Baculovirus Molecular Biology
Figures and Tables
(revised 5/10/09)
Copyright G. F. Rohrmann
These figures are freely available for non commercial use:
For other use contact:
George F. Rohrmann
Department of Microbiology
Oregon State University
rohrmanng@orst.edu
Citation:
Rohrmann GF. Baculovirus Molecular Biology. Bethesda (MD): National Library of Medicine (US), National Center for
Biotechnology Information; 2008 November. http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=bacvir
Fig. 1.1 Baculovirus Occlusion Bodies.
Scanning EM by K. Hughes and R.B. Addison
Fig. 1.2. Phylogenetic relatedness of LEF8 from
selected baculoviruses. Neighbor joining;
bootstrap analysis (1000 reps).
Table 1.1. Distribution1 of baculoviruses in insect
orders {Martignoni, 1986 #254}.
Insect orders
NPVs
GVs
Diptera
27
Hymenoptera
30
Lepidoptera
456
148
1
Indicates the number of species that have been
reported to be infected
1
Table 1.2 Genera of the Baculoviridae
Genus
Members
Alphabaculovirus
Lepidopteran NPVs
Betabaculovirus
Lepidopteran GVs
Gammabaculovirus
Hymenopteran NPVs
Deltabaculovirus
Dipteran NPVs
Table 1.3. Genes1 found in and unique to all sequenced Group I NPV genomes
Ac1 (ptp), Ac16 (BV-ODV26), Ac27 (iap-1), Ac30, Ac42 (gta), Ac72, Ac73,
Ac114, Ac124, Ac128 (gp64), Ac 132, Ac151 (ie2)
1
1
Genes are designated by their AcMNPV orf number
Table 1.4. Genome size and predicted ORF content* of selected baculoviruses
Virus type
Name of Virus
Size (kb)
Orfs
(>50 aa)
Group I (12 members)**
Group II (16 members)
GV (10 members)
Hymenopt. NPV (3 members)
Dipteran NPV (1 member)
EppoMNPV (45)
AnpeNPV (46)
AcMNPV (47)
AdhoNPV (48)
SeMNPV(49)
AgseNPV (50)
LdMNPV(29)
LeseNPV (51)
AdorGV (52)
CrleGV (31)
CpGV (33)
XecnGV (30)
NeleNPV (53)
NeabNPV (54)
NeseNPV (55)
CuniNPV (56)
119
126
134
113
136
148
161
168
100
111
124
179
82
84
86
108
*Selected f r om over 40 gen ome sequences ( 200 8)
** The n u mbers in brackets indicate the total nu mber o f genomes in the catego ry.
136
145
~150
125
139
153
163
169
119
124
143
181
89
93
90
109
Table 1.5. Conserved genes in baculoviruses and nudiviruses
Conserved in baculovirus
AcMNPV ORF
Conserved in Nudivirus
Ac6 Lef2
Ac14 Lef1
Ac22 PIF2
+
Ac40 p47
Ac50 LEF-8
+
Ac54 vp1054
Ac62 LEF9
+
Ac65 DNA pol
+
Ac66
+(GbNV)
Ac68
Ac77 VLF-1
+
Ac80 gp41
+
Ac81
Ac83 VP91
+
Ac89 VP39
Ac90 LEF-4
+
Ac92 p33
Ac95 DNA helicase
+
Ac96
+
Ac98
+
Ac99 LEF-5
+
Ac100 p6.9
Ac109
Ac115 PIF-3
+
Ac119 PIF-1
+
Ac133 Alkaline nuclease
Ac138 p74
+
Ac142
Ac143 odv-E18
Ac144
Ac148 odv-e56
+
Table 2.1 Proteins Associated with Baculovirus Occlusion Bodies
AcMNPV orf # and name
Distribution in the Baculoviridae
1
Effect of Deletion2
Viable
Ac8 Polyhedrin
All
Ac131 Polyhedron
envelope/Calyx
All except CuniNPV
Viable
Enhancin
A few NPVs and GVs
Viable
Ac137 p10
Group I/II; some GVs
Viable
Alkaline proteases
Non baculovirus contaminants
1
CuniNPV polyhedrin is unrelated to that of other baculoviruses.
