Regulation of expression of three genes in the polyhedron
envelope protein (PEP) gene region of the Orgyia pseudotsugata
multinucleocapsid nuclear polyhedrosis virus (OpMNPV) was
examined. These genes include open reading frame (ORF) 1 (encoding
p21), ORF 2 (encoding gp16), and ORF 3 (encoding the polyhedron
envelope protein). The effect of elimination of the late promoter
elements of each ORF or both ORFs 1 and 2 on ORF 3 expression was
examined by using an ORF 3 promoter-CAT gene fusion. The data
indicated that the ORF 3 promoter was essential for the expression of
the PEP. Destruction of ORF 1 caused no effect whereas destruction of
ORF 2 promoter resulted in a 29% increase in CAT activity.
To characterize the role of ORF 1 and 2 in the viral life cycle and
the location of the proteins in virions and infected cells, antisera were
produced against these proteins. The 21 kDa protein was present in
both purified budded and occluded virions as demonstrated by
Western blot analysis. Immunoelectron microscopy showed that the
21 kDa protein was a capsid-associated protein in both phenotypes.
The ORF 2 gene encodes a 12 kDa protein that is N-glycosylated,
migrates at a MW of 16 kDa, and is not present in budded or occluded
virions. Immunoelectron microscopy indicated that gp16 is associated
with lamellar-like membranous structures in close association with the
nuclear membrane. It was also found associated with envelopes of
virions that had budded from the nucleus into the cytoplasm.
A gene that reportedly has a similar role to ORF 3 (polyhedron
envelope protein) has been described in Autographa californica
MNPV. This gene encodes a protein called the spheroidin-like protein
(SLP) because of its sequence similarity to the spheroidin inclusion
protein of the Choristoneura biennis entomopox virus. The gene was
located, sequenced, transcriptionally mapped in OpMNPV and an
antiserum was produced against a fusion protein containing most of
the SLP ORF. Immunoelectron microscopy showed that the protein
was concentrated in cytoplasmic inclusion bodies and was not
associated with the polyhedron envelope structure in OpMNPV. It was
found to be associated with polyhedra of AcMNPV, but no specific
association with the polyhedron envelope was found.
The role of the PEP and the p10 protein in polyhedron
morphogenesis was examined using deletion mutants of OpMNPV
and immunoelectron microscopy. The p10 deletion mutant produced
polyhedra with patchy and poorly attached polyhedron envelopes,
suggesting p10 has a direct or indirect role in the proper formation of
the polyhedron envelope. The PEP deletion mutant showed that PEP
was an essential component in the formation of the polyhedron
envelope. The mutant with both p10 and PEP deleted had polyhedra
that showed a distinct cubic morphology. These data suggest that these
two proteins may affect polyhedra morphology. / Graduation date: 1993
| Identifer | oai:union.ndltd.org:ORGSU/oai:ir.library.oregonstate.edu:1957/36737 |
| Date | 12 May 1992 |
| Creators | Gross, Christian Hans |
| Contributors | Rohrmann, George F. |
| Source Sets | Oregon State University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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