Limits to the Independence of Bottom Component RNA of Cowpea Mosaic Virus

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SUMMARYCowpea mosaic virus (CPMV) M RNA is translated in vitro into two polyproteins of Mr values 105000 (105K) and 95K. Using antiserum against the small capsid protein VP23, these proteins have now been detected in cowpea protoplasts, a few hours after inoculation with CPMV. These proteins could also be detected at later stages of infection, but only when proteolytic processing was inhibited by the addition of ZnCI> Using antiserum against a synthetic peptide, corresponding to a part of the overlapping C-terminal ends of the 58K and 48K proteins, the 58K protein, which is the aminoterminal cleavage product of the 105K protein, was found in the cytoplasmic fraction of infected protoplasts. The 48K protein, derived from the 95K protein, was detected in both the cytoplasmic and membrane fractions of protoplasts. The presence of the 105K, 95K, 58K and 48K proteins in CPMV-infected protoplasts indicates that separate initiation codons on the M RNA are used in vivo to produce the 105K and 95K polyproteins, as demonstrated in vitro.

Section: Discussionmentioning

confidence: 99%

Expression of Cowpea Mosaic Virus M RNA in Cowpea Protoplasts

Rezelman

Kammen

Wellink

1989

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“…This M RNA is translated into two overlapping polyproteins of 105K and 95K (Pelham, 1979;Vos et al, 1984;Rezelman et al, 1989), which are proteolytically processed to give the overlapping 58K and 48K nonstructural proteins and, via a 60K precursor, the 37K and 23K coat proteins . The B RNA encodes all the functions necessary for RNA replication (Goldbach et al, 1980;Eggen & van Kammen, 1988) and is able to replicate independently of the M RNA in cowpea protoplasts (Rezelman et al, 1982). Using infectious transcripts derived from cDNA clones of the M RNA it has been shown that both 58K/48K movement and the capsid proteins are required for cellto-cell movement .…”

Section: Introductionmentioning

confidence: 99%

Tubular structures involved in movement of cowpea mosaic virus are also formed in infected cowpea protoplasts

Lent¹,

Storms²,

Meer³

et al. 1991

167

104

In cowpea plant cells infected with cowpea mosaic virus, tubular structures containing virus particles are formed in the plasmodesmata between adjacent cells; these structures are supposedly involved in cell-to-cell spread of the virus. Here we show that similar tubular structures are also formed in cowpea protoplasts, from which the cell wall and plasmodesmata are absent. Between 12 and 21 h post-inoculation, tubule formation starts in the periphery of the protoplast at the level of the plasma membrane. Upon assembly, the viruscontaining tubule is enveloped by the plasma membrane and extends into the culture medium. This suggests that the tubule has functional polarity and makes it likely that a tubule 'grows' into a neighbouring cell in vivo. On average, 75 % of infected protoplasts were shown to possess tubular structures extending from their surface. The tubule wall was 3 to 4 nm thick and they were up to 20 gm in length, as shown by fluorescent light microscopy and negative staining electron microscopy. By analogy to infected plant cells, both the viral 58K/48K movement and capsid proteins were located in these tubules, as determined by immunofluorescent staining and immunogold labelling using specific antisera against these proteins. These results demonstrate that the formation of tubules is not necessarily dependent on the presence of plasmodesmata or the cell wall, and that they are composed, at least in part, of virus-encoded components.

“…However for a successful infection of plants, M RNA is essential. Therefore it probably encodes proteins involved in transport of the virus (Rezelman et al, 1982).…”

Section: Introductionmentioning

confidence: 99%

Cell-to-cell Transport of Cowpea Mosaic Virus Requires Both the 58K/48K Proteins and the Capsid Proteins

Wellink

Kammen

1989

Self Cite

119

SUMMARYInsertions and deletions have been introduced into an infectious cDNA clone of M RNA of cowpea mosaic virus (CPMV), in the coding regions of the 58K/48K and capsid proteins. Transcripts derived from these mutant clones appeared to be replicated in cowpea protoplasts as detected by immunofluorescent staining and Northern blotting. However in cowpea plants, mutations in either region restricted the replication of the viral RNAs to the inoculated cells and thus prevented a successful systemic infection of the plant. These results indicate that the M RNA-encoded 58K/48K proteins are involved in cell-to-cell transport of CPMV, and that the virus can spread only if the RNA is encapsidated in particles.