Date of Award
Doctor of Philosophy
Parameters controlling the initiation of replication of the neurotropic Coranavirus JHMV were investigated to further elucidate the determinants conferring permissiveness or resistance of cells from the central nervous system. Two fines of enquiry pertaining to the early events of JHMV infection were investigated.;One aspect, the mechanism of JHMV penetration into host cells was examined with the fully permissive L-2 fibroblasts as the host. Several inhibitors including phenylarsine oxide, chlorpromazine, trifluoperazine, hypertonic solution and bafilomycin A1 affecting various stages of the endocytic pathway were used to affect penetration. These experimental approaches revealed that most probably JHMV is internalized through receptor-mediated endocytosis so as to initiate the infectious process within early rather than late acidic endosomes.;The second topic dealt with an early event restricting infection in mature oligodendrocytes. Upregulation in expression of the regulatory R1 subunit of cAMP-dependent protein kinase A; associated with maturation of these host cells, caused inhibition of a phosphoprotein phosphatase (PPPase) associated with endosomes. This activity can dephosphorylate in vitro nucleocapsid (N) protein of JHMV leading to an investigation of a modulation of JHMV replication by R1 and PPPase. Inhibition with calyculin A, revealed that dephosphorylation of N is catalysed by a type 2A PPPaase and occurs during penetration and uncoating of inoculum virions. A correlation between high levels of R1 in L-2 and inhibition of dephosphorylation of N with coincident reduction of JHMV replication, suggests that normally JHMV inoculum is uncoated after entering early endosomes due to dephosphorylation of N. By preventing the uncoating process the R subunit interferes with initiation of replication.;Additional but unrelated studies examined the significance of the amino acid sequence relatedness between N, and microtubule-associated protein (MAP) tau which is axon-specific, and another, MAP4, present in neurites in the trafficking of JHMV within neurons. Using material obtained from the neuronal cell line OBL-21, it was shown that sequence relatedness between N, the MAPs extended to immunological cross-reactivity. By demonstrating that N can interact in vitro with purified tubulin, due to a mimicry there may exist a functional basis for the association between N and microtubules within neurons, as revealed by the electron microscope.;The results of this study imply that, due to their nature as obligatory parasites, viruses have evolved strategies adapted towards utilization of host specific structures and functions necessary for their replication in selected organs such as the CNS.
Kalicharran, Kishna Kumar, "Parameters Controlling The Initiation Of Replication Of Coronavirus Jhm In Cells Of The Central Nervous System" (1996). Digitized Theses. 2702.