title: Fate of HIV-1 cDNA intermediates during reverse transcription is dictated by transcription initiation site of virus genomic RNA.
abstract: Retroviral reverse transcription is accomplished by sequential strand-transfers of partial cDNA intermediates copied from viral genomic RNA. Here, we revealed an unprecedented role of 5'-end guanosine (G) of HIV-1 genomic RNA for reverse transcription. Based on current consensus for HIV-1 transcription initiation site, HIV-1 transcripts possess a single G at 5'-ends (G1-form). However, we found that HIV-1 transcripts with additional Gs at 5'-ends (G2- and G3-forms) were abundantly expressed in infected cells by using alternative transcription initiation sites. The G2- and G3-forms were also detected in the virus particle, although the G1-form predominated. To address biological impact of the 5'-G number, we generated HIV clone DNA to express the G1-form exclusively by deleting the alternative initiation sites. Virus produced from the clone showed significantly higher strand-transfer of minus strong-stop cDNA (-sscDNA). The in vitro assay using synthetic HIV-1 RNAs revealed that the abortive forms of -sscDNA were abundantly generated from the G3-form RNA, but dramatically reduced from the G1-form. Moreover, the strand-transfer of -sscDNA from the G1-form was prominently stimulated by HIV-1 nucleocapsid. Taken together, our results demonstrated that the 5'-G number that corresponds to HIV-1 transcription initiation site was critical for successful strand-transfer of -sscDNA during reverse transcription.
authors.affiliation: Department of Immunotherapeutics, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Masuda
authors.affiliation: Department of Immunotherapeutics, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Sato
authors.affiliation: Department of Immunotherapeutics, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Yu-Lun
authors.initials: YL
authors.lastname: Huang
authors.affiliation: Department of Immunotherapeutics, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Koi
authors.affiliation: Department of Immunotherapeutics, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Tatsuro
authors.initials: T
authors.lastname: Takahata
authors.affiliation: Department of Immunotherapeutics, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Atsuhiko
authors.initials: A
authors.lastname: Hasegawa
authors.affiliation: Department of Life and Environmental Sciences, Faculty of Engineering, Chiba Institute of Technology, 2-17-1 Tsudanuma, Narashino-shi, Chiba 275-0016, Japan.
authors.forename: Gota
authors.initials: G
authors.lastname: Kawai
authors.affiliation: Department of Immunotherapeutics, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
authors.forename: Mari
authors.initials: M
authors.lastname: Kannagi
mesh_terms: D018076:DNA, Complementary / Q000235:genetics*; D057809:HEK293 Cells / Q000821:virology; D015497:HIV-1 / Q000235:genetics* / Q000472:pathogenicity; D006801:Humans; D019590:Nucleocapsid Proteins / Q000235:genetics / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics*; D048348:Reverse Transcription; D024363:Transcription Initiation Site
chemical_list: D018076:DNA, Complementary; D019590:Nucleocapsid Proteins; D012367:RNA, Viral