Low-level Hypermutation in T Cell–independent Germinal Centers Compared with High Mutation Rates Associated with T Cell–dependent Germinal Centers

Exceptionally germinal center formation can be induced without T cell help by polysaccharide-based antigens, but these germinal centers involute by massive B cell apoptosis at the time centrocyte selection starts. This study investigates whether B cells in germinal centers induced by the T cell–independent antigen (4-hydroxy-3-nitrophenyl)acetyl (NP) conjugated to Ficoll undergo hypermutation in their immunoglobulin V region genes. Positive controls are provided by comparing germinal centers at the same stage of development in carrier-primed mice immunized with a T cell–dependent antigen: NP protein conjugate. False positive results from background germinal centers and false negatives from non-B cells in germinal centers were avoided by transferring B cells with a transgenic B cell receptor into congenic controls not carrying the transgene. By 4 d after immunization, hypermutation was well advanced in the T cell–dependent germinal centers. By contrast, the mutation rate for T cell–independent germinal centers was low, but significantly higher than in NP-specific B cells from nonimmunized transgenic mice. Interestingly, a similar rate of mutation was seen in extrafollicular plasma cells at this stage. It is concluded that efficient activation of hypermutation depends on interaction with T cells, but some hypermutation may be induced without such signals, even outside germinal centers

CDK Inhibitor p18INK4c Is Required for the Generation of Functional Plasma Cells

AbstractB cell terminal differentiation is associated with the onset of high-level antibody secretion and cell cycle arrest. Here the cyclin-dependent kinase (CDK) inhibitor p18INK4c is shown to be required within B cells for both terminating cell proliferation and differentiation of functional plasma cells. In its absence, B cells hyperproliferate in germinal centers and extrafollicular foci in response to T-dependent antigens but serum antibody titers are severely reduced, despite unimpaired germinal center formation, class switch recombination, variable region-directed hypermutation, and differentiation to antibody-containing plasmacytoid cells. The novel link between cell cycle control and plasma cell differentiation may, at least in part, relate to p18INK4c inhibition of CDK6. Cell cycle arrest mediated by p18INK4C is therefore requisite for the generation of functional plasma cells

Viral Superantigen Drives Extrafollicular and Follicular B Cell Differentiation Leading to Virus-specific Antibody Production

Mouse mammary tumor virus (MMTV[SW]) encodes a superantigen expressed by infected B cells. It evokes an antibody response specific for viral envelope protein, indicating selective activation of antigen-specific B cells. The response to MMTV(SW) in draining lymph nodes was compared with the response to haptenated chicken gamma globulin (NP-CGG) using flow cytometry and immunohistology. T cell priming occurs in both responses, with T cells proliferating in association with interdigitating dendritic cells in the T zone. T cell proliferation continues in the presence of B cells in the outer T zone, and B blasts then undergo exponential growth and differentiation into plasma cells in the medullary cords. Germinal centers develop in both responses, but those induced by MMTV(SW) appear later and are smaller. Most T cells activated in the T zone and germinal centers in the MMTV(SW) response are superantigen specific and these persist for weeks in lymph nodes draining the site MMTV(SW) injection; this contrasts with the selective loss of superantigen-specific T cells from other secondary lymphoid tissues. The results indicate that this viral superantigen, when expressed by professional antigen-presenting cells, drives extrafollicular and follicular B cell differentiation leading to virus-specific antibody production

Germinal Centers without T Cells

Germinal centers are critical for affinity maturation of antibody (Ab) responses. This process allows the production of high-efficiency neutralizing Ab that protects against virus infection and bacterial exotoxins. In germinal centers, responding B cells selectively mutate the genes that encode their receptors for antigen. This process can change Ab affinity and specificity. The mutated cells that produce high-affinity Ab are selected to become Ab-forming or memory B cells, whereas cells that have lost affinity or acquired autoreactivity are eliminated. Normally, T cells are critical for germinal center formation and subsequent B cell selection. Both processes involve engagement of CD40 on B cells by T cells. This report describes how high-affinity B cells can be induced to form large germinal centers in response to (4-hydroxy-3-nitrophenyl) acetyl (NP)-Ficoll in the absence of T cells or signaling through CD40 or CD28. This requires extensive cross-linking of the B cell receptors, and a frequency of antigen-specific B cells of at least 1 in 1,000. These germinal centers abort dramatically at the time when mutated high-affinity B cells are normally selected by T cells. Thus, there is a fail-safe mechanism against autoreactivity, even in the event of thymus-independent germinal center formation

T Helper 1 (Th1) and Th2 Characteristics Start to Develop During T Cell Priming and Are Associated with an Immediate Ability to Induce Immunoglobulin Class Switching

The respective production of specific immunoglobulin (Ig)G2a or IgG1 within 5 d of primary immunization with Swiss type mouse mammary tumor virus [MMTV(SW)] or haptenated protein provides a model for the development of T helper 1 (Th1) and Th2 responses. The antibody-producing cells arise from cognate T cell B cell interaction, revealed by the respective induction of Cγ2a and Cγ1 switch transcript production, on the third day after immunization. T cell proliferation and upregulation of mRNA for interferon γ in response to MMTV(SW) and interleukin 4 in response to haptenated protein also starts during this day. It follows that there is minimal delay in these responses between T cell priming and the onset of cognate interaction between T and B cells leading to class switching and exponential growth. The Th1 or Th2 profile is at least partially established at the time of the first cognate T cell interaction with B cells in the T zone

STATHMIN EXPRESSION IS ASSOCIATED WITH THE ABILITY OF CELLS TO PROGRESS THROUGH THE CELL CYCLE

Stathmin is a prominent cytosolic protein which can be phosphorylated by MAP kinase, cAMP-dependent protein kinase and p34cdc2 kinase. Iinmunoslaining of human and mouse tissues has revealed that stathmin is expressed in the proliferative compartment of cells of most, if not all, eel! lineages. The myeloid cell lines HL60 and K562 express stathmin at very high levels. When these cells are induced to differentiate, stathmin expression is down-regulated as cells go out of cycle. A decrease in the proportion of stathmin that is phosphorylated was also observed. To investigate the role that stathmin plays in the control of cell proliferation, we have transfected the promonocytoid cell line U937 with the pMep4 vector containing stathmin cDNA in an antisense orientation. Antisense stathmin transfected cells were unable to undergo normal cell division and became multinucleate. Belmont and Mitchison (Cell 19%, 84, 623-631) have shown in vitro that stathmin interacts with tubulin dimers to increase the catastrophe rate of microtubules and suggest that stathmin may increase the catastrophe rate of microtubules during mitosis. The above findings show that stathmin plays an important role in the progression of cells through cycle.</p