Identification and characterization of the IE-N gene from the Autographa californica Nuclear Polyhedrosis Virus /
| Main Author: | |
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| Other Authors: | , , |
| Format: | Thesis Book |
| Language: | English |
| Published: |
1990.
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| Online Access: | Link to ProQuest copy Link to OAKTrust copy |
| Abstract: | To study temporal regulation of Autographa californica Nuclear Polyhedrosis Virus (AcMNPV) genes, the delayed early 39K gene was cloned from AcMNPV and linked to the chloramphenicol acetyltransferase (CAT) reporter gene. Co-transfection with viral DNA digested with BglII suggested that a viral element was involved in the activation of the delayed early 39K promoter. This BglII-sensitive element, was localized to the PstI-N fragment of AcMNPV DNA. A 1330 nucleotide RNA spanning the BglII site was transcribed when the PstI-N fragment was transiently expressed in S. frugiperda cells. This gene encoded by the PstI-N fragment was designated IE-N. The nucleotide sequence of the IE-N gene was determined and shown to encode a serine- and glutamine-rich protein with a predicted molecular weight of 47,000. A polypeptide produced by the IE-N coding region can be detected by in vitro translation, radio-immunoprecipitation detection of an IE-NCAT fusion protein, and by protein activity. IE-N mRNA was abundantly expressed early, but not late, after infection with AcMNPV. In addition, IE-N mRNA and activity can be detected in S. frugiperda cells transiently expressing the PstI-N fragment. Because IE-N did not require the synthesis of AcMNPV gene products to be expressed, IE-N is an immediate early gene. Three viral factors regulated the transient expression of IE-N. These factors were the viral enhancer hr1, immediate early gene IE-1, and the gene product of IE-N. The hr1 stimulated expression when linked in cis to the IE-N gene. This enhancement was independent of position and orientation with respect to the IE-N gene. Co-expression of IE-1 with IE-N reduced the overall expression of IE-N. This decrease in IE-N expression by IE-1 was mediated by the hr1 enhancer sequences linked to the IE-N gene. The IE-N gene product increased expression from the IE-N gene. This stimulation was dependent upon the promoter of IE-N. In addition to increasing its own expression and that of the delayed early gene 39K, IE-N also stimulated expression of the immediate early IE-1 gene. IE-N, therefore, appears to produce a protein which functions in activating AcMNPV immediate early gene expression and possibly delayed early gene expression, as well. |
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| Item Description: | "Major subject: Biochemistry." Typescript (photocopy). Vita. |
| Physical Description: | x, 105 leaves : illustrations ; 29 cm |
| Bibliography: | Includes bibliographical references. |