ALPHAVIRUS-BASED EXPRESSION VECTORS - STRATEGIES AND APPLICATIONS

Alphaviruses are positive-strand RNA viruses that can mediate efficent cytoplasmic gene expression in insect and vertebrate cells. Through r ecombinant DNA technology, the alphavirus RNA replication machinery ha s been engineered for high-level expression of heterologous RNAs and p roteins. Amplification of replication-competent alphavirus RNAs (repli cons) can be initiated by RNA or DNA transfection and a variety of pac kaging systems have been developed for producing high titers of infect ious viral particles. Although normally cytocidal for vertebrate cells , variants with adaptive mutations allowing noncytopathic replication have been isolated from persistently infected cultures or selected usi ng a dominant selectable marker. Such mutations have been mapped and u sed to create new alphavirus vectors for noncytopathic gene expression in mammalian cells. These vectors allow long-term expression at moder ate levels and complement previous vectors designed for short-term hig h-level expression. Besides their use for a growing number of basic re search applications, recombinant alphavirus RNA replicons may also fac ilitate genetic vaccination and transient gene therapy.