During morphogenesis, poxviruses undergo a remarkable transition from spherical immature forms to brick-shaped infectious particles lacking helical or icosahedral symmetry. In this study, we show that the transitory honeycomb lattice coating the lipoprotein membrane of immature vaccinia virus particles is formed from trimers of a 62-kD protein encoded by the viral D13L gene. Deep-etch electron microscopy demonstrated that anti-D13 antibodies bound to the external protein coat and that lattice fragments were in affinity-purified D13 preparations. Soluble D13 appeared mostly trimeric by gel electrophoresis and ultracentrifugation, which is consistent with structural requirements for a honeycomb. In the presence or absence of other virion proteins, a mutated D13 with one amino acid substitution formed stacks of membrane-unassociated flat sheets that closely resembled the curved honeycombs of immature virions except for the absence of pentagonal facets. A homologous domain that is present in D13 and capsid proteins of certain other lipid-containing viruses support the idea that the developmental stages of poxviruses reflect their evolution from an icosahedral ancestor.
Infectious poxvirus particles are unusual in that they are brick shaped and lack symmetry. Nevertheless, an external honeycomb lattice comprised of a capsid-like protein dictates the spherical shape and size of immature poxvirus particles. In the case of vaccinia virus, trimers of 63-kDa D13 polypeptides form the building blocks of the lattice. In the present study, we addressed two questions: how D13, which has no transmembrane domain, associates with the immature virion (IV) membrane to form the lattice structure and how this scaffold is removed during the subsequent stage of morphogenesis. Interaction of D13 with the A17 membrane protein was demonstrated by immunoaffinity purification and Western blot analysis. In addition, the results of immunogold electron microscopy indicated a close association of A17 and D13 in crescents, as well as in vesicular structures when crescent formation was prevented. Further studies indicated that binding of A17 to D13 was abrogated by truncation of the N-terminal segment of A17. The N-terminal region of A17 was also required for the formation of crescent and IV structures. Disassembly of the D13 scaffold correlated with the processing of A17 by the I7 protease. When I7 expression was repressed, D13 was retained on aberrant virus particles. Furthermore, the morphogenesis of IVs to mature virions was blocked by mutation of the N-terminal but not the C-terminal cleavage site on A17. Taken together, these data indicate that A17 and D13 interactions regulate the assembly and disassembly of the IV scaffold.The assembly and morphogenesis of vaccinia virus (VACV) and other poxviruses occurs in specialized regions of the cytoplasm called factories. The first distinctive viral forms discerned by transmission electron microscopy are spherical immature virions (IVs) and their membrane crescent precursors, which appear to be covered by a layer of spicules (14). Morerecent studies employing three-dimensional deep-etch electron microscopy revealed that the "spicule coat" of IVs is actually a continuous honeycomb lattice (20). The IVs enclose dense granular material comprising the core precursors and a DNA nucleoid. The "spicule coat" is lost as the IVs undergo a remarkable transition into dense, brick-shaped infectious mature virions (MVs).Several studies led to the identification of D13 protein trimers as the building blocks of the scaffold: (i) single amino acid changes in D13 are responsible for VACV mutants that are resistant to the drug rifampin (rifampicin) (4, 11, 42), which causes reversible formation of irregular membranes lacking the "spicule coat" (18, 29, 30); (ii) repression of D13 expression results in a phenotype identical to that caused by the drug rifampin (50); (iii) antibody to D13 labels IVs (40) on the outer surface (28, 41); (iv) in the presence of rifampin, D13 antibodies label cytoplasmic inclusions that are distinct from aberrant viral membranes (40); and (v) the results of physical and microscopic studies indicate that D13 exists as trimers of 63-kDa subunits arran...
Background: ␣1-Antitrypsin (␣1AT) deficiency (␣1ATD) is a consequence of defective folding, trafficking, and secretion of ␣1AT. Results: SAHA restores the secretion of an active form of Z-␣1AT in part through a calnexin-and HDAC7-sensitive dependent mechanism(s). Conclusion: SAHA may represent a potential therapeutic approach for ␣1ATD. Significance: SAHA is a regulator of the proteostasis biology of Z-␣1AT, favoring export of a functional form to serum.
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