BIOPL 652.1 Molecular Plant
Pathology
30
January 2002 S. Lazarowitz
A. Important
questions
1. What
are the molecular and cellular events that allow virus multiplication?
2. What
steps are virus-specific (potential targets for resistance)?
3. c.
How do viruses modify host cells to promote their own multiplication?
B. Virus Definitions and Basic Principles (Lazarowitz 2001).
1. Unique
properties of viruses
a. Obligate
intracellular parasites that do not
undergo binary fission
b. Parasitic
genomes related to plasmids, but that can exist in 2 forms: (1) extracellular
metabolically inert form (the virion); and (2) intracellular replicating form.
2. Virus
families defined by genome structure/replication strategy (primarily) plus virion
morphology and diseases they cause.
3. Structure
of viral genomes: DNA or RNA, single
(ss) or double (ds) strand
a. ssRNA
viruses defined as positive-sense or negative-sense.
C. The
pathogenic process as a competition between virus and host
1. Nonhost: not permissive for virus replication
2. Host: non-specific or specific defenses
a.
Specific plant defenses: R genes (HR),
RNAi
b.
Virus counterstrategy
i.
Mutate Avr recognized by plant R gene
product
ii. Encode
suppressor of RNAi defense
c.
Viral Avr can be CP, Rep or MP
D. Virus
multiplication: Viruses do it from
within.
1. Stages
of viral life cycle: (1) attachment; (2) penetration and uncoating; (3)
synthesis and replication; (4) progeny virus assembly; (5) release
2. Based
originally on studies of animal viruses and phage
3. Getting
In and Out: Consequences of the cell
wall, plant viruses differ at stages (1) attachment, and (5) release.
a.
No evidence for specific receptors on
plant cell walls.
b.
Move cell-to-cell without going through
an extracellular phase.
E. Intracellular
replication. Viruses play by the rules:
Consequence of being obligate intracellular parasites -- it’s all quite
logical, really. (Knipe & Howley, Chapters 5, 6 and 14).
1. DNA viruses: genes organized and expressed as
are host genes, encode own replication initiation protein, but use host DNA
replicating enzymes for synthesis
2. RNA viruses: provide their own RNA-dependent
RNA polymerase
3. Eukaryotes:
Genes each have their own promoter and termination sequences, mRNAs are
polycistronic, with 5’-caps and
3’-polyA tails.
a. Animal
and plant DNA viruses replicate in the nucleus (exception: poxviruses)
i.
Viral proteins required for
transcription, replication and encapsidation will have nuclear localization
signals (NLSs)
ii. Some
viral proteins may shuttle between the nucleus and cytoplasm. These will contain nuclear export sequences
(NESs) in addition to NLSs.
b. Animal
and plant RNA viruses replicate in cytoplasm (exceptions: influenza viruses,
retroviruses):
i.
Reconcile genome structure with
eukaryotic requirement for monocistronic mRNAs
c. Viruses
have evolved novel expression strategies to maximize their coding capacity
while ‘playing by the rules’.
F. Specific examples of plant virus
multiplication.
1. Potyviruses (Tobacco Etch Virus, TEV): polyprotein (Kasschau and Carrington 1995; Li and Carrington 1995; Verchot
and Carrington 1995; Verchot and Carrington 1995; Schaad et al. 1996; Daros and
Carrington 1997; Haldeman-Cahill et al. 1998)
2. Tobamoviruses (Tobacco Mosaic Virus,
TMV): subgenomic RNA and readthrough (Young et al. 1987; Valle et al. 1992; Bao
et al. 1996; Schmitz et al. 1996; Urban et al. 1996)
3. Geminiviruses (Squash Leaf Curl, Tomato
Golden Mosaic, and Wheat Dwarf): DNA virus (Heyraud
et al. 1993; Eagle et al. 1994; Fontes et al. 1994; Pascal et al. 1994;
Heyraud-Nitschke et al. 1995; Laufs et al. 1995; Sanderfoot et al. 1996;
Lazarowitz et al. 1998; Qin et al. 1998; Ward and Lazarowitz 1999)
G. Viruses promote their own multiplication,
usually at the expense of the host.
This can involve redirecting host metabolic pathways and using cell
macromolecular trafficking pathways. Specific examples for plant viruses:
1. Inhibit/redirect DNA/RNA synthesis (direct or
indirect): Geminiviruses and the
requirement for host DNA synthesizing enzymes (Nagar,
et al. 1995; Xie et al. 1995; Ach et al. 1997; Gutierrez 2000; Kong et al.
