Role of cellular actin in human parainfluenza virus type 3 genome transcription.
The transcribing ribonucleoprotein (RNP) complex of human parainfluenza virus type 3 (HPIV-3) requires cellular actin for transcription of viral genome in vitro (De, B. P., Lesoon, A., and Banerjee, A. K. (1991) J. Virol. 65, 3268-3275). In this communication, we have studied the interactions between different molecular forms of actin and the RNP of HPIV-3 to understand the role of actin in mRNA synthesis. We demonstrate that both polymeric and monomeric forms of actin (obtained by DNase I treatment) bind strongly to the RNP at 100 mM KCl concentration (polymerizing buffer). The binding was virtually abolished at zero KCl concentration (depolymerizing buffer). Isolation of the RNP-actin complex and subsequent use in a transcription reaction showed that the bound actin alone was sufficient for mRNA synthesis in vitro. Interestingly, the DNase I-arrested monomeric form of actin failed to activate mRNA synthesis, indicating a requirement of polymerization of the bound actin during HPIV-3 transcription. Electron microscopic analyses revealed that a drastic structural modification of the RNP occurred because of the polymerization of actin from a loosely coiled and irregular structure to a condensed and flexible structure. Activation of transcription was observed also with poly-L-glutamic acid, a highly acidic polypeptide. However, unlike cellular actin, poly-L glutamic acid was able to activate only 10% of the input RNP. These results suggest that cellular actin activates HPIV-3 transcription by polymerizing specifically on the RNP complex. This event results in an alteration of the RNP structure that enhances its suitability for efficient transcription. The acidic domain of actin may play an important role in this process.
De
B P
BP
Department of Molecular Biology, Cleveland Clinic Foundation, Ohio 44195.
Burdsall
A L
AL
Banerjee
A K
AK
D000199:Actins / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D016679:Genome, Viral; D006801:Humans; D010224:Parainfluenza Virus 3, Human / Q000235:genetics*; D011099:Polyglutamic Acid / Q000494:pharmacology; D011485:Protein Binding; D012333:RNA, Messenger / Q000096:biosynthesis / Q000187:drug effects; D012261:Ribonucleoproteins / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D014764:Viral Proteins / Q000378:metabolism*
D000199:Actins; D012333:RNA, Messenger; D012261:Ribonucleoproteins; D014764:Viral Proteins; D011099:Polyglutamic Acid; D003850:Deoxyribonuclease I