CITED1 is a novel binding partner of MITF that influences the MITF-directed transcriptional profile in melanoma
- Published
- Accepted
- Subject Areas
- Cell Biology, Genomics, Molecular Biology, Oncology
- Keywords
- MITF, CITED1, Melanoma, Phenotype-switching model, proximity ligation assay, ChIP-seq
- Copyright
- © 2017 Lettiero et al.
- Licence
- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ Preprints) and either DOI or URL of the article must be cited.
- Cite this article
- 2017. CITED1 is a novel binding partner of MITF that influences the MITF-directed transcriptional profile in melanoma. PeerJ Preprints 5:e3146v1 https://doi.org/10.7287/peerj.preprints.3146v1
Abstract
We previously demonstrated how CITED1 knockdown in melanoma cells had the capacity to perturb expression of a significant number of genes that comprised MITF and several of its known transcriptional targets. This manifest as a switch from a more invasive to a more proliferative gene signature phenotype. We now demonstrate by using MITF ChIP-seq, that altered CITED1 expression affects MITF transcription factor binding to its targets across the genome. We show that silencing CITED1 effectively amplifies the MITF chromatin-binding signal response while we also demonstrate for the first time that CITED1 and MITF co-localise in a nuclear complex using an in-situ ligation proximity assay. We propose that CITED1-MITF binding is capable of altering both the affinity of chromatin association and transcriptional response to MITF at the target regions in the genome where MITF is either directly or indirectly bound to DNA. As CITED1/SMAD2 has been shown to mediate TGFβ-driven transcription that induces amoeboid-like invasion in melanoma cells we hypothesis that the MITF/CITED1 driven transcriptional response dominates in MITF-high/low-invasive environment or proliferative signature cell phenotype, whereas the SMAD2/CITED1 transcriptional response is dominant in a low-MITF/ high-invasive signature environment.
Author Comment
This is a preprint submission to PeerJ Preprints.
Supplemental Information
Supplementary Figure S1a
(i). The total protein of each cell fraction visualized on a stain free gel cytoplasmic, (C); nucleoplasmic /nuclear soluble, (NS); nuclear chromatin bound pellet, (NCBp). The strong band in the nuclear bound fraction is the micrococcal nuclease added to degrade the associated DNA before SDS-PAGE. (ii). Validation of the antibodies used to detect CITED1 using Duolink/IF and western blots. AB15096 used in figure 1a has been previously validated using siRNA (Howlin et al., 2015) .
Supplementary Figure S1b
The cell fixation protocol provided by Active Motif
Supplementary Data S2a
The ordered lists of genes from the ChIP-seq heatmap presented in figure 2a
Supplementary Data S2b
Summary statistics for the MITF ChIP-seq data analysis
Supplementary Data S2c
MITF ChIP-seq lists of occupied genes and active regions respectively
Supplementary Data S3a
Known transcription factor motif identification for siNEG MITF ChIP-seq
Supplementary Data S3b
De novo transcription factor motif identification for siNEG MITF ChIP-seq
Supplementary Data S3c
Known transcription factor motif identification for siCITED1 MITF ChIP-seq
Supplementary Data S3d
De novo transcription factor motif identification for siITED1 MITF ChIP-seq
Supplementary Data S4a
UCSC tracks BED file for the MITF-ChIP-seq experiment (siNEG and siCITED1)