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Mechanism of start site selection by RNA polymerase II: interplay between TFIIB and Ssl2/XPB helicase subunit of TFIIH.

ABSTRACT: TFIIB is essential for transcription initiation by RNA polymerase II. TFIIB also cross-links to terminator regions and is required for gene loops that juxtapose promoter-terminator elements in a transcription-dependent manner. The Saccharomyces cerevisiae sua7-1 mutation encodes an altered form of TFIIB (E62K) that is defective for both start site selection and gene looping. Here we report the isolation of an ssl2 mutant, encoding an altered form of TFIIH, as a suppressor of the cold-sensitive growth defect of the sua7-1 mutation. Ssl2 (Rad25) is orthologous to human XPB and is a member of the SF2 family of ATP-dependent DNA helicases. The ssl2 suppressor allele encodes an arginine replacement of the conserved histidine residue (H508R) located within the DEVH-containing helicase domain. In addition to suppressing the TFIIB E62K growth defect, Ssl2 H508R partially restores both normal start site selection and gene looping. Moreover, Ssl2, like TFIIB, associates with promoter and terminator regions, and the diminished association of TFIIB E62K with the PMA1 terminator is restored by the Ssl2 H508R suppressor. These results define a novel, functional interaction between TFIIB and Ssl2 that affects start site selection and gene looping.

SUBMITTER: Goel S

PROVIDER: S-EPMC3249109 | biostudies-literature | 2012 Jan

REPOSITORIES: biostudies-literature

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Mechanism of start site selection by RNA polymerase II: interplay between TFIIB and Ssl2/XPB helicase subunit of TFIIH.

Goel Shivani S Krishnamurthy Shankarling S Hampsey Michael M

The Journal of biological chemistry 20111111 1

TFIIB is essential for transcription initiation by RNA polymerase II. TFIIB also cross-links to terminator regions and is required for gene loops that juxtapose promoter-terminator elements in a transcription-dependent manner. The Saccharomyces cerevisiae sua7-1 mutation encodes an altered form of TFIIB (E62K) that is defective for both start site selection and gene looping. Here we report the isolation of an ssl2 mutant, encoding an altered form of TFIIH, as a suppressor of the cold-sensitive g ...[more]

PMID: 22081613

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Project description:AimsSpironolactone (SPL) improves endothelial dysfunction and survival in heart failure. Immune modulation, including poorly understood mineralocorticoid receptor (MR)-independent effects of SPL might contribute to these benefits and possibly be useful in other inflammatory cardiovascular diseases such as pulmonary arterial hypertension.Methods and resultsUsing human embryonic kidney cells (HEK 293) expressing specific nuclear receptors, SPL suppressed NF-κB and AP-1 reporter activity independent of MR and other recognized nuclear receptor partners. NF-κB and AP-1 DNA binding were not affected by SPL and protein synthesis blockade did not interfere with SPL-induced suppression of inflammatory signalling. In contrast, proteasome blockade to inhibit degradation of xeroderma pigmentosum group B complementing protein (XPB), a subunit of the general transcription factor TFIIH, or XPB overexpression both prevented SPL-mediated suppression of inflammation. Similar to HEK 293 cells, a proteasome inhibitor blocked XPB loss and SPL suppression of AP-1 induced target genes in human pulmonary artery endothelial cells (PAECs). Unlike SPL, eplerenone (EPL) did not cause XPB degradation and failed to similarly suppress inflammatory signalling. SPL combined with siRNA XPB knockdown further reduced XPB protein levels and had the greatest effect on PAEC inflammatory gene transcription. Using chromatin-immunoprecipitation, PAEC target gene susceptibility to SPL was associated with low basal RNA polymerase II (RNAPII) occupancy and TNFα-induced RNAPII and XPB recruitment. XP patient-derived fibroblasts carrying an N-terminal but not C-terminal XPB mutations were insensitive to both SPL-mediated XPB degradation and TNFα-induced target gene suppression. Importantly, SPL treatment decreased whole lung XPB protein levels in a monocrotaline rat model of pulmonary hypertension and reduced inflammatory markers in an observational cohort of PAH patients.ConclusionSPL has important anti-inflammatory effects independent of aldosterone and MR, not shared with EPL. Drug-induced, proteasome-dependent XPB degradation may be a useful therapeutic approach in cardiovascular diseases driven by inflammation.

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Mechanism of start site selection by RNA polymerase II: interplay between TFIIB and Ssl2/XPB helicase subunit of TFIIH.

Publications

Mechanism of start site selection by RNA polymerase II: interplay between TFIIB and Ssl2/XPB helicase subunit of TFIIH.

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