2
For details see Chapter 11
Fig. 2.1. Two adjacent dissolved polyhedra showing rod-shaped virions trapped by the collapsed
polyhedron envelope. Photo by K. Hughe s
Fig. 2.2. Fibrous p10-containing material aligned with the calyx/polyhedron envelope.
Photo courtesy of G. Williams. From (133), with permission.
Fig. 2.3. Polyhedra from OpMNPV with the polyhedron envelope protein and p10 genes deleted. From
(8), with permission
Fig. 2.4. Distribution of envelope fusion proteins. Group I have homologs of both GP64 and F,
but F is not a fusion protein. Group II, GVs and dipteran viruses have homologs of F, whereas
the hymenopteran viruses have homologs of neither GP64 nor F.
Fig. 2.5. Structure of the baculovirus F (fusion) protein (Ld130) from LdMNPV and AcMNPV GP64.
A) Ld130 F protein. Shown is a predicted signal peptide (SP), fusion peptide (FP) and transmembrane
domain (TM) including the amino acid coordinates. The cleavage site is indicated by the arrow. A
predicted coiled coil domain is also indicated. The disulfide bond is predicted from (134). B) GP64.
This figure is derived from Kadlec (55).
Table 2.2. Occlusion Derived Virus Envelope Proteins and Per os infectivity factors
AcMNPV orf # and name
Distribution in the
Baculoviridae
Effect of Deletion
Ac16, BV/ODV-E26
Lep. I
Viable (74)
Ac46, ODV-E66
Lep. I, II, GV
Viable (82)
Ac94, ODV-E25
Lep. I, II, GV
Not viable (82)
Ac109, ODV-EC43
All
Not viable (82)
Ac143 ODV-E18
Lep. I, II
Not viable (85)
Ac148, ODV-E56
All
Severely compromised (82)
Ac22, pif-2
All
Viable (not per os) (88)
Ac115, pif-3
All
Viable (not per os) (88)
Ac119, pif-1
All
Viable (not per os) (88)
Ac138, p74
All
Viable (not per os) (135)
Ac145
All but CuniNPV
Viable (reduced per os) (94)
Ac150
Lep. I (a few)
Viable (94)
ODV envelope proteins
Per os infectivity factors
Fig. 2.6. Selected structural proteins of ODV. Shown are envelope proteins, the tegument
protein, gp41, the DNA binding protein, p6.9, the PIF proteins, and two basal end-associated
proteins, pp78/83 and VLF-1. There are a variety of other capsid proteins (see text), but they
appear to have a more generalized distribution.
Table 2.3 Selected Proteins Associated with Baculovirus Nucleocapsids
Name and AcMNPV orf #
Ac100, P6.9 DNA binding
Distribution in the
Baculoviridae
All
Effect of Deletion or
mutation
Not viable (82)
Ac90, VP39
All
Not viable (82)
Ac80 GP41 tegument
All
Not viable (112)
Ac142
All
Not viable (113, 115)
Ac144
All
Not viable (113)
Ac66
All
Severely compromised (126)
Ac92 (P33)
All
Not viable(82)
Ac54 (VP1054)
All
Not viable (112)
Ac77 VLF-1
All
Not viable (136)
Ac104, VP80
Lep. I and II NPV
Not viable (82)
Ac9, PP78/83
Lep. I and II NPV
Not viable(124)
Ac129, P24
Lep. I, II, GV
Viable (132)
Fig. 3.1. A life cycle of a baculovirus causing systemic infection. Occlusion bodies ingested by an
insect, dissolve in the midgut and ODV are released which then infect epithelial cells (A). The virion
buds out of the cell in a basal direction and initiate a systemic infection (B). Early in the systemic
infection more BV are produced which spread the infection throughout the insect (C). Late in infection
occluded virions are produced, and the cell then dies releasing the occlusion bodies (D). The virogenic
stroma (VS) is indicate d .