2000).
2. ER and Golgi:
a. Cell secretory pathway for modification,
processing, and membrane targeting of glycoproteins
b. Replication of RNA viruses occurs in
association with cytoplasmic membranes (ER).
This has been known since the mid-1970s for poliovirus. Recent studies have shown specific targeting
of RNA replication complexes to and remodeling of ER membranes for TMV and
TEV. (Schaad
et al. 1997; Reichel and Beachy 1998).
c Plant virus movement involves ER: Viral MPs
target to ER (Ward et al. 1997;
Heinlein et al. 1998; Lazarowitz and Beachy 1999).
3. Cytoskeleton: It has recently been shown that
DNA animal viruses move along actin filaments and microtubules to the nucleus
during uncoating (e.g. Adenovirus, Herpes simplex virus) (Greber et al. 1993; Sodeik et al. 1997;
Li et al. 1998).
a. This is likely to be true for plant DNA
viruses (geminiviruses) and pararetroviruses (caulimoviruses), although it has
not as yet been demonstrated.
b. The TMV MP tracks on actin filaments and
microtubules through the cytoplasm (Heinlein,
Padgett et al. 1998).
4. Nuclear import and nuclear export:
a. Plant DNA virus proteins required for
replication, transcription, encapsidation and movement have been shown to be
nuclear targeted or nuclear shuttle proteins.
For some plant viruses RNA nuclear targeting may be a mechanism to ‘get
rid’ of proteins as part of the temporal control for viral replication. (CaMV, SqLCV, TEV) (Gorlich and Mattaj 1996; Sanderfoot et al. 1996; Schaad,
Haldeman-Cahill et al. 1996; Lartey and Citovsky 1997; Li et al. 1997; Smith
and Raikhel 1998; Weis 1998; Ward and Lazarowitz 1999)
5. Plasmodesmata:
a. Plant virus MPs modify plasmodesmata so that
viral genomes can be transported across the cell wall (Carrington et al. 1998; Santa Cruz et al. 1998; Lazarowitz and
Beachy 1999; Yang et al. 2000).
Additional Web Resource Sites:
PLPA 705 (Phytovirology): http://ppathw3.cals.cornell.edu/Virology/pp705/PP705.htm
BIOMI 409 (Principles of Virology): http://ppathw3.cals.cornell.edu/Virology/BioMi%20409/BioMi409.htm
General Background Reading (for those who are interested):
*Flint,
S.J. et al. (1999). “Principles of
Virology”
*Knipe,
D.M. and Howley, P.M. (2001). "Fundamental Virology" 4th Edition
Agrios,
G.N. (1997). “Plant Pathology”, 4th Edition
*Up to date, comprehensive, and highly
recommended. Flint et al. has excellent figures that illustrate many principles and
techniques. Knipe and Howley covers all
virus families in terms of replication and pathogenesis, and has much not
included in Flint et al.
In particular:
Virus structure,
multiplication and genetics
Flint Chapter 1 “Foundations of Virology” (some virus history!)
Chapter
2 “Virus Cultivation, Detection and Genetics”
Knipe & Chapter 3 (Harrison) “Principles of Virus
Structure
Howley Chapter 14 (Lazarowitz) “Plant Viruses” (a must, obviously!!)
Chapter
5 (Ball) “Replication Strategies of RNA Viruses”
Chapter
6 (DiMaio and Coen) “Replication Strategies of RNA Viruses”
Insect transmission
of plant viruses
Gray,
S. M. and N. Banerjee (1999). “Mechanisms of arthropod transmission of plant
and animal viruses.” Microbiol. Mol. Biol. Rev. X: xx-yy.
Gray, S. M. and D.
Rochon (1999). Vector transmission of plant viruses. Encyclopedia of
Virology. A. Granoff and R. G. Webster. San Diego, Academic Press. 3: 1899-1910.
References (cited in
outline for January 30, 2002): Plant Virus Specific
Ach,
R. A., T. Durfee, et al. (1997). “RRB1 and RRB2 encode maize
retinoblastoma-related proteins that interact with a plant D-type cyclin and
geminivirus replication protein.” Mol Cell Biol 17(9): 5077-86.
