Human cytomegalovirus IE2 drives transcription initiation from a select subset of late infection viral promoters by host RNA polymerase II

Author summary The herpesvirus subfamilies differ in the viral proteins used in generating the cascade of viral immediate-early, early, early-late, or late gene transcription. With the application of advanced technologies, we discovered that the betaherpesvirus, human cytomegalovirus, has evolved strategies analogous to those used by both alpha- and gammaherpesviruses to bring about RNA Pol II transcription from its late infection promoters. Like alphaherpesviruses, human cytomegalovirus purposes a pivotal immediate-early viral transcription factor to initiate transcription from early, early-late, and late viral promoters. However, the cytomegalovirus transcription factor only targets a select set of viral early-late and late promoters without appreciably affecting host promoters at late times. Most of these late infection viral promoters are structurally and mechanistically different from promoters activated by the 6-member viral transcription factor complex that is analogous to the transcription factor complex utilized by gammaherpesviruses. Human cytomegalovirus genome amplification must first take place, but need not continue, to enable the two different mechanisms of late viral promoter activation. Herpesvirus late promoters activate gene expression after viral DNA synthesis has begun. Alphaherpesviruses utilize a viral immediate-early protein to do this, whereas beta- and gammaherpesviruses primarily use a 6-member set of viral late-acting transcription factors (LTF) that are drawn to a TATT sequence in the late promoter. The betaherpesvirus, human cytomegalovirus (HCMV), produces three immediate-early 2 protein isoforms, IE2-86, IE2-60, IE2-40, late in infection, but whether they activate late viral promoters is unknown. Here, we quickly degrade the IE2 proteins in late infection using dTag methodology and analyze effects on transcription using customized PRO-Seq and computational methods combined with multiple validation methods. We discover that the IE2 proteins selectively drive RNA Pol II transcription initiation at a subset of viral early-late and late promoters common to different HCMV strains, but do not substantially affect Pol II transcription of the 9,942 expressed host genes. Most of the IE2-activated viral late infection promoters lack the TATT sequence bound by the HCMV UL87-encoded LTF. The HCMV TATT-binding protein is not mechanistically involved in late RNA expression from the IE2-activated TATT-less UL83 (pp65) promoter, as it is fo