Limits...
Human ISWI complexes are targeted by SMARCA5 ATPase and SLIDE domains to help resolve lesion-stalled transcription.

Aydin ÖZ, Marteijn JA, Ribeiro-Silva C, Rodríguez López A, Wijgers N, Smeenk G, van Attikum H, Poot RA, Vermeulen W, Lans H - Nucleic Acids Res. (2014)

Bottom Line: Using live cell imaging, we identify a novel function for two distinct mammalian ISWI adenosine triphosphate (ATP)-dependent chromatin remodeling complexes in resolving lesion-stalled transcription.After initial recruitment to UV damage, SMARCA5 re-localizes away from the center of DNA damage, requiring its HAND domain.Our studies support a model in which SMARCA5 targeting to DNA damage-stalled transcription sites is controlled by an ATP-hydrolysis-dependent scanning and proofreading mechanism, highlighting how SWI2/SNF2 chromatin remodelers identify and bind nucleosomes containing damaged DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Medical Genetics Cluster, Cancer Genomics Netherlands, Erasmus MC, Rotterdam, 3015 GE, The Netherlands.

Show MeSH

Related in: MedlinePlus

ACF1 and WSTF function in the transcription-coupled response to UV. Depletion of ACF1 and WSTF renders cells hypersensitive to UV and impairs RRS. Colony survival of U2OS cells treated with siRNAs against ACF1 and CSB (A) and HeLa cells stably expressing shRNAs against WSTF and CSB (B). The percentage of surviving cells is plotted against the applied UV-C dose (J/m2). (C,D) Impaired RRS, 16 h after 6 J/m2 UV-C irradiation, in U2OS cells treated with siRNA against ACF1 or WSTF as measured by EU incorporation. (E) siRNA treatment against ACF1 or WSTF in primary C5RO fibroblasts does not affect UDS, as measured by EdU incorporation after 16 J/m2 UV-C irradiation. Error bars denote standard error of the mean. All results were confirmed using independent, duplicate experiments.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4117783&req=5

Figure 6: ACF1 and WSTF function in the transcription-coupled response to UV. Depletion of ACF1 and WSTF renders cells hypersensitive to UV and impairs RRS. Colony survival of U2OS cells treated with siRNAs against ACF1 and CSB (A) and HeLa cells stably expressing shRNAs against WSTF and CSB (B). The percentage of surviving cells is plotted against the applied UV-C dose (J/m2). (C,D) Impaired RRS, 16 h after 6 J/m2 UV-C irradiation, in U2OS cells treated with siRNA against ACF1 or WSTF as measured by EU incorporation. (E) siRNA treatment against ACF1 or WSTF in primary C5RO fibroblasts does not affect UDS, as measured by EdU incorporation after 16 J/m2 UV-C irradiation. Error bars denote standard error of the mean. All results were confirmed using independent, duplicate experiments.

Mentions: SMARCA5 is the catalytic subunit of several ISWI family ATP-dependent chromatin remodeling complexes including ATP-utilizing Chromatin assembly and remodeling Factor (ACF) (54) and WSTF-ISWI CHromatin remodeling complex (WICH) (55). Both complexes were previously shown to be involved in DSB repair (28,32), while Drosophila ACF complex was also shown to facilitate NER of DNA damage in linker DNA in vitro (56). The human ACF complex consists of SMARCA5 and ACF1 (57), whereas the WICH complex consists of SMARCA5 and WSTF (55) (Supplementary Figure S5A and B). As shown in Figure 6A and B, knock-down of ACF1 and WSTF (Supplementary Figure S5C) rendered cells hypersensitive to UV. In addition, loss of ACF1 and WSTF clearly reduced RRS (Figure 6C and D) but not UDS (Figure 6E). This was achieved using different siRNAs, ruling out off-target effects (Supplementary Figure S5C). These results indicate that ACF1 and WSTF are both involved in TC-NER but not GG-NER, consistent with a function in complex with SMARCA5. Based on these results, we hypothesize that both the ACF and the WICH complex may remodel chromatin during initiation of TC-NER.


