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A conserved alpha-herpesvirus protein necessary for axonal localization of viral membrane proteins.

Tomishima MJ, Enquist LW - J. Cell Biol. (2001)

Bottom Line: We conclude that the Us9 membrane protein controls axonal localization of diverse viral membrane proteins but not that of capsid or tegument proteins.The data support a model where virion subassemblies but not complete virions are transported in the axon.Our results provide new insight into the process of virion assembly and exit from neurons that leads to directional spread of herpesviruses in the nervous system.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA.

ABSTRACT
Pseudorabies virus, an alpha-herpesvirus, is capable of infecting the nervous system and spreading between synaptically connected neurons in diverse hosts. At least three viral membrane proteins (gE, gI, and Us9) are necessary for the spread of infection from presynaptic to postsynaptic neurons (anterograde spread) in infected rodents. To understand how these proteins effect anterograde spread between neurons, we analyzed the subcellular localization of viral proteins after infection of cultured rat sympathetic neurons with wild-type or mutant viruses. After Us9- mutant infections but not gE- mutant infections, only a subset of the viral structural proteins had entered axons. Surprisingly, capsid and tegument proteins but not viral membrane proteins were detected in axons. The spread of Us9 missense mutants in the rodent nervous system correlated with the amount of viral membrane proteins localized to axons. We conclude that the Us9 membrane protein controls axonal localization of diverse viral membrane proteins but not that of capsid or tegument proteins. The data support a model where virion subassemblies but not complete virions are transported in the axon. Our results provide new insight into the process of virion assembly and exit from neurons that leads to directional spread of herpesviruses in the nervous system.

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Related in: MedlinePlus

Characterization of cultured SCG neurons. Phase–contrast images of neurons after 3 h (A), 5 d (B), or 21 d (C) in culture. The establishment of cell polarity of SCG neurons after 21 d in culture is demonstrated in D–K. After fixing and permeabilizing, the neurons were labeled with antibodies that recognized the axonal markers tau-1 (D and H) and phospho-M and H neurofilament (E and I) or the somatodendritic markers MAP2 (F and J) and nonphospho-M and H neurofilament (G and K). Two confocal images of each field are shown: D–G have image planes at the level of the cell bodies, whereas H–K show the same neuron at the level of the substratum so that axons can be visualized. Bar, 25 μm.
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fig11: Characterization of cultured SCG neurons. Phase–contrast images of neurons after 3 h (A), 5 d (B), or 21 d (C) in culture. The establishment of cell polarity of SCG neurons after 21 d in culture is demonstrated in D–K. After fixing and permeabilizing, the neurons were labeled with antibodies that recognized the axonal markers tau-1 (D and H) and phospho-M and H neurofilament (E and I) or the somatodendritic markers MAP2 (F and J) and nonphospho-M and H neurofilament (G and K). Two confocal images of each field are shown: D–G have image planes at the level of the cell bodies, whereas H–K show the same neuron at the level of the substratum so that axons can be visualized. Bar, 25 μm.

Mentions: Many types of neurons can not sort their proteins to axonal or dendritic compartments until at least 2 wk of growth in vitro (Caceres et al., 1986; Goslin et al., 1990; Ledesma et al., 1999). Rat sympathetic neurons derived from the SCG are reported to sort endogenous proteins to axonal and somatodendritic compartments after 3 wk in vitro (Bruckenstein and Higgins, 1988). The polarity of cultured neurons used in this report was determined by analysis of three control proteins: (a) the axonal markers tau-1 (Boehringer) and SMI-31 (Sternberger-Meyer Immunochemicals), and (b) the cell body/dendrite markers MAP2 (Boehringer) and SMI-32 (Sternberger-Meyer Immunochemicals). The antibody SMI-31 recognizes the phosphoforms of the M and H neurofilament, whereas the antibody SMI-32 reacts with the nonphosphoforms of the M and H neurofilament (Sternberger and Sternberger, 1983). Since neuronal cell bodies are raised as much as 20 μm from the floor of the tissue culture dish where the axons grow, two different image planes are shown in Fig. 11, D–K : the top panels (Fig. 11, D–G) show an image plane at the center of the cell bodies (above the substratum), whereas the bottom panels (Fig. 11, H–K) have the image plane at the substratum where the neurites can be visualized.


