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Chemical perturbation of an intrinsically disordered region of TFIID distinguishes two modes of transcription initiation.

Zhang Z, Boskovic Z, Hussain MM, Hu W, Inouye C, Kim HJ, Abole AK, Doud MK, Lewis TA, Koehler AN, Schreiber SL, Tjian R - Elife (2015)

Bottom Line: They are abundant in eukaryotic proteomes and are often associated with human diseases, but their biological functions have been elusive to study.Binding arrests an isomerization of promoter-bound TFIID that is required for the engagement of Pol II during the first (de novo) round of transcription initiation.This work also suggests a new avenue for targeting the elusive IDRs by harnessing certain features of metal-based complexes for mechanistic studies, and for the development of novel pharmaceutical interventions.

View Article: PubMed Central - PubMed

Affiliation: Transcription Imaging Consortium, Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States.

ABSTRACT
Intrinsically disordered proteins/regions (IDPs/IDRs) are proteins or peptide segments that fail to form stable 3-dimensional structures in the absence of partner proteins. They are abundant in eukaryotic proteomes and are often associated with human diseases, but their biological functions have been elusive to study. In this study, we report the identification of a tin(IV) oxochloride-derived cluster that binds an evolutionarily conserved IDR within the metazoan TFIID transcription complex. Binding arrests an isomerization of promoter-bound TFIID that is required for the engagement of Pol II during the first (de novo) round of transcription initiation. However, the specific chemical probe does not affect reinitiation, which requires the re-entry of Pol II, thus, mechanistically distinguishing these two modes of transcription initiation. This work also suggests a new avenue for targeting the elusive IDRs by harnessing certain features of metal-based complexes for mechanistic studies, and for the development of novel pharmaceutical interventions.

No MeSH data available.


Related in: MedlinePlus

The originally purchased lead compound specifically blocks de novo transcription initiation at the step of Pol II engagement.Refer to Figure 6 legends for more details. The inhibitor used here was ChemDiv 7241-4207 (original purchase).DOI:http://dx.doi.org/10.7554/eLife.07777.015
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fig6s1: The originally purchased lead compound specifically blocks de novo transcription initiation at the step of Pol II engagement.Refer to Figure 6 legends for more details. The inhibitor used here was ChemDiv 7241-4207 (original purchase).DOI:http://dx.doi.org/10.7554/eLife.07777.015

Mentions: Interestingly, pre-incubation of TFIID or TFIID together with TFIIB with the DNA template led to a more severe inhibition (16-fold, Figure 6C, lanes 4 and 6) when compared to adding all the protein factors after the inhibitor (6.5-fold, lane 2). This suggests that a TFIID-TFIIB-DNA sub-complex is likely the most susceptible substrate for inhibition, while the presence of other core promoter factors may facilitate the progression of PIC assembly and reduce the time interval of this vulnerable stage. Addition of TFIIF partially alleviated the inhibition (comparing lanes 6 and 8), suggesting that TFIIF may play a necessary but insufficient role in driving TFIID into an inhibitor-resistant state. Upon further addition of Pol II to the assembly, the system became resistant to the inhibitor to a level comparable to that of preformed PICs (lanes 10 and 14). In contrast to the essential role of TFIID, TFIIB, TFIIF, and Pol II (the same set of factors required for Pol II engagement) in forming the minimal PIC intermediate resistant to inhibition, adding TFIIE and TFIIH had no effect on inhibition (comparing lane 10, 12, and 14), even though these two factors are required for robust transcription. Very similar results were observed when individual protein factors were left out one by one from GTF set 1 as an alternative strategy to arrest PIC assembly at specific stages (Figure 6C lanes 11–24 for the pure tin(IV) oxochloride compound; and Figure 6—figure supplement 1 for the original lead compound ChemDiv 7241-4207). Taken together, these results strongly suggest that the step most sensitive to inhibition involves the initial binding of Pol II to a DNA-TFIID complex that must then transition into a conformation compatible with productive Pol II engagement. Importantly, after Pol II engagement takes place during de novo PIC assembly, the system becomes resistant to the inhibitor.


