Short poly(C)-tract Mengoviruses have proven vaccine efficacy in many species of animals. A novel vector for the delivery of foreign proteins was created by insertion of a second autoproteolytic primary cleavage cassette linked to a multiple cloning site (MCS) into an attenuated variant of Mengo. Nineteen cDNAs from foreign sequences that ranged from 39 to 1653 bases were cloned into the MCS. The viral reading frame was maintained and translation resulted in dual, autocatalytic excision of the foreign peptides without disruption of any Mengo proteins. All cDNAs except those with the largest insertions produced viable virus. Active proteins such as GFP, CAT, and SIV p27 were expressed within infected cells. Relative to parental Mengo, the growth kinetics and genetic stability of each vector was inversely proportional to the size of the inserted sequence. While segments up to 1000 bases could be carried, inserts greater than 500-600 bases were usually reduced in size during serial passage. The limit on carrying capacity was probably due to difficulties in virion assembly or particle stability. Yet for inserts less than 500-600 bases, the Mengo vectors provided an effective system for the delivery of foreign epitopes into cells and mice.

Interferons induced by viral infections can have powerful immuno- modulatory effects, and several epidemiologic studies have found an association between certain viral infections and reduced prevalence of allergy. We hypothesized that allergenic proteins could be synthesized by a replicating virus, and this construct could be useful as an immunomodulator. To test this hypothesis, we cloned an allergenic protein (ovomucoid [OVM]) into a murine picornavirus (Mengo virus) vector. This plasmid has a multicloning site surrounded by auto-catalytic sequences so that a foreign protein will be cleaved from viral proteins during replication. OVM sequences were cloned in the context of full-length viral genome cDNA, T7 RNA transcripts of this plasmid were transfected into HeLa cells, and recombinant virus plaques appeared on the second passage. Sequence analysis of recombinant viruses derived from individual plaques demonstrated that three viral isolates contained up to 2/3 of the OVM coding sequence, which was retained by the viruses after 5 additional passages in HeLa cells. The experiments verify the stable expression of immunoreactive OVM subunits by replicating viruses. These virus/allergen constructs could provide a tool to evaluate whether intracellular presentation of allergenic proteins in the context of a viral infection could prevent allergic sensitization upon re-challenge.

The translation of maternal glp-1 mRNA is regulated both temporally and spatially in the early Caenorhabditis elegans embryo (T. C. Evans, S. L. Crittenden, V. Kodoyianni, and J. Kimble, Cell 77, 183-194, 1994). To investigate the control of embryonic glp-1 expression, we have examined the distribution of GLP-1 protein in selected maternal effect mutants that affect pattern or fate in the early embryo. We find that mutants that disrupt anterior-posterior asymmetry in the early embryo (par-1-par-6, emb-8, Par(q537)) disrupt the spatial but not temporal control of GLP-1 expression: GLP-1 is observed at the normal stage of embryogenesis in par-like mutants; however, it is uniformly distributed. In contrast, mutants that alter blastomere identity (skn-1, pie-1, mex-1, apx-1) do not affect the normal GLP-1 pattern. We conclude that genes controlling the asymmetry of cellular components, including P granules, also control GLP-1 asymmetry in the early embryo. The finding that mutants that disrupt anterior-posterior asymmetry translate GLP-1 in all blastomeres suggests that loss of embryonic asymmetry causes translational activation of GLP-1 in the posterior.

ABSTRACT:The world collection of sugarcane (Saccharum hybrids) and related grasses (WCSRG) is an important source of genes for sugarcane and energycane breeding. The core collection or diversity panel of the WCSRG was created in Canal Point with 300 accession and 10 checks to evaluate its diversity. The fiber components of the species and accessions within the WCSRG are unknown, so a shredded and dried fiber sample was taken from each accession and sent for fiber analysis. The acetyl groups, acid insoluble lignin, acid soluble lignin, arabinan, glucan, holocellulose, total lignin, structural ash, and xylan were quantified on a % fiber basis and nonstructural ash on a % total basis. There were significant, but not large differences between species for holocellulose, lignin, acetyl, acid soluble lignin, PLANT BREEDING -Article

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The development of therapeutic cancer vaccines remains an active area, although previous approaches have yielded disappointing results. We have built on lessons from previous cancer vaccine approaches and immune checkpoint inhibitor research to develop VBIR, a vaccine-based immunotherapy regimen. Assessment of various technologies led to selection of a heterologous vaccine using chimpanzee adenovirus (AdC68) for priming followed by boosts with electroporation of DNA plasmid to deliver T cell antigens to the immune system. We found that priming with AdC68 rapidly activates and expands antigen-specific T cells and does not encounter pre-existing immunity as occurs with the use of a human adenovirus vaccine. The AdC68 vector does, however, induce new anti-virus immune responses, limiting its use for boosting. To circumvent this, boosting with DNA encoding the same antigens can be done repetitively to augment and maintain vaccine responses. Using mouse and monkey models, we found that the activation of both CD4 and CD8 T cells was amplified by combination with anti-CTLA-4 and anti-PD-1 antibodies. These antibodies were administered subcutaneously to target their distribution to vaccination sites and to reduce systemic exposure which may improve their safety. VBIR can break tolerance and activate T cells recognizing tumor-associated self-antigens. This activation lasts more than a year after completing treatment in monkeys, and inhibits tumor growth to a greater degree than is observed using the individual components in mouse cancer models. These results have encouraged the testing of this combination regimen in cancer patients with the aim of increasing responses beyond current therapies.