Theiler's murine encephalomyelitis protein 2C and its effect on membrane trafficking
Abstract
Picornaviruses replicate in association with cytoplasmic membranes of infected cells.
Poliovirus 2C and 2BC play an important role in the formation of membranous vesicles, and
induce dramatic changes in membrane trafficking. Theiler’s murine encephalomyelitis virus
protein 2C was localized in infected cells using an anti-TMEV-2C antibody. Early upon
infection, TMEV 2C was localized in the cytoplasm in an ER-like pattern. At later stages, 2C
redistributed to a juxtanuclear site, which represents the viral replication site. Co-localization
with the Golgi complex could not be observed. TMEV 2C seems to interact in vitro with
reticulon 3, a highly conserved ER-associated protein. It was not possible to confirm a
previously identified interaction with AKAP10, a protein kinase anchoring protein, presumably
reflecting conformational constraints of the interaction. Two mutations in the AKAP10 binding
site of TMEV 2C were identified, which inhibit the completion of the infectious cycle of
TMEV. The intracellular changes that occur during TMEV infection were observed. Both actin
filaments and microtubules may be used at early stages of infection; however both cytoskeleton
components accumulate at the periphery of the cell during late stages of infection. A computer-
based analysis has demonstrated that TMEV 2C is highly similar to katanin, a microtubule-
severing protein, and may play a similar role in the reorganization of microtubules during
infection. The Golgi complex turns from a solid, crescent-shaped organelle, into a series of
punctuate fluorescent points forming an expanding balloon-like structure surrounding the
concomitantly expanding site of virus replication. The remnants of the Golgi complex are
finally dispersed throughout the cytoplasm. Live imaging confirmed these findings. It was
observed that PKA also undergoes displacement to the cell periphery during infection.
However, BIG1 seems to locate to the viral replication site during infection, suggesting it may
play a role during viral replication. The localization of PKA and BIG1 in the infected cell may
in part explain the observed dispersion of the Golgi complex.
Type
Thesis, PhD Doctor of Philosophy
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