Spherical virus capsids are large, multimeric protein shells whose assembly and stability depend on the establishment of multiple non-covalent interactions between many polypeptide subunits. In a foot-and-mouth disease virus capsid, 42 amino acid side chains per protomer are involved in noncovalent interactions between pentameric subunits that function as assembly/disassembly intermediates. We have individually truncated to alanine these 42 side chains and assessed their relevance for completion of the virus life cycle and capsid stability. Most mutations provoked a drastic reduction in virus yields. Nearly all of these critical mutations led to virions whose thermal inactivation rates differed from that of the parent virus, and many affected also early steps in the viral cycle. Rapid selection of genotypic revertants or variants with forward or compensatory mutations that restored viability was occasionally detected. The results with this model virus indicate the following. (i) Most of the residues at the interfaces between capsid subunits are critically important for viral function, in part but not exclusively because of their involvement in intersubunit recognition. Each hydrogen bond and salt bridge buried at the subunit interfaces may be important for capsid stability. (ii) New mutations able to restore viability may arise frequently at the subunit interfaces during virus evolution. (iii) A few interfacial side chains are functionally tolerant to truncation and may provide adequate mutation sites for the engineering of a thermostable capsid, potentially useful as an improved vaccine.

Toxic shock syndrome toxin 1 (TSST1)t is a 22-kD exotoxin produced by most strains of Staphylococcus aureus isolated from patients with toxic shock syndrome (TSS) (1) . TSST 1 is a potent activator of monocytes and T cells . It acts on monocytes to induce the synthesis of IL-1 and TNF (2-4) . TSSTl also triggers T cell activation and proliferation (5), as well as the production of large amounts of various lymphokines such as IL-2 (6) and IFN-y (7) . The massive induction of monokine and lymphokine production by TSST1 is thought to play an important role in the pathogenesis of TSS.A significant insight into the mechanism of action of TSST 1, as well as other related staphylococcal exotoxins, came with the observation that these toxins bind directly to MHC class II molecules (8-11). Among the MHC class II molecules, TSST1 binds equally well with high affinity and saturation kinetics to HLA-DR and -DQ antigens (8) . TSST 1 also binds to HLA-DP alleles, albeit with a much lower affinity than is observed for HLA-DR and -DQalleles (Scholl, P, unpublished data) . The MHC class II-bound toxin behaves as a superantigen that interacts with T cells via the TCR )3 chain (12, 13) to induce MHC-unrestricted T cell activation and proliferation .The ability ofMHC class II-bound TSST1 to engage T cells raises the possibility that TSST 1 may mimic nominal antigen in initiating cognate interaction between T and B cells resulting in B cell proliferation and Ig production . We demonstrate here that TSST1 induces both the proliferation of resting human B cells and their differentiation into Ig-secreting lymphocytes . Triggering of B cell proliferation and differentiation by TSST1 is dependent on the presence of T cells, and proceeds via This work was supported by U. S .

Apicomplexan parasites exhibit tremendous diversity in much of their fundamental cell biology, but study of these organisms using light microscopy is often hindered by their small size. Ultrastructural expansion microscopy (U-ExM) is a microscopy preparation method that physically expands the sample ∼4.5x. Here, we apply U-ExM to the human malaria parasite Plasmodium falciparum during the asexual blood stage of its lifecycle to understand how this parasite is organized in three-dimensions. Using a combination of dye-conjugated reagents and immunostaining, we have catalogued 13 different P. falciparum structures or organelles across the intraerythrocytic development of this parasite and made multiple observations about fundamental parasite cell biology. We describe that the microtubule organizing center (MTOC) and its associated proteins anchor the nucleus to the parasite plasma membrane during mitosis. Furthermore, the rhoptries, Golgi, basal complex, and inner membrane complex, which form around this anchoring site while nuclei are still dividing, are concurrently segregated and maintain an association to the MTOC until the start of segmentation. We also show that the mitochondrion and apicoplast undergo sequential fission events while maintaining an MTOC association during cytokinesis. Collectively, this study represents the most detailed ultrastructural analysis of P. falciparum during its intraerythrocytic development to date, and sheds light on multiple poorly understood aspects of its organelle biogenesis and fundamental cell biology.