Bao,
Y., S. A. Carter, et al. (1996). “The 126- and 183-kilodalton proteins of
tobacco mosaic virus, and not their common nucleotide sequence, control mosaic
symptom formation in tobacco.” J Virol 70(9):
6378-83.
Carrington,
J. C., P. E. Jensen, et al. (1998). “Genetic evidence for an essential role for
potyvirus CI protein in cell-to-cell movement.” Plant J 14(4): 393-400.
Daros,
J. A. and J. C. Carrington (1997). “RNA binding activity of NIa proteinase of
tobacco etch potyvirus.” Virology 237(2):
327-36.
Eagle,
P. A., B. M. Orozco, et al. (1994). “A DNA sequence required for geminivirus
replication also mediates transcriptional regulation.” Plant Cell 6(8): 1157-1170.
Fontes,
E. P., P. A. Eagle, et al. (1994). “Interaction between a geminivirus
replication protein and origin DNA is essential for viral replication.” J Biol
Chem 269(11): 8459-8465.
Gutierrez,
C. (2000). “DNA replication and cell cycle in plants: learning from geminiviruses.”
Embo J 19(5): 792-9.
Haldeman-Cahill,
R., J. A. Daros, et al. (1998). “Secondary structures in the capsid protein
coding sequence and 3' nontranslated region involved in amplification of the
tobacco etch virus genome.” J Virol 72(5):
4072-9.
Heinlein,
M., H. S. Padgett, et al. (1998). “Changing patterns of localization of the
tobacco mosaic virus movement protein and replicase to the endoplasmic
reticulum and microtubules during infection.” Plant Cell 10(7): 1107-1120.
Heyraud,
F., V. Matzeit, et al. (1993). “The conserved nonanucleotide motif of the
geminivirus stem-loop sequence promotes replicational release of virus
molecules from redundant copies.” Biochimie 75(7): 605-15.
Heyraud-Nitschke,
F., S. Schumacher, et al. (1995). “Determination of the origin cleavage and
joining domain of geminivirus Rep proteins.” Nucl. Acids Res. 23(6): 910-916.
Kasschau,
K. D. and J. C. Carrington (1995). “Requirement for HC-Pro processing during
genome amplification of tobacco etch potyvirus.” Virology 209(1): 268-273.
Kong,
L. J., B. M. Orozco, et al. (2000). “A geminivirus replication protein
interacts with the retinoblastoma protein through a novel domain to determine
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Lartey,
R. and V. Citovsky (1997). “Nucleic acid transport in plant-pathogen
interactions.” Genet Eng (N Y) 19:
201-14.
Laufs,
J., I. Jupin, et al. (1995). “Geminivirus replication: genetic and biochemical
characterization of Rep protein function, a review.” Biochimie 77(10): 765-773.
Lazarowitz,
S. G. (2001). Plant Viruses. Virology. P. M. H. a. D. M. Knipe. Philadelphia,
Lippincott, Williams and Wilkins: Chapter
14.
Lazarowitz,
S. G. and R. N. Beachy (1999). “Viral movement proteins as probes for
investigating intracellular and intercellular trafficking in plants.” Plant
Cell 11: 535-548.
Li,
X. H. and J. C. Carrington (1995). “Complementation of tobacco etch potyvirus
mutants by active RNA polymerase expressed in transgenic cells.” Proc Natl Acad
Sci U S A 92(2): 457-61.
Li,
X. H., P. Valdez, et al. (1997). “Functions of the tobacco etch virus RNA
polymerase (NIb): subcellular transport and protein-protein interaction with
VPg/proteinase (NIa).” J Virol 71(2):
1598-607.
Nagar,
S., T. J. Pedersen, et al. (1995). “A geminivirus induces expression of a host
DNA synthesis protein in terminally differentiated plant cells.” Plant Cell 7(6): 705-19.
Pascal,
E., A. A. Sanderfoot, et al. (1994). “The geminivirus BR1 movement protein
binds single-stranded DNA and localizes to the cell nucleus.” Plant Cell 6: 995-1006.
Qin,
S., B. M. Ward, et al. (1998). “The bipartite geminivirus coat protein aids BR1
function in viral movement by affecting the accumulation of viral
single-stranded DNA.” J. Virol. 72(11):
9247-9256.
