DNA REPLICATION AND GENE EXPRESSION OF CHLORELLA VIRUSES

DESCRIPTION During the past 10
years, the applicant has isolated and partially characterized many large,
polyhedral, dsDNA (>300kb) containing, plaque forming viruses which
infect certain unicellular, eucaryotic, chlorella-like green algae.
These viruses have several unique features, two of which are the focus
of this proposal. (i) They encode DNA methyltransferases and DNA site-
specific (restriction) endonucleases. Some of these endonucleases
recognize and cleave DNA at the same position as bacterial restriction
endonucleases, whereas others have novel specificities. The viruses
also contain nonfunctional DNA methyltransferase genes; the applicant
suspects they contain nonfunctional restriction endonuclease genes as
well. (ii) The viruses contain at least three glycoproteins, including
he major capsid protein.However, unlike other glycoprotein containing
viruses, the chlorella viruses appear to encode putative
glycosyltransferases involved in their glycosylation. The objectives of this proposal are: (i) To continue to isolate and
characterize virus encoded DNA methyltransferases and DNA restriction
endonucleases suspected of recognizing unique DNA sequences and to clone
and sequence their genes. (ii) To determine important domains such as
the target recognition domain in selected DNA methyltransferases and DNA
restriction endonucleases by comparing functional and nonfunctional
genes and making fusion proteins and site-directed mutants. (iii) To
determine if the high homology between certain chlorella virus and
bacterial DNA methyltransferases indicates natural interkingdom gene
exchange between bacteria and the chlorella viruses. (iv)To determine
how the chlorella viruses maintain a constant G + C content despite
having vastly different levels of 5-methylcytosine. (v) To determine by
genetic, molecular genetic, immunological, and biochemical methods those
genes and gene products encoded by the prototype virus, PBCV-1, which
glycosylate the virus major capsid protein.