By using a Reynolds-averaged two-dimensional computation of a turbulent flow over an airfoil at post-stall angles of attack, we show that the massively separated and disordered unsteady flow can be effectively controlled by periodic blowing-suction near the leading edge with low-level power input. This unsteady forcing can modulate the evolution of the separated shear layer to promote the formation of concentrated lifting vortices, which in turn interact with trailing-edge vortices in a favourable manner and thereby alter the global deep-stall flow field. In a certain range of poststall angles of attack and forcing frequencies, the unforced random separated flow can become periodic or quasi-periodic, associated with a significant lift enhancement. This opens a promising possibility for flight beyond the static stall to a much higher angle of attack. The same local control also leads, in some situations, to a reduction of drag. On a part of the airfoil the pressure fluctuation is suppressed as well, which would be beneficial for high-α buffet control. The computations are in qualitative agreement with several recent post-stall flow control experiments. The physical mechanisms responsible for post-stall flow control, as observed from the numerical data, are explored in terms of nonlinear mode competition and resonance, as well as vortex dynamics. The leading-edge shear layer and vortex shedding from the trailing edge are two basic constituents of unsteady post-stall flow and its control. Since the former has a rich spectrum of response to various disturbances, in a quite wide range the natural frequency of both constituents can shift and lock-in to the forcing frequency or its harmonics. Thus, most of the separated flow becomes resonant, associated with much more organized flow patterns. During this nonlinear process the coalescence of small vortices from the disturbed leading-edge shear layer is enhanced, causing a stronger entrainment and hence a stronger lifting vortex. Meanwhile, the unfavourable trailing-edge vortex is pushed downstream. The wake pattern also has a corresponding change: the shed vortices of alternate signs tend to be aligned, forming a train of close vortex couples with stronger downwash, rather than a Kármán street.

The production of high-affinity protective antibodies requires somatic hypermutation (SHM) of the antibody variable (V)-region genes. SHM is characterized by a high frequency of point mutations that occur only during the centroblast stage of B-cell differentiation. Activation-induced cytidine deaminase (AID), which is expressed specifically in germinal-centre centroblasts, is required for this process, but its exact role is unknown. Here we show that AID is required for SHM in the centroblast-like Ramos cells, and that expression of AID is sufficient to induce SHM in hybridoma cells, which represent a later stage of B-cell differentiation that does not normally undergo SHM. In one hybridoma, mutations were exclusively in G*C base pairs that were mostly within RGYW or WRCY motifs, suggesting that AID has primary responsibility for mutations at these nucleotides. The activation of SHM in hybridomas indicates that AID does not require other centroblast-specific cofactors to induce SHM, suggesting either that it functions alone or that the factors it requires are expressed at other stages of B-cell differentiation.

Affinity for DNA and cross-reactivity with renal antigens are associated with enhanced renal pathogenicity of lupus autoantibodies. In addition, certain IgG subclasses are enriched in nephritic kidneys, suggesting that isotype may determine the outcome of antibody binding to renal antigens. To investigate if the isotype of DNA antibodies affects renal pathogenicity by influencing antigen binding, we derived IgM, IgG1, IgG2b and IgG2a forms of the PL9–11 antibody (IgG3 anti-DNA) by in vitro class switching or PCR cloning. The affinity and specificity of PL9–11 antibodies for nuclear and renal antigens were analyzed using ELISA, Western blotting, surface plasmon resonance (SPR), binding to mesangial cells, and glomerular proteome arrays. Renal deposition and pathogenicity were assayed in mice injected with PL9–11 hybridomas. We found that PL9–11 and its isotype-switched variants had differential binding to DNA and chromatin (IgG3 > IgG2a > IgG1 > IgG2b > IgM) by direct and competition ELISA, and SPR. In contrast, in binding to laminin and collagen IV the IgG2a isotype actually had the highest affinity. Differences in affinity of PL9–11 antibodies for renal antigens were mirrored in analysis of specificity for glomeruli, and were associated with significant differences in renal pathogenicity in vivo and survival. Our novel findings indicate that the constant region plays an important role in the nephritogenicity of antibodies to DNA by affecting immunoglobulin affinity and specificity. Increased binding to multiple glomerular and/or nuclear antigens may contribute to the renal pathogenicity of anti-DNA antibodies of the IgG2a and IgG3 isotype. Finally, class switch recombination may be another mechanism by which B cell autoreactivity is generated.

After encounter with antigen, the antibody repertoire is shaped by somatic hypermutation (SHM), which leads to an increase in the affinity of antibodies for the antigen, and class-switch recombination (CSR), which results in a change in the effector function of antibodies. Both SHM and CSR are initiated by activation-induced cytidine deaminase (AID), which deaminates deoxycytidine to deoxyuridine in single-stranded DNA (ssDNA). The precise mechanism responsible for the formation of ssDNA in V regions undergoing SHM has yet to be experimentally established. In this study, we searched for ssDNA in mutating V regions in which DNA–protein complexes were preserved in the context of chromatin in human B cell lines and in primary mouse B cells. We found that V regions that undergo SHM were enriched in short patches of ssDNA, rather than R loops, on both the coding and noncoding strands. Detection of these patches depended on the presence of DNA-associated proteins and required active transcription. Consistent with this, we found that both DNA strands in the V region were transcribed. We conclude that regions of DNA that are targets of SHM assemble protein–DNA complexes in which ssDNA is exposed, making it accessible to AID.

Antibodies to the Cryptococcus neoformans capsular glucuronoxylomannan (GXM) form the basis of two potential therapeutic intervention strategies, i.e., conjugate vaccines and passive antibody therapy. To better understand the molecular basis of the antibody response, the heavy-and light-chain immunoglobulin variable region (V. and VL, respectively) sequences of seven monoclonal antibodies (MAbs) to GXM were determined. Rabbit anti-idiotypic serum was made to the previously characterized murine MAb 2H1 and used to study MAb 2H1 idiotype expression in other GXM-binding MAbs and immune sera. MAb El originated from a C3H/HeJ mouse immunized with C. neoformans serotype A polysaccharide. MAbs 471, 1255, 339, 3C2, 386, and 302 originated from BALB/c mice immunized with polysaccharide of serotypes A, A, B, C, D, and D, respectively, conjugated to sheep erythrocytes. In the El, VH uses V,1 from the T15 gene family and JH3 and has a D segment of three amino acids, and the VL uses a V,Ser-like gene family element and JK5. In MAbs 471 and 3C2, the VH uses VH7183-like gene family elements and JH2 and has D segments of seven amino acids, and the VL uses V,5.1 and J,1. In MAbs 1255 and 339, the V. uses VH10-like gene elements and JH4 and has six codon D segments, and the VL uses a V.21-like gene element and J.5. In MAbs 302 and 386, respectively, the VH uses V,GAM-like gene elements and JH2 and JH3 and has six and four codon D segments, and VL uses V,4/5-like gene elements and J,1. VH usage, MAb 2H1 idiotype expression, and fine specificity mapping define a minimum of three GXM epitopes which elicit protective antibodies. The results confinn that the antibody response is highly restricted, suggest a close relationship between molecular structure and serological properties, and provide insight into protein structural motifs important for GXM binding. Cryptococcus neofornans is an encapsulated fungus that causes life-threatening meningoencephalitis in up to 9% of patients with AIDS (53). Treatment of cryptococcal meningitis in patients with AIDS is difficult because antifungal drugs often