Helper T (Th) cell differentiation is highly regulated by cytokines but initiated by mitogens. By examining gene expression in discrete generations of dividing cells, we have delineated the relationship between proliferation and differentiation. Initial expression of IL-2 is cell cycle-independent, whereas effector cytokine expression is cell cycle-dependent. IFNgamma expression increases in frequency with successive cell cycles, while IL-4 expression requires three cell divisions. Cell cycle progression and cytokine signaling act in concert to relieve epigenetic repression and can be supplanted by agents that hyperacetylate histones and demethylate DNA. Terminally differentiated cells exhibit stable epigenetic modification and cell cycle-independent gene expression. These data reveal a novel mechanism governing Th cell fate that initially integrates proliferative and differentiative signals and subsequently maintains stability of the differentiated state.

To determine whether DNA immunization could elicit protective immunity to Leishmania major in susceptible BALB/c mice, cDNA for the cloned Leishmania antigen LACK was inserted into a euykaryotic expression vector downstream to the cytomegalovirus promoter. Susceptible BALB/c mice were then vaccinated subcutaneously with LACK DNA and challenged with L. major promastigotes. We compared the protective efficacy of LACK DNA vaccination with that of recombinant LACK protein in the presence or absence of recombinant interleukin (rIL)-12 protein. Protection induced by LACK DNA was similar to that achieved by LACK protein and rIL-12, but superior to LACK protein without rIL-12. The immunity conferred by LACK DNA was durable insofar as mice challenged 5 wk after vaccination were still protected, and the infection was controlled for at least 20 wk after challenge. In addition, the ability of mice to control infection at sites distant to the site of vaccination suggests that systemic protection was achieved by LACK DNA vaccination. The control of disease progression and parasitic burden in mice vaccinated with LACK DNA was associated with enhancement of antigen-specific interferon-γ (IFN-γ) production. Moreover, both the enhancement of IFN-γ production and the protective immune response induced by LACK DNA vaccination was IL-12 dependent. Unexpectedly, depletion of CD8+ T cells at the time of vaccination or infection also abolished the protective response induced by LACK DNA vaccination, suggesting a role for CD8+ T cells in DNA vaccine induced protection to L. major. Thus, DNA immunization may offer an attractive alternative vaccination strategy against intracellular pathogens, as compared with conventional vaccination with antigens combined with adjuvants.

Significance After infection of Escherichia coli by bacteriophage T7, the host RNA polymerase (RNAP) produces early phage transcription products that encode the phages own RNAP (that transcribes subsequent phage genes) as well as Gp2, an essential inhibitor of the host RNAP. X-ray crystal structures of E. coli RNAP define the structure and location of the RNAP σ 70 subunit domain 1.1 inside the RNAP active site channel, where it must be displaced by the DNA upon formation of the transcription complex. Gp2 binds inside the RNAP active site channel and also interacts with , preventing from exiting the RNAP active site channel. Gp2 thus misappropriates a domain of the RNAP, , to inhibit the function of the enzyme.