Regulation of gene expression by the CD44-Intracytoplasmic domain (CD44-ICD)
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Abstract
CD44 is a major cell membrane hyaluronan receptor and cell adhesion glycoprotein. It is involved in multiple cellular processes including cell migration and invasion, apoptosis, stemness and cell proliferation. It exists in a variety of isoforms mainly due to the alternative splicing of ten of its 20 exons and posttranslational modifications. CD44 also undergoes proteolytic cleavages that generate the CD44 intracytoplasmic domain (CD44-ICD). The CD44- ICD is translocated to the nucleus where it can regulate gene expression. The CD44-ICD- dependent regulation of gene expression is likely to be mediated by protein-protein interactions (PPI) with factors of the transcriptional machinery, which makes of the CD44-ICD a transcriptional co-regulator. However, the existence of the CD44-ICD response element (CIRE) and its reported binding to it makes of the CD44-ICD a transcription factor. We hypothesize that the CD44-ICD is a novel transcriptional regulator with properties of a co-regulator and a transcription factor. To test this hypothesis we carried out PPI assays such as proximity ligation assays (PLA) and plate array PPI assays. We also analyzed the expression of genes related with CD44-associated cellular events such as apoptosis, oxidative stress and stemness using proteome profiler assays and transcription factor activation plate arrays. PLA data showed that the CD44- ICD interacts with the transcription factor Runx2 in the nucleus as well as in the cytoplasm. We also found that the CD44-ICD interacts with more transcription factors. The expression of CD44 in MCF-7 cells, which is expected to generate the CD44-ICD, inhibited the expression of Nrf2, caspase 3 and E-cadherin, important genes in oxidative stress, apoptosis and stemness, respectively. The inhibition of Nrf2 in MCF-7/CD44 cells, promoted a higher sensitivity to oxidative stress caused by hydrogen peroxide compared to MCF-7 cells. Altogether, these data
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supports that CD44 can act as a regulator of gene expression and that this regulation is in part via its intracytoplasmic domain once released into the cytoplasm and nucleus. The CD44-ICD appears to be able to act as a co-activator as well as a co-repressor. Because the CD44-ICD size of only 72 residues, the CIRE sequence to which it can bind is unusual. This type of protein- DNA interaction is usually associated with larger DNA-binding proteins such as bona fide transcription factors. The dual capacity of the CD44-ICD to interact with proteins of the transcriptional machinery as well as to interact with its own response element on promoter sequences, might place the CD44-ICD in a novel category of transcriptional regulators.