PLANT VIRUSES II

Some plant viruses have a very limited host range

and others attack numerous species.
Plant viruses can multiply only within living cells.
Plant viruses usually multiply only within living
plant cells, but some may be able to multiply
within the bodies of aphids and nematodes.
A given plant virus may be able to multiply only
within the living cells of one species or genera of
plants, but some can multiply within the cells of a
wide group of plant families.

Plant viruses do not attack animals and vice
versa.
Temperate viruses embed themselves within
the hosts nucleic acid and are transmitted
generation to generation just like genes of the
host.
Often viruses reside in their host without
causing any disease or symptom. Such latent
viruses are undiscovered.

Plant viruses vary in their mode of transmission. Usually
only a single mode of transmission is important, but some
viruses are transmitted by more than one mode.
Aphids or other sap-suckers are the most common mode of
transmission
Nematodes living in the soil transmit some viruses.
Sap transmission is important for only a few viruses and
occurs on cultivators, pruning, hands of workers, and
clothing of workers. Important for some potato viruses.
Pollen transmission from male flower to female occurs for a
few viruses. Such viruses are seed borne.
Most viruses are do not get into the gametes and therefore
are not seed-borne.

Detection
Until a virus is detected, its presence is not
known. Nearly everyone who received an
early polio vaccine is now a carrier of Simian
Virus 40.(SV40). The monkey cells used to
grow that vaccine were infected by SV40 but
no one knew SV40 existed.
Many plants are carriers of plant viruses but
show no disease. The yield might be reduced
but that would not be visible either.

The methods for detecting plant viruses
include:
Use of antibodies to the virus in the ELISA
assay.
Graft a leaf from the suspected plant to an
indicator plant. The virus moves into the host
plant and causes symptoms in the indicator
plant.

Sap transmission. Place a drop of sap from the
suspected plant on an intact leaf of an indicator
plant. Add some grit to the sap and rub so the
leaf is scratched and the virus, if any, can get into
the leaf. If the indicator plant show symptoms
then we know the virus came from the donor
plant. The indicator plant for sweetpotato viruses
is Brazilian Morning glory, tabacco, tomato,
lambsquarter, and other plants are used as
indicators in the sap and grafting assays.

It is not possible to cure a plant of viruses, but one of
the following methods may give a virus free clone.
Since the "virus-free" plant might contain a virus you
did not know about, it is proper to call them virus-
tested or virus indexed plants.
Often the terminal bud of a plant is free of virus.
Remove the 0.03 to 0.05 mm bud tip and grow it in
sterile agar contain sucrose and everything the bud
needs. 70 to 99.9% of the buds will rot or fail. If some
grow to plants, test that plant for virus as above and
you might find one which is free of virus. Use it to start
more plants. Be sure to test the plant for every known
virus.

Some viruses infect the tip also and the above
method will not work. In such cases try growing
the plant in an incubator that is so hot the plant
barely grows. Often you must supply extra carbon
dioxide at a very precise level. Then try to start a
new plant as above and test it for all the known
viruses.
Most viruses are not seed-borne. Therefore, a
new seedling may be free of virus. Test the
seedling for all known viruses.

Once you have a virus-indexed plant keep it in
a screened cage where it can't be infected via
a vector. Alternatively, keep it on sugar agar in
a sterile glass jar under fluorescent light at a
low temperature where it grows slowly. Use
scions from this microplant to grow plants for
the field.

Expression strategies in plant RNA viruses
Genome of plus strand viruses can be
translated directly upon 80 S ribosomes to
produce virus specific polypeptides.
The basic anatomy of these are similar to host
mRNA with which they must compete.
Variations are seen in the modifications to
RNA termini, in translation and protein
processing strategy.
3 different structures have been identified at the
5end.
m
7
G
5
ppp
5
seen in at least nine groups of plant
viruses, similar to those seen in eukaryotic mRNAs.
Existence of a small protein of mol.wt 3000-7000 ,
covalently linked to the 5 end, called VPg.( polio
virus also has this.)
Unmodified 5 termini with di or triphosphate
groups.