Fig. 3.2. The insect midgut and virus infection. The midgut cells generate the peritrophic membrane
(PM) by the synthesis and secretion of chitin, muccopolysaccharides and proteins. They also secrete
digestive enzymes and ions that regulate the pH. Occlusion bodies are dissolved by the high pH in the
midgut lumen, and are further degraded by proteinases that may also digest the PM. The three major
types of midgut cells are indicated, columnar
Fig. 3.3 PH profiles along the gut lumens of two lepidoperan species. The pH of the
hemolymph was 6.7. The species shown are Lichnoptera felina (circles) and Manduca sexta
(triangles). This figure is reproduced with permission of the Company of Biologists from (18).
Fig. 3.4. Possible interactions of selected ODV envelope proteins and vp91 with midgut cells.
VP91, Ac145 and Ac150 all have chitin binding domains suggesting that they may interact with
chitin synthesizing cells. P74, PIF-1 and -2 bind to midgut cells. The role of PIF-3 is not
known. ODV-E66 has an enzymatic activity (haluronan lyase) that may assist in initiating the
infection.
Fig. 3.5. Budded virus infection of a Group I virus. BV attach to receptors located in clathrin
coated pits via GP64 and are endocytosed (A). The endocytic vesicle is acidified and this
changes the conformation of GP64 and causes the virion envelope to fuse with the endosomal
membrane releasing the nucleocapsid into the cytoplasm (B). The nucleocapsid may enter the
nucleus or insert its DNA through a nuclear pore complex (C), genes are transcribed, DNA is
replicated and nucleocapsids are assembled in the virogenic stroma (D). In Group I virus, at
least two envelope proteins are synthesized, GP64 and F. They are likely translated in
association with the endoplasmic reticulum, glycosylated and transported to and incorporated
into the cytoplasmic membrane via the Golgi apparatus (E). Nucleocapsids destined to
become BV exit the nucleus and are thought to transiently obtain an envelope that is lost (F).
They move to the cytoplasmic membrane at the site of concentrations of GP64 and F proteins,
bud through, and obtain envelopes (G).
Fig. 3.6. A hypothetical diagram of ODV membrane morphogenesis. In this diagram mRNA
encoding ODV envelope proteins are transcribed (A) and exported (B) to the cytoplasm for
translation (C) and the proteins are then targeted to the nucleus (D). Some of these proteins
may be targeted to the inner nuclear membrane and induce it to invaginate thereby forming
microvesicles (E). The microvesicles may be further modified by the incorporation of additional
virally encoded ODV envelope proteins and then virions become enveloped (F, G). Shown
are the outer nuclear membrane (onm), the inner nuclear membrane (inm) and nuclear pore
complexes (npc).
Fig. 3.7. A granulovirus life cycle with systemic infection. Many features of a systemic GV
infection are likely to be similar to that of NPVs, including the infection of insect midgut and the
systemic spread to other tissues (A, B). However, the GV infection leads to the clearing of the
nucleus with nuclear material (N) locating to the margins (C) and the virogenic stroma (VS)
distributed throughout the nucleus. Later in the infection the nuclear membrane becomes
fragmented and the nuclear and cytoplasmic regions merge (D). This figure is interpreted from
(75, 91).
Fig. 4.1. The baculovirus transcriptional cascade showing the interrelationship of host and viral
RNA polymerases and DNA replication and VLF-1.
Fig. 4.2. Diagram of the AcMNPV genome showing hrs. The numbers in circles indicate the
number of repeats in each hr. Below is shown a representative palindrome with mismatches
shown as underlined italicized larger type. The EcoRI site at the center of the palindrome is
also underlined. At the bottom is a schematic showing the CRE- and TRE-like sequences that
are located between the major palindromes.
Fig. 4.3. Activation of baculovirus early gene transcription by hrs and IE1. This diagram
shows two possible mechanism by which IE1 activates transcription. Transcription might be
activated directly by IE1 as shown in the bottom of the diagram, or it can interact with both
RNA polymerase II (in blue) and hr sequences thereby bringing hr-bound transcriptional
activators in close proximity to RNA polymerase II as shown in the top of the figure.
