Abstract
CITED1 is a non-DNA binding transcriptional co-regulator whose expression can distinguish the ‘proliferative’ from ‘invasive’ signature in the phenotype-switching model of melanoma. We have found that, in addition to other ‘proliferative’ signature genes, CITED1 expression is repressed by TGFβ while the ‘invasive’ signature genes are upregulated. In agreement, CITED1 positively correlates with MITF expression and can discriminate the MITF-high/pigmentation tumour molecular subtype in a large cohort (120) of melanoma cell lines. Interestingly, CITED1 overexpression significantly suppressed MITF promoter activation, mRNA and protein expression levels while MITF was transiently upregulated following siRNA mediated CITED1 silencing. Conversely, MITF siRNA silencing resulted in CITED1 downregulation indicating a reciprocal relationship. Whole genome expression analysis identified a phenotype shift induced by CITED1 silencing and driven mainly by expression of MITF and a cohort of MITF target genes that were significantly altered. Concomitantly, we found changes in the cell-cycle profile that manifest as transient G1 accumulation, increased expression of CDKN1A and a reduction in cell viability. Additionally, we could predict survival outcome by classifying primary melanoma tumours using our in vitro derived ‘CITED1-silenced’ gene expression signature. We hypothesize that CITED1 acts a regulator of MITF, functioning to maintain MITF levels in a range compatible with tumourigenesis.
Highlights
CITED1 is the founding member of the CITED (CBP/p300-interacting transactivator with glutamic acid [E]/aspartic acid [D]-rich C-terminal domain) family of transcriptional coregulators and was originally cloned from a differential display screen between pigmented mouse B16 melanoma cells and their dedifferentiated weakly-pigmented derivative, How to cite this article Howlin et al (2015), Loss of CITED1, an MITF regulator, drives a phenotype switch in vitro and can predict clinical outcome in primary melanoma tumours
By 2005, as gene expression profiling became relatively commonplace, CITED1 was identified in several new screens of tumours and cell lines: two studies identified CITED1 as a gene whose expression distinguished nevi from primary melanoma, another found CITED1 to be upregulated in advanced stage melanomas in comparison to benign nevi or melanoma in situ, while expression profiling of an in vitro progression model identified CITED1 among a signature of genes lost in aggressive melanoma lines relative to primary melanocytes in culture (Ryu et al, 2007; Haqq et al, 2005; Talantov, 2005; Smith, Hoek & Becker, 2005)
The original signature set defined by Hoek et al, was redefined as more public datasets became available and has a slightly different but overlapping gene profile based on the top ranked differentially expressed genes (Fig. 1B). Both MITF and CITED1 are in the proliferative cohort and their response to TGFβ treatment was confirmed at protein level in A2058 cells (Fig. 1C)
Summary
CITED1 is the founding member of the CITED (CBP/p300-interacting transactivator with glutamic acid [E]/aspartic acid [D]-rich C-terminal domain) family of transcriptional coregulators and was originally cloned from a differential display screen between pigmented mouse B16 melanoma cells and their dedifferentiated weakly-pigmented derivative, How to cite this article Howlin et al (2015), Loss of CITED1, an MITF regulator, drives a phenotype switch in vitro and can predict clinical outcome in primary melanoma tumours. This led to speculation that CITED1 or msg (melanocyte specific gene 1) as it was known at that time, was involved in the process of pigmentation (Shioda, Fenner & Isselbacher, 1996). By 2005, as gene expression profiling became relatively commonplace, CITED1 was identified in several new screens of tumours and cell lines: two studies identified CITED1 as a gene whose expression distinguished nevi from primary melanoma, another found CITED1 to be upregulated in advanced stage melanomas in comparison to benign nevi or melanoma in situ, while expression profiling of an in vitro progression model identified CITED1 among a signature of genes lost in aggressive melanoma lines relative to primary melanocytes in culture (Ryu et al, 2007; Haqq et al, 2005; Talantov, 2005; Smith, Hoek & Becker, 2005)
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