A conserved alpha-herpesvirus protein necessary for axonal localization of viral membrane proteins.

Tomishima MJ, Enquist LW - J. Cell Biol. (2001)

Characterization of cultured SCG neurons. Phase–contrast images of neurons after 3 h (A), 5 d (B), or 21 d (C) in culture. The establishment of cell polarity of SCG neurons after 21 d in culture is demonstrated in D–K. After fixing and permeabilizing, the neurons were labeled with antibodies that recognized the axonal markers tau-1 (D and H) and phospho-M and H neurofilament (E and I) or the somatodendritic markers MAP2 (F and J) and nonphospho-M and H neurofilament (G and K). Two confocal images of each field are shown: D–G have image planes at the level of the cell bodies, whereas H–K show the same neuron at the level of the substratum so that axons can be visualized. Bar, 25 μm.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196449&req=5

fig11: Characterization of cultured SCG neurons. Phase–contrast images of neurons after 3 h (A), 5 d (B), or 21 d (C) in culture. The establishment of cell polarity of SCG neurons after 21 d in culture is demonstrated in D–K. After fixing and permeabilizing, the neurons were labeled with antibodies that recognized the axonal markers tau-1 (D and H) and phospho-M and H neurofilament (E and I) or the somatodendritic markers MAP2 (F and J) and nonphospho-M and H neurofilament (G and K). Two confocal images of each field are shown: D–G have image planes at the level of the cell bodies, whereas H–K show the same neuron at the level of the substratum so that axons can be visualized. Bar, 25 μm.
Mentions: Many types of neurons can not sort their proteins to axonal or dendritic compartments until at least 2 wk of growth in vitro (Caceres et al., 1986; Goslin et al., 1990; Ledesma et al., 1999). Rat sympathetic neurons derived from the SCG are reported to sort endogenous proteins to axonal and somatodendritic compartments after 3 wk in vitro (Bruckenstein and Higgins, 1988). The polarity of cultured neurons used in this report was determined by analysis of three control proteins: (a) the axonal markers tau-1 (Boehringer) and SMI-31 (Sternberger-Meyer Immunochemicals), and (b) the cell body/dendrite markers MAP2 (Boehringer) and SMI-32 (Sternberger-Meyer Immunochemicals). The antibody SMI-31 recognizes the phosphoforms of the M and H neurofilament, whereas the antibody SMI-32 reacts with the nonphosphoforms of the M and H neurofilament (Sternberger and Sternberger, 1983). Since neuronal cell bodies are raised as much as 20 μm from the floor of the tissue culture dish where the axons grow, two different image planes are shown in Fig. 11, D–K : the top panels (Fig. 11, D–G) show an image plane at the center of the cell bodies (above the substratum), whereas the bottom panels (Fig. 11, H–K) have the image plane at the substratum where the neurites can be visualized.

Bottom Line: We conclude that the Us9 membrane protein controls axonal localization of diverse viral membrane proteins but not that of capsid or tegument proteins.The data support a model where virion subassemblies but not complete virions are transported in the axon.Our results provide new insight into the process of virion assembly and exit from neurons that leads to directional spread of herpesviruses in the nervous system.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA.

ABSTRACT
Pseudorabies virus, an alpha-herpesvirus, is capable of infecting the nervous system and spreading between synaptically connected neurons in diverse hosts. At least three viral membrane proteins (gE, gI, and Us9) are necessary for the spread of infection from presynaptic to postsynaptic neurons (anterograde spread) in infected rodents. To understand how these proteins effect anterograde spread between neurons, we analyzed the subcellular localization of viral proteins after infection of cultured rat sympathetic neurons with wild-type or mutant viruses. After Us9- mutant infections but not gE- mutant infections, only a subset of the viral structural proteins had entered axons. Surprisingly, capsid and tegument proteins but not viral membrane proteins were detected in axons. The spread of Us9 missense mutants in the rodent nervous system correlated with the amount of viral membrane proteins localized to axons. We conclude that the Us9 membrane protein controls axonal localization of diverse viral membrane proteins but not that of capsid or tegument proteins. The data support a model where virion subassemblies but not complete virions are transported in the axon. Our results provide new insight into the process of virion assembly and exit from neurons that leads to directional spread of herpesviruses in the nervous system.

Show MeSH
Related in: MedlinePlus