Many viral proteins that contain MHC class I-restricted peptides are long-lived, and it is elusive how they can give rise to class I epitopes. Recently, we showed that direct presentation of an epitope of the long-lived lymphocytic choriomeningitis virus nucleoprotein (LCMV-NP) required neosynthesis in accordance with the defective ribosomal products hypothesis. In this study, we report that LCMV-NP can be cross-primed in mice using either LCMV-NP-transfected human HEK293 or BALB/c-derived B8 cells as Ag donor cells. In addition, we establish that contrary to direct presentation, cross-presentation required accumulation of the mature LCMV-NP and could not be sustained by the newly synthesized LCMV-NP protein, intermediate proteasomal degradation products, or the minimal NP396 epitope. Nevertheless, NP cross-presentation was enhanced by heat shock and was blunted by inhibitors of heat shock protein 90 and gp96. We propose that cross-presentation has evolved to sustain the presentation of stable viral proteins when their neosynthesis has ceased in infected donor cells.

This study shows that the Bacillus anthracis pXO1 virulence plasmid carries a Rap-Phr system, BXA0205, which regulates sporulation initiation in this organism. The BXA0205Rap protein was shown to dephosphorylate the Spo0F response regulator intermediate of the phosphorelay signal transduction system that regulates the initiation of the developmental pathway in response to environmental, metabolic, and cell cycle signals. The activity of the Rap protein was shown to be inhibited by the carboxy-terminal pentapeptide generated through an export-import processing pathway from the associated BXA0205Phr protein. Deregulation of the Rap activity by either overexpression or lack of the Phr pentapeptide resulted in severe inhibition of sporulation. Five additional Rap-Phr encoding systems were identified on the chromosome of B. anthracis, one of which, BA3790-3791, also affected sporulation initiation. The results suggest that the plasmid-borne Rap-Phr system may provide a selective advantage to the virulence of B. anthracis.Bacillus anthracis, the etiological agent of anthrax, is a grampositive spore-forming organism that primarily infects ruminants but can also be highly pathogenic to other mammals, including humans. The intrinsic spore resistance to extreme stresses such as desiccation, solvents, extreme pH, temperature, UV, and ionizing radiation plays a major role in anthrax pathogenesis. Sporulation is essential for survival in the environment, and it evidently contributes to anthrax diffusion, because spores are usually present when the infection is initiated (27).The process of sporulation has been extensively studied in Bacillus subtilis and shown to be the result of a complex differentiation pathway that has its onset in a signal transduction system called phosphorelay. The phosphorelay is a more complex version of the two-component signal transduction systems, because it is composed of multiple central elements and it is modulated by a variety of ancillary factors (40,41).In B. subtilis, five histidine sensor kinases (KinA, -B, -C, -D, and -E) can respond to a multiplicity of inducing signals and activate the pathway by autophosphorylating and transferring the activating phosphoryl group to an intermediate response regulator acceptor called Spo0F. From Spo0F, the phosphoryl group is then transferred to the Spo0A response regulator through the Spo0B phosphotransferase. Spo0A is the critical transcription regulator for sporulation initiation. Accumulation of its activated form, Spo0AϳP, during growth progressively results in the repression of genes not required for sporulation and the activation of genes necessary for spore formation (7,19,28,53).Negative inputs into the phosphorelay are brought about mainly by aspartyl phosphate phosphatases that specifically dephosphorylate the response regulator components of the system. The three members of the Spo0E family of phosphatases dephosphorylate Spo0AϳP, while three members of the Rap family of phosphatases dephosphorylate the Spo0FϳP intermediate (18,36,38). Rap p...

The initiation of sporulation in