated with respect to each other by base pair wise increasing insertions. The electrophoretic mobilities on polyacrylamide gels of these DNA fragments were used to identify insertion sizes with cis conformations of the bent ends. These experiments revealed a helical repeat in solution of d(C).d(G) tracts of 11.1 +/- 0.08 bp. The electrophoretic mobilities of ligation ladders with properly phased d(C)5 and d(C)16 runs demonstrate a small but clearly detectable curvature of these fragments.