A chimeric gene containing a cloned cDNA of the coat protein (CP) gene of tobacco mosaic virus (TMV) was introduced into tobacco cells on a Ti plasmid of Agrobacterium tumefaciens from which tumor inducing genes had been removed. Plants regenerated from transformed cells expressed TMV mRNA and CP as a nuclear trait. Seedlings from self-fertilized transgenic plants were inoculated with TMV and observed for development of disease symptoms. The seedlings that expressed the CP gene were delayed in symptom development and 10 to 60 percent of the transgenic plants failed to develop symptoms for the duration of the experiments. Increasing the concentration of TMV in the inoculum shortened the delay in appearance of symptoms. The results of these experiments indicate that plants can be genetically transformed for resistance to virus disease development.

https://science.sciencemag.org/content/sci/232/4751/738.full.pdf

Delay of disease development in transgenic plants that express the tobacco mosaic virus coat protein gene.

The proposed role of the 30-kilodalton(kD) protein of tobacco mosaic virus is to facilitate cell-to-cell spread of the virus-during infection. To directly define the function of the protein, a chimeric gene containing a cloned complementary DNA of the 30-kD protein gene was introduced into tobacco cells via a Ti plasmid-mediated transformation system of Agrobacterium tumefaciens. Transgenic plants regenerated from transformed tobacco cells expressed the 30-kD protein messenger RNA and accumulated 30-kD protein. Seedlings expressing the 30-kD protein gene complemented the Lsl mutant of TMV, a mutant that is temperature-sensitive in cell-to-cell movement. In addition, enhanced movement of the Lsl virus at the permissive temperature was detected in seedlings that express the 30-kD protein gene. These results conclusively demonstrate that the 30-kD protein of tobacco mosaic virus potentiates the movement of the virus from cell to cell.

https://science.sciencemag.org/content/sci/237/4813/389.full.pdf

The 30-kilodalton gene product of tobacco mosaic virus potentiates virus movement.

A remarkable number of cellular activities are directly dependent on ion-pumping ATPases. In addition to the fundamental processes of active transport and ATP synthesis, recent evidence has shown an essential role for ion-pumping ATPases in cell motility, development, ligand-receptor uncoupling, and protein sorting [71, 111,147]. Despite the multiplicity of functions for ATPases, the basic design of these enzymes has been highly conserved. In both eucaryotic and procaryotic organisms, most ionpumping ATPases that have been characterized fit into one of three structural types. In this review we wish to compare these three types of ATPases, drawing upon recent work with fungal cells.

H+-ATPases from mitochondria, plasma membranes, and vacuoles of fungal cells

Publisher Summary The transport of infection is a distinct virus-specific function—transport function (TF)—performed by virus-coded (or virus-induced) proteins. The TF can be complemented in mixed infection with a helper virus; a temperature-resistant helper assists the systemic spreading of the virus temperature-sensitive in TF. Complementation can be achieved with some unrelated viruses—that is, the TF is nonspecific. Systemic spreading of infection takes place by two ways: (1) slow cell-to-cell movement (short distance transport) in the parenchyma and (2) rapid migration over long distances via the conducting tissues (long-distance transport). The long-distance transport of plant viruses occurs usually in the phloem and, in a few cases, in the xylem. The rate of virus transport in the infected tissue appears to be determined by the number of plasmodesmata connecting the neighboring cells; the increase in the number or size or structural alterations of plasmodesmata may favor enhanced transport of infection. Resistance of nonhost plants to viruses can be because of the impossibility of the infection transport (subliminal infection) and the virus being confined to the primarily infected cells. The virus genetic material is transported through the infected plant in the form of special transport particles—virus-specific informosome-like ribonucleoproteins (vRNP) differing from mature virions. These vRNP contain viral genomic and subgenomic RNAs and virus-specific and cell proteins.

Plant Virus-Specific Transport Function And Resistance Of Plants To Viruses

A temperature-sensitive strain of tobacco mosaic virus defective in cell-to-cell movement generates an altered viral-coded protein.

A maize-root microsomal fraction was enriched in ATPase by treatment with Triton X-100. This activity, which reached 1.2–2.0/μmol Pi× min−1× gm protein−1, was specific for ATP, very slightly stimulated by K+, inhibited by orthovanadate and diethylstibestrol, resistant to oligomycin and azide, and had a Km of 1.2 mM MgATP.

Incubation of th emicrosomal fraction with [y32-P]ATP followed by electrophoresis in acid conditions revealed the presence of several phosphoproteins. The phosphorylation of a 110000-M4 polypeptide reached the steady-state level in less than 5 s and rapidly turned over the phosphate group. The phosphorylation level was an hyperbolic function of the [ATP] with a Km of 0.6 mM, suggesting that the rat e of Pi production was proportional to the phosphoprotein concentration. The extent of phosphoprotein was decreased by vanadate and diethylstilbestrol. The phosphorylation level was 30% decreased by 50 mM K+ or Na+ while the ATPase activity was slightly stimulated (12% and 5%. respectively). The polypeptide could not bephosphorylated in reverse by Pi. This phosphorylated intermediate from maize-root microsomes exhibits molecular properties characteristic of transport ATPases such as the yeast plasma membrance H+ -translocating ATPase. This similarity indicates existence of a transport ATPase in plant plasma membranes.

Three other plant microsomal polypeptides (Mr= 52000, 17000 and 16000) and a low molecular weight component (Mr < 1000) were phosphorylated much more slowly, were not undergoing a rapid turnover and were not hydrolysed by hydroxylamine. These phosphoproteins and the Mr < 1000 phosphorylated component were inhibited by vanadate and diethylstilbestrol. These properties are similar to those of the protein kinase activity recently described in yeast plasma membranes.

Phosphorylated Intermediate of a Transport ATPase and Activity of Protein Kinase in Membranes from Corn Roots

An in vivo study of a nonstructural polypeptide synthesized upon TMV infection and its identification with a polypeptide synthesized in vitro from TMV RNA

Sodium dodecyl sulphate-polyacrylamide-gel electrophoretic detection of two high molecular weight proteins associated with tobacco mosaic virus infection in tobacco.

Jacqueline Rigaud

Papers

Phosphorylated Intermediate of a Transport ATPase and Activity of Protein Kinase in Membranes from Corn Roots

An in vivo study of a nonstructural polypeptide synthesized upon TMV infection and its identification with a polypeptide synthesized in vitro from TMV RNA

Sodium dodecyl sulphate-polyacrylamide-gel electrophoretic detection of two high molecular weight proteins associated with tobacco mosaic virus infection in tobacco.