Human ISWI complexes are targeted by SMARCA5 ATPase and SLIDE domains to help resolve lesion-stalled transcription.

Aydin ÖZ, Marteijn JA, Ribeiro-Silva C, Rodríguez López A, Wijgers N, Smeenk G, van Attikum H, Poot RA, Vermeulen W, Lans H - Nucleic Acids Res. (2014)

ACF1 and WSTF function in the transcription-coupled response to UV. Depletion of ACF1 and WSTF renders cells hypersensitive to UV and impairs RRS. Colony survival of U2OS cells treated with siRNAs against ACF1 and CSB (A) and HeLa cells stably expressing shRNAs against WSTF and CSB (B). The percentage of surviving cells is plotted against the applied UV-C dose (J/m2). (C,D) Impaired RRS, 16 h after 6 J/m2 UV-C irradiation, in U2OS cells treated with siRNA against ACF1 or WSTF as measured by EU incorporation. (E) siRNA treatment against ACF1 or WSTF in primary C5RO fibroblasts does not affect UDS, as measured by EdU incorporation after 16 J/m2 UV-C irradiation. Error bars denote standard error of the mean. All results were confirmed using independent, duplicate experiments.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4117783&req=5

Figure 6: ACF1 and WSTF function in the transcription-coupled response to UV. Depletion of ACF1 and WSTF renders cells hypersensitive to UV and impairs RRS. Colony survival of U2OS cells treated with siRNAs against ACF1 and CSB (A) and HeLa cells stably expressing shRNAs against WSTF and CSB (B). The percentage of surviving cells is plotted against the applied UV-C dose (J/m2). (C,D) Impaired RRS, 16 h after 6 J/m2 UV-C irradiation, in U2OS cells treated with siRNA against ACF1 or WSTF as measured by EU incorporation. (E) siRNA treatment against ACF1 or WSTF in primary C5RO fibroblasts does not affect UDS, as measured by EdU incorporation after 16 J/m2 UV-C irradiation. Error bars denote standard error of the mean. All results were confirmed using independent, duplicate experiments.
Mentions: SMARCA5 is the catalytic subunit of several ISWI family ATP-dependent chromatin remodeling complexes including ATP-utilizing Chromatin assembly and remodeling Factor (ACF) (54) and WSTF-ISWI CHromatin remodeling complex (WICH) (55). Both complexes were previously shown to be involved in DSB repair (28,32), while Drosophila ACF complex was also shown to facilitate NER of DNA damage in linker DNA in vitro (56). The human ACF complex consists of SMARCA5 and ACF1 (57), whereas the WICH complex consists of SMARCA5 and WSTF (55) (Supplementary Figure S5A and B). As shown in Figure 6A and B, knock-down of ACF1 and WSTF (Supplementary Figure S5C) rendered cells hypersensitive to UV. In addition, loss of ACF1 and WSTF clearly reduced RRS (Figure 6C and D) but not UDS (Figure 6E). This was achieved using different siRNAs, ruling out off-target effects (Supplementary Figure S5C). These results indicate that ACF1 and WSTF are both involved in TC-NER but not GG-NER, consistent with a function in complex with SMARCA5. Based on these results, we hypothesize that both the ACF and the WICH complex may remodel chromatin during initiation of TC-NER.

Bottom Line: Using live cell imaging, we identify a novel function for two distinct mammalian ISWI adenosine triphosphate (ATP)-dependent chromatin remodeling complexes in resolving lesion-stalled transcription.After initial recruitment to UV damage, SMARCA5 re-localizes away from the center of DNA damage, requiring its HAND domain.Our studies support a model in which SMARCA5 targeting to DNA damage-stalled transcription sites is controlled by an ATP-hydrolysis-dependent scanning and proofreading mechanism, highlighting how SWI2/SNF2 chromatin remodelers identify and bind nucleosomes containing damaged DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Medical Genetics Cluster, Cancer Genomics Netherlands, Erasmus MC, Rotterdam, 3015 GE, The Netherlands.

Show MeSH
Related in: MedlinePlus