Tobacco mosaic virus,
Monopartite genome, 6395 nt,rod shaped
particle.
Has a cap at the 5 end of RNA , the 3 end has
a tRNA like structure , which can be
aminoacylated with histidine or valine.
Encloses 4 polypeptides.



68 nt leader seq precedes 2 long ORFs .
First ORF is translated into a polypeptide of
mol wt ,126000, P126.
Suppression of amber termination codon at
the end of the first ORF permits translation
readthrough into the second , producing P183.
2 smaller ORFs towards the 3 end , P30, P18.
P126, P183 appear early in the viral life cycle
and thought to constitute the viral replicase.
The internal ORFs coding for P30 , P 18, are
not expressed as translation readthro.
Instead subgenomic mRNAs are generated
from each gene by RNA transcription from
specific promoters.
P30 is supposed to be the involved in cell to
cell movement of the virus.
Cowpea Mosaic virus

An RNA-containing virus with isometric particles
about 28 nm in diameter.
Has a limited host range, is transmitted mainly by
beetles and readily by sap inoculation.
Infected plants contain two kinds of
nucleoprotein particle similar in size but differing
in RNA content.
Particles containing no RNA are also produced by
most isolates. The RNA species in different
particle types represent separate parts of the
viral genome.

Has a bipartite genome with 2 plus sense
strand RNA molecules.
Ultracentifuged purified CPMV shows 3
distinct spherical particles.
Fast sedimenting Bottom (B ) component with
single RNA species 5889 nt long.
Intermediate sedimenting M component 3481
nt long


Slow sedimenting top component has empty
viral capsids.
5 end of each RNA has a VPg attached to it
and a poly A tail at the 3end.
Both the RNA s are translated into large
polypeptides which are subsequently cleaved.
Both the RNA s are required for infectivity


B RNA can support its own replication
independantly of MRNA( viral replicase)
BRNA cannot move to adjacent plant cells in
the absence of MRNA.
The structure of CPMV is similar and has
resemblance to that of animal picarnoviruses
Both genome RNAs contain a small protein
(VPg) at their 5'-end and a poly A tail at their
3'-end.
Each RNA species contains one large open
reading frame, and they are translated in
vitro as well as in vivo into one (RNA1) or two
(RNA2) polyproteins that are cleaved by a viral
proteinase (encoded by RNA1) to give 15
intermediate and final processing products.

RNA1 carries all the information for RNA
replication, including the polymerase, VPg and
a protein containing a RNA replication,
including the polymerase, VPg and a protein
containing a nucleotide binding site, whereas
RNA2 codes for the two capsid proteins and
the movement protein, which are involved in
cell-to-cell transport of the virus.

Viral movement in plant cell
Upon infection, viral RNA (vRNA) initiates
synthesis of replicase, plus and minus
vRNA stands, subgenomic RNAs and
movement protein (MP), and coat
protein.
In response to infection,
callose accumulates in the wall region
surrounding the plasmodesmata (Pd)
restricting the cytoplasmic sleeve.

Stage II: MP, an integral endoplasmic reticulum
(ER) membrane protein, functions as a protein
raft binding vRNA on its cytoplasmic domains
forming a replication complex (VRC) that may
also contain replicase.
Intracellular trafficking of the VRC to the cortical
ER is either by diffusion in the ER lipids or by
vesicular trafficking.


Stress-induced class I beta-1,3,glucanase
traffics in the lumen of VRC vesicles to plasma
membrane (PM) with requisite docking
protein (filled arrowheads).
Stage III:
Cycling vesicles containing beta-1,3-glucanase
cargo fuse to PM and deliver beta-1,3-
glucanase to the cell wall.
Callose is hydrolyzed, allowing Pd to dilate.
Vesicles with attached VRC recycle back to the
cortical ER, in which vesicles fuse to cortical
ER. VRC diffuses through the Pd to adjacent
cells by diffusion in ER desmotubule
continuum motivated by the concentration
gradient between a viral-infect cell
and adjacent noninfected cells.