- 作者: Edward D. Blair; Christopher M. Roberts; Ian Clark-Lewis
- 作者服務機構: a Gene Targets Group, Wellcome Research; Laboratories, Beckenham, UK, and; b The Biomedical Research Centre, Department of Biochemistry, University of; British Columbia, Vancouver, Canada
- 中文摘要: --
- 英文摘要: A 'G-less' cassette was cloned downstream of the human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) to facilitate rapid identification of authentic LTR- directed transcription in T cell nuclear extracts. Despite a high constitutive level of transcrip- tion, addition of chemically synthesized full-length HIV-1bru tat (amino acids 1- 86) stimulat- ed transcription 3-fold but only if the template included the TAR region of the LTR. Suppres- sion of basal transcription in T cell nuclear extracts by sodium citrate increased the relative level of tat stimulation, but neither potassium chloride nor histone H1 had an effect. A mutant synthetic tat polypeptide, lacking residues 22-32 in the cysteine-rich domain, was inactive in uptake assays and failed to stimulate transcription. Short basic peptides that com- peted full-length tat from complexes with TAR RNA also inhibited tat stimulation of tran- scription, whereas short basic peptide unable to bind TAR or compete tat from complexes were also unable to inhibit tat stimulation of transcription. These data confirm that active HIV-1 tat must first interact with TAR RNA via basic amino acid residues in order to stimu- late transcription of downstream sequences.
- 中文關鍵字: --
- 英文關鍵字: tat; TAR RNA; Transcription; G-less