Reichel,
C. and R. N. Beachy (1998). “Tobacco mosaic virus infection induces severe
morphological changes of the endoplasmic reticulum.” Proc. Natl. Acad. Sci. USA
95(19): 11169-11174.
Sanderfoot,
A. A., D. J. Ingham, et al. (1996). “A viral movement protein as a nuclear
shuttle: The geminivirus BR1 movement
protein contains domains essential for interaction with BL1 and nuclear
localization.” Plant Physiol. 110:
23-33.
Santa
Cruz, S., A. G. Roberts, et al. (1998). “Cell to cell and phloem-mediated
movement of potato virus X: the role of virions.” Plant Cell: in press.
Schaad,
M. C., R. Haldeman-Cahill, et al. (1996). “Analysis of the VPg-proteinase (NIa)
encoded by tobacco etch potyvirus: effects of mutations on subcellular
transport, proteolytic processing, and genome amplification.” J Virol 70(10): 7039-48.
Schaad,
M. C., P. E. Jensen, et al. (1997). “Formation of plant RNA virus replication
complexes on membranes: role of an endoplasmic reticulum-targeted viral
protein.” Embo J 16(13): 4049-59.
Schmitz,
J., D. Prufer, et al. (1996). “Non-canonical translation mechanisms in plants:
efficient in vitro and in planta initiation at AUU codons of the tobacco mosaic
virus enhancer sequence.” Nucleic Acids Res 24(2): 257-63.
Urban,
C., K. Zerfass, et al. (1996). “UGA suppression by tRNACmCATrp occurs in
diverse virus RNAs due to a limited influence of the codon context.” Nucleic
Acids Res 24(17): 3424-30.
Valle,
R. P., G. Drugeon, et al. (1992). “Codon context effect in virus translational
readthrough. A study in vitro of the determinants of TMV and Mo-MuLV amber
suppression.” FEBS Lett 306(2-3):
133-9.
Verchot,
J. and J. C. Carrington (1995). “Debilitation of plant potyvirus infectivity by
P1 proteinase-inactivating mutations and restoration by second-site
modifications.” J Virol 69(3):
1582-90.
Verchot,
J. and J. C. Carrington (1995). “Evidence that the potyvirus P1 proteinase
functions in trans as an accessory factor for genome amplification.” J. Virol. 69(6): 3668-3674.
Ward,
B. M. and S. G. Lazarowitz (1999). “Nuclear export in plants: Use of geminivirus movement proteins for an in vivo cell based export assay.” Plant
Cell 11: 1267-1276.
Ward,
B. M., R. Medville, et al. (1997). “The geminivirus BL1 movement protein is
associated with endoplasmic reticulum-derived tubules in developing phloem
cells.” J. Virology 71: 3726-3733.
Xie,
Q., P. Suarez-Lopez, et al. (1995). “Identification and analysis of a
retinoblastoma binding motif in the replication protein of a plant DNA virus:
requirement for efficient viral DNA replication.” Embo J 14(16): 4073-82.
Yang,
Y., B. Ding, et al. (2000). “Cell-to-cell movement of the 25K protein of potato
virus X is regulated by three other viral proteins.” MPMI 13: 599-605.
Young,
N. D., J. Forney, et al. (1987). “Tobacco mosaic virus replicase and replicative
structures.” J. Cell Sci. 7:
277-285.
Additional
Cited General Reviews and Animal Virus References
Gorlich,
D. and I. W. Mattaj (1996). “Nucleocytoplasmic transport.” Science 271(5255): 1513-8.
Greber,
U. F., M. Willetts, et al. (1993). “Stepwise dismantling of adenovirus 2 during
entry into cells.” Cell 75: 477-486.
Li,
E., D. Stupack, et al. (1998). “Adenovirus endocytosis requires actin
cytoskeleton reorganization mediated by Rho family GTPases.” J Virol 72(11): 8806-12.
Smith,
H. M. and N. V. Raikhel (1998). “Nuclear localization signal receptor importin
alpha associates with the cytoskeleton.” Plant Cell 10(11): 1791-9.
Sodeik,
B., M. W. Ebersold, et al. (1997). “Microtubule-mediated transport of incoming
herpes simplex virus 1 capsids to the nucleus.” J Cell Biol 136(5): 1007-21.
Weis,
K. (1998). “Importins and exportins: how to get in and out of the nucleus.”
Trends Biochem Sci 23(5): 185-9.