The nucleus is known to be compartmentalized into units of function, but the processes leading to the spatial organization of chromosomes and nuclear compartments are not yet well defined. Here we report direct quantitative analysis of the global structural perturbations of interphase chromosome and interchromosome domain distribution caused by infection with herpes simplex virus-1 (HSV-1). Our results show that the peripheral displacement of host chromosomes that correlates with expansion of the viral replication compartment (VRC) is coupled to a twofold increase in nuclear volume. Live cell dynamic measurements suggest that viral compartment formation is driven by the functional activity of viral components and underscore the significance of spatial regulation of nuclear activities.

PECAM-1/CD31 is a cell adhesion and signaling molecule that is enriched at the endothelial cell junctions. Alternative splicing generates multiple PECAM-1 splice variants, which differ in their cytoplasmic domains. It has been suggested that the extracellular ligand-binding property, homophilic versus heterophilic, of these isoforms is controlled by their cytoplasmic tails. To determine whether the cytoplasmic domains also regulate the cell surface distribution of PECAM-1 splice variants, we examined the distribution of CD31-EGFPs (PECAM-1 isoforms tagged with the enhanced green fluorescent protein) in living Chinese hamster ovary cells and in PECAM-1-deficient endothelial cells. Our results indicate that the extracellular, rather than the cytoplasmic domain, directs PECAM-1 to the cell-cell borders. Furthermore, coculturing PECAM-1 expressing and deficient cells along with transfection of CD31-EGFP cDNAs into PECAM-1 deficient cells reveal that this PECAM-1 localization is mediated by homophilic interactions. Although the integrin ␣v␤3 has been shown to interact with PECAM-1, this trans-heterophilic interaction was not detected at the borders of endothelial cells. However, based on cocapping experiments performed on proT cells, we provide evidence that the integrin ␣v␤3 associates with PECAM-1 on the same cell surface as in a cis manner.

Recently, we demonstrated that experimental autoimmune encephalomyelitis (EAE) in the rat, passively transferred using myelin basic protein (MBP)-reactive encephalitogenic CD4+ T cells, was preventable by administration of a p55-tumor necrosis factor-IgG fusion protein (TNFR-IgG). This was despite quantitatively and qualitatively normal movement of these MBP-specific T cells to the central nervous system (CNS). To extend these findings, the effect of TNFR-IgG on EAE actively induced by injection of MBP in complete Freund's adjuvant was examined. This form of EAE in the rat typically involves an acute, self-limiting neurological deficit, substantial CNS inflammation, but minimal demyelination. Here we show that administration of TNFR-IgG prior to onset of disease signs completely prevented the neurological deficit or markedly reduced its severity. This blockade of clinical disease was dissociated from weight loss which occurred at the same tempo and magnitude as in control rats exhibiting neurological signs of disease such as paralysis. The timing of TNF blockade was critical as established clinical disease was relatively refractory to TNFR-IgG treatment. Activated CD4+ T cells expressing normal or elevated levels of VLA4, major histocompatibility complex class II, MRC OX40 and CD25 were isolated from or immunohistochemically localized in the CNS of clinically healthy rats treated before disease onset. There was a reduction of the amount of other inflammatory leukocytes in the CNS of these treated animals but, more importantly, the activation state of inflammatory leukocytes, as well as that of microglia isolated from treated animals, was reduced. Thus, TNFR-IgG, when administered before disease onset, appears to act by inhibiting an effector function of activated T cells and possibly other inflammatory leukocytes necessary to bring about the neurological deficit. However, while TNF is a critically important cytokine for the early events leading to initiation of EAE, it is not a necessary factor in the acute neurological deficit characteristic of this form of EAE, once disease onset has occurred.

The E3/19K protein of human adenovirus type 2 binds to class I MHC Ags thereby interfering with their cell surface expression and Ag presentation function. Currently, it is unclear exactly which structure of MHC molecules is recognized by the E3/19K protein. We have previously demonstrated that the murine H-2Kd Ag is able to associate with E3/19K, whereas the allelic H-2Kk molecule is not. By using exon shuffling between Kd and Kk molecules, the alpha 1 and alpha 2 domains of MHC class I molecules were identified as essential structures for binding the viral protein. In this report, we have examined the contribution of individual amino acids within the alpha 2 domain of MHC for binding E3/19K. First, we show that within this domain the alpha-helical part is most important for the interaction with E3/19K. By using site-directed mutagenesis, Kd-specific amino acids were introduced into the alpha-helix of the alpha 2 domain of Kk. By using the expression of mutagenized proteins in E3/19K+ cells, we have identified Tyr 156 and Leu 180 as being essential for the association with the E3/19K protein. In addition, Kd residue Glu 163 seems to contribute to the complex formation. Furthermore, analysis of a panel of Kd/Dd recombinants indicates that a similar region in the Dd molecule, namely, the C-terminal half of the alpha 2 domain, affects binding to E3/19K. Combining these results with Ab binding data, we present two alternative models of how the adenovirus protein may bind to the alpha 1 and alpha 2 domains.