Dataset Information

KLF8 recruits the p300 and PCAF co-activators to its amino terminal activation domain to activate transcription.

ABSTRACT: Krüppel-like factor 8 (KLF8) regulates critical cellular processes including cell cycle progression, transformation, epithelial-to-mesenchymal transition, migration and invasion by either repressing or activating target gene promoters. As a repressor, KLF8 recruits the CtBP co-repressor via its PVDLS repression motif. However, how KLF8 acts as an activator has not been determined. Here we report the identification of both the KLF8 activation domain and associated co-activators. By site-directed mutagenesis and cyclin D1 promoter reporter assays using both mouse fibroblasts and human epithelial cells, we determined that deletion of residues 100-260 or mutation of Q118-Q248 abolished KLF8 transactivity. this transactivity was dramatically reduced in p300(-/-), CBP(-/-) or PCAF(-/-) cells and could be restored by re-expressing p300 or PCAF, but not CBP. Co-immunoprecipitation analyses demonstrated that KLF8 interacted with these co-activators whereas the Q118N-Q248N mutant did not. Chromatin immunoprecipitation experiments showed that KLF8 promoted histone acetylation at the promoter whereas the Q118N-Q248N mutant had a dramatic loss of this function. Western blotting revealed that unlike wild-type KLF8 the Q118N-Q248N was no longer able to upregulate cyclin D1 protein level. BrdU incorporation assays showed that the Q118N-Q248N mutant also lost the ability to promote DNA synthesis. Taken together, these results identified the KLF8 activation domain located between residues 101-260 where the well-conserved Q118 and Q248 are essential for recruiting p300 and PCAF to activate target gene transcription.

SUBMITTER: Urvalek AM

PROVIDER: S-EPMC2888133 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Human telomerase reverse transcriptase (hTERT) plays an extremely important role in cancer initiation and development, including colorectal cancer (CRC). However, the precise upstream regulatory mechanisms of hTERT in different cancer types remain poorly understood. Here, we uncovered the candidate transcriptional factor of hTERT in CRC and explored its role and the corresponding molecular mechanisms in regulating hTERT expression and CRC survival with an aim of developing mechanism-based combinational targeting therapy. The possible binding proteins at the hTERT promoter were uncovered using pull-down/mass spectrometry analysis. The regulation of SPT6 on hTERT expression and CRC survival was evaluated in human CRC cell lines and mouse models. Mechanistic studies focusing on the synergy between SPT6 and staphylococcal nuclease and Tudor domain containing 1 (SND1) in controlling hTERT expression and CRC progression were conducted also in the above two levels. The expression correlation and clinical significance of SPT6, SND1, and hTERT were investigated in tumor tissues from murine models and patients with CRC in situ. SPT6 was identified as a possible transcriptional factor to bind to the hTERT promoter. SPT6 knockdown decreased the activity of hTERT promoter, downregulated the protein expression level of hTERT, suppressed proliferation, invasion, and stem-like properties, promoted apoptosis induction, and enhanced chemotherapeutic drug sensitivity in vitro. SPT6 silencing also led to the delay of tumor growth and metastasis in mice carrying xenografts of human-derived colon cancer cells. Mechanistically, SND1 interacted with SPT6 to co-control hTERT expression and CRC cell proliferation, stemness, and growth in vitro and in vivo. SPT6, SND1, and hTERT were highly expressed simultaneously in CRC tissues, both from the murine model and patients with CRC in situ, and pairwise expression among these three factors showed a significant positive correlation. In brief, our research demonstrated that SPT6 synergized with SND1 to promote CRC development by targeting hTERT and put forward that inhibiting the SPT6-SND1-hTERT axis may create a therapeutic vulnerability in CRC.

Dataset Information

KLF8 recruits the p300 and PCAF co-activators to its amino terminal activation domain to activate transcription.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets