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Identification and characterization of small rnas and proteins expressed by the human parvovirus B19

University of British Columbia

The human pathogenic parvovirus B19 has a strict tissue tropism and will only replicate in a subset of erythroid progenitor cells. However, it is shown that COS-7 cells transfected with SV4O-B19 hybrid vectors express the major B19 RNAs and proteins. In addition, capsid proteins synthesized in these cells self-assemble into virus particles that by EM are morphologically very similar to native B19 virions. Cytoplasmic RNA from transfected COS-7 cells was used to prepare a cDNA library using a method which enriched the library for B19 cDNAs. A second cDNA library was prepared from B19 infected human bone marrow cells that had been isolated from a patient with a chronic myelogenous leukemia using PCR to amplify B19-specific cDNAs. The libraries were probed with a series of RNA probes derived from different regions of the B19 genome in pYT1O3 and selected cDNAs were sequenced and compared. Two size classes of small, abundant, polyadenylated RNAs were identified; the 700 and 800 nt size class of RNA is the product of transcriptional processing in the middle of the B19 genome downstream from an unusual polyadenylation signal, ATTAAA or AATAAC (map unit 49). These transcripts contain an ORF within their second exon which is the same reading frame used for the translation of the nonstructural proteins; however, there are no AUG translation initiation codons within this region of the ORF. The second most abundant size class of RNA in B19 infected cells are the 500 and 600 nt transcripts which are made from three exons and terminated at a normal polyadenylation signal (AATAAA) near the right hand end of the genome (map unit 97). A 94 aa ORF within the third exon is invariant in the two RNA species. [more abstract]

Thesis/Dissertation

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2009-01-05

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http://hdl.handle.net/2429/3329

Biochemistry and Molecular Biology

University of British Columbia

UBCV

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