Chemical perturbation of an intrinsically disordered region of TFIID distinguishes two modes of transcription initiation.

Zhang Z, Boskovic Z, Hussain MM, Hu W, Inouye C, Kim HJ, Abole AK, Doud MK, Lewis TA, Koehler AN, Schreiber SL, Tjian R - Elife (2015)

The originally purchased lead compound specifically blocks de novo transcription initiation at the step of Pol II engagement.Refer to Figure 6 legends for more details. The inhibitor used here was ChemDiv 7241-4207 (original purchase).DOI:http://dx.doi.org/10.7554/eLife.07777.015
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4582147&req=5

fig6s1: The originally purchased lead compound specifically blocks de novo transcription initiation at the step of Pol II engagement.Refer to Figure 6 legends for more details. The inhibitor used here was ChemDiv 7241-4207 (original purchase).DOI:http://dx.doi.org/10.7554/eLife.07777.015
Mentions: Interestingly, pre-incubation of TFIID or TFIID together with TFIIB with the DNA template led to a more severe inhibition (16-fold, Figure 6C, lanes 4 and 6) when compared to adding all the protein factors after the inhibitor (6.5-fold, lane 2). This suggests that a TFIID-TFIIB-DNA sub-complex is likely the most susceptible substrate for inhibition, while the presence of other core promoter factors may facilitate the progression of PIC assembly and reduce the time interval of this vulnerable stage. Addition of TFIIF partially alleviated the inhibition (comparing lanes 6 and 8), suggesting that TFIIF may play a necessary but insufficient role in driving TFIID into an inhibitor-resistant state. Upon further addition of Pol II to the assembly, the system became resistant to the inhibitor to a level comparable to that of preformed PICs (lanes 10 and 14). In contrast to the essential role of TFIID, TFIIB, TFIIF, and Pol II (the same set of factors required for Pol II engagement) in forming the minimal PIC intermediate resistant to inhibition, adding TFIIE and TFIIH had no effect on inhibition (comparing lane 10, 12, and 14), even though these two factors are required for robust transcription. Very similar results were observed when individual protein factors were left out one by one from GTF set 1 as an alternative strategy to arrest PIC assembly at specific stages (Figure 6C lanes 11–24 for the pure tin(IV) oxochloride compound; and Figure 6—figure supplement 1 for the original lead compound ChemDiv 7241-4207). Taken together, these results strongly suggest that the step most sensitive to inhibition involves the initial binding of Pol II to a DNA-TFIID complex that must then transition into a conformation compatible with productive Pol II engagement. Importantly, after Pol II engagement takes place during de novo PIC assembly, the system becomes resistant to the inhibitor.

Bottom Line: They are abundant in eukaryotic proteomes and are often associated with human diseases, but their biological functions have been elusive to study.Binding arrests an isomerization of promoter-bound TFIID that is required for the engagement of Pol II during the first (de novo) round of transcription initiation.This work also suggests a new avenue for targeting the elusive IDRs by harnessing certain features of metal-based complexes for mechanistic studies, and for the development of novel pharmaceutical interventions.

View Article: PubMed Central - PubMed

Affiliation: Transcription Imaging Consortium, Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States.

ABSTRACT
Intrinsically disordered proteins/regions (IDPs/IDRs) are proteins or peptide segments that fail to form stable 3-dimensional structures in the absence of partner proteins. They are abundant in eukaryotic proteomes and are often associated with human diseases, but their biological functions have been elusive to study. In this study, we report the identification of a tin(IV) oxochloride-derived cluster that binds an evolutionarily conserved IDR within the metazoan TFIID transcription complex. Binding arrests an isomerization of promoter-bound TFIID that is required for the engagement of Pol II during the first (de novo) round of transcription initiation. However, the specific chemical probe does not affect reinitiation, which requires the re-entry of Pol II, thus, mechanistically distinguishing these two modes of transcription initiation. This work also suggests a new avenue for targeting the elusive IDRs by harnessing certain features of metal-based complexes for mechanistic studies, and for the development of novel pharmaceutical interventions.

No MeSH data available.


Related in: MedlinePlus