Table 5.1. Viral genes essential for DNA replication
Gene
DNA pol
Ssb
primase
Primase accessory factor
helicase
Origin binding protein
Processivity factor
Baculovirus
+
+ (LEF-3)
+ (LEF1)
+ (LEF2)
+ (p143)
+ IE1 (?)
-
Herpesvirus
+ (UL30)
+ (ICP8)
+ (UL52)
+ (UL8)
+ (UL5)
+ (UL9)
UL42
Fig. 5.2 Theoretical diagram of different stages of DNA replication and
packaging. Shown is replication coordinated with packaging (a and b) and DNA
replication that is independent of packaging showing extensive recombination
(f,g). The large stippled area is the virogenic stroma (VS). The circularization of
the genome by recombination is also indicated (c). Mature virions are
represented by (d and e).
Fig. 5.3 Hypothetical diagram for roles of two types of baculovirus DNA; genomic
DNA that is packaged and DNA that is not packaged and is essential for very late
transcription.
Fig. 6.1. Homology of baculovirus RNA polymerase subunits. The similarity of
the LEF-8 and -9 subunits to the two largest RNA polymerase subunits of
Drosophila melanogaster RNA polymerase II is shown. In addition, the
relatedness of P47 to the alpha subunit of B. subtilis RNA polymerase as
determined by HHpred is indicated (see text). The numbers at the end of the
lines indicate the size of each protein in amino acids. The numbers before the
sequences indicates the location of the domain within the sequence. The
underlined amino acids are not conserved.
Fig. 6.2. Relationships of RNA polymerase subunits. The large subunits, ’ and
in some
archaea have undergone fission and are present as either 3 or 4 components. The subunits are
color-coded indicating relatedness except for the auxillary subunits shown in white. This figure is
based on the data of . For description of baculovirus homology, see text.
Fig. 6.3. Capping baculovirus mRNA. The four modifications that occur during
capping are summarized in the following steps: 1; LEF-4 has the potential to
dephosphorylate the 5' end of mRNA; 2, and to transfer guanosine to the 5' end
of the mRNA; 3, the guanosine is methylated--enzyme is unknown; 4. The 2' OH
group of the terminal A is methylated by an unknown methytransferase. The
final structure is shown at the bottom.
Fig. 6.4. Comparison of late and very late gene transcription. Shown is late
gene transcription (a), and a hypothetical mechanism for the shutoff of late
transcription (b), and activation of very late gene transcription involving VLF-1
interacting with the burst sequence (shown as a red rectangle) (c and d). Late
transcription initiates within the late promoter element. Therefore all late
messages likely begin with the sequence UAAG.
Fig. 7.1. The cell cycle is regulated by the phosphorylation state of different cyclin molecules.
Baculovirus infection appears to block the cell cycle and prevents cells from undergoing
mitosis. They likely induce a 'pseudo' S phase-like environment in which the virus
transcriptional activator IE1, along with hr enhancer sequences preferentially activate early
viral genes leading to DNA replication and the production of the baculovirus RNA polymerase.
Fig. 7.2. Cell death regulation by baculovirus apoptotic suppressors. a) Baculovirus apoptotic
inhibitors P49 and P35 prevent apoptosis by blocking caspase activity. b) IAP-3 (vIAP) may
block apoptosis by interfering with cellular antagonists that interfere with cellular IAP function.
By blocking these antagonists, the cellular IAPs are free to block apoptosis.
Table 10.1 Major Factors influencing high levels of baculovirus very late gene transcription
Factor
Ability of virus to cause systemic
infection
Result
Allows virus to exploit insect synthetic systems,
e.g., the fat body
2 Shut off of early baculovirus and host
Makes host cell biosynthetic systems available
3 Shut off of baculovirus late gene
Makes baculovirus RNA polymerase available
4 A high concentration of unpackaged viral
High copy numbers of very late genes are
accessible for transcription
5 Efficiency of the baculovirus RNA
Facilitates high level mRNA production and
RNA capping
6 The biosynthetic capacity of insect cells
Allows high level very late gene expression
1
gene transcription
expression
DNA
polymerase