Various complexes that contain the core subunits of RNA polymerase II associated with different transcription factors have been isolated from eukaryotes; their precise molecular constitution depends on the purification procedure. We estimated the numbers of various components of such complexes in an HeLa cell by quantitative immunoblotting. The cells were lysed with saponin in a physiological buffer; ϳ140,000 unengaged polymerases (mainly of form IIA) were released. Only ϳ4,000 of these soluble molecules sedimented in glycerol gradients as holoenzyme-sized complexes. About 180,000 molecules of polymerases (ϳ110,000 molecules of form IIO) and 10,000 to 30,000 molecules of each of TFIIB, TFIIE␣, TFIIE␤, TFIIF-RAP74, TFIIF-RAP30, and TFIIH-MAT1 remained tightly associated with the nuclear substructure. Most proteins and run-on activity were retained when ϳ50% of the chromatin was detached with a nuclease, but ϳ45,000 molecules of bound TATA binding protein (TBP) were detached. Similar results were obtained after cross-linking living cells with formaldehyde. The results provide little support for the existence of a large pool of soluble holoenzyme; they are consistent with TBP-promoter complexes in nuclease-sensitive chromatin being assembled into preinitiation complexes attached to the underlying structure.

Sulfur oxides (SO x ) are important atmospheric trace species in both gas and particulate phases, and sulfate is a major component of atmospheric aerosol. One potentially important source of particulate sulfate formation is the oxidation of dissolved SO2 by organic peroxides, which comprises a major fraction of secondary organic aerosol (SOA). In this study, we investigated the reaction kinetics and mechanisms between SO2 and condensed-phase peroxides. pH-dependent aqueous phase reaction rate constants between S­(IV) and organic peroxide standards were measured. Highly oxygenated organic peroxides with O/C > 0.6 in α-pinene SOA react rapidly with S­(IV) species in the aqueous phase. The reactions between organic peroxides and S­(IV) yield both inorganic sulfate and organosulfates (OS), as observed by electrospray ionization ion mobility mass spectrometry. For the first time, 34S-labeling experiments in this study revealed that dissolved SO2 forms OS via direct reactions without forming inorganic sulfate as a reactive intermediate. Kinetics of OS formation was estimated semiquantitatively, and such reaction was found to account for 30–60% of sulfur reacted. The photochemical box model GAMMA was applied to assess the implications of the measured SO2 consumption and OS formation rates. Our findings indicate that this novel pathway of SO2–peroxide reaction is important for sulfate formation in submicron aerosol.

Replication of HIV-1 requires the viral Tat protein, which increases the extent of transcription elongation by RNA polymerase II after activation at the single viral long terminal repeat (LTR) promoter. This effect of Tat on transcription requires Tat interactions with a 5 region (TAR) in nascent transcripts as well as Tat-specific cofactors. The present study identifies a cellular protein, TIP30, that interacts with Tat and with an SRB-containing RNA polymerase II complex both in vivo and in vitro. Coexpression of TIP30 specifically enhances transactivation by Tat in transfected cells, and immunodepletion of TIP30 from nuclear extracts abolishes Tat-activated transcription without affecting Tatindependent transcription. These results implicate TIP30 as a specific coactivator that may enhance formation of a Tat-RNA polymerase II holoenzyme complex.