Logy, P-cadherin is involved in homeostatic processes, such as cell differentiation, development and embryogenesis [32]. We have recently found that P-cadherin enriched cell populations show enhanced mammosphere forming efficiency (MFE), as well as increased expression of CD24, CD44 and CD49f, already described as normal or cancer stem cell markers. These results allowed to link P-cadherin expression to the luminal progenitor phenotype of the normal breast hierarchy and established an indirect effect of P-cadherin in stem cell biology [33]. Interestingly, these findings come along with observations that C/EBPb regulates stem cell activity and specifies luminal cell fate in the mammary gland, categorizing C/EBPb as one of the several critical transcription factors that specifies mammary stem cells fate during mammary gland development [34]. In a breast cancer biology setting, another interesting finding is related to the fact that P-cadherin, like C/EBPb, is not mutated in breast tumours, but its overexpression has been widely described in a subset of aggressive breast cancers [5]. Importantly, at a clinicopathological level, some C/EBPb isoforms, especially C/EBPb-LIP, correlates with an ER-negative breast cancer phenotype, highly proliferative and high grade lesions and poor patient outcome [8,35]. All these characteristics overlap with the ones observed in highly malignant breast tumours overexpressing P-cadherin. The present work demonstrates for the first time that Pcadherin and C/EBPb co-localize in the same breast cancer cells, and that there is a buy I-BET151 physical interaction between this transcription factor and CDH3 gene promoter. Herein, in addition to the identification of the promoter binding sites that are relevant for the ICG-001 site transcriptional modulation of CDH3 gene activity by C/EBPb, we still tested the relevance of the different C/EBPb isoforms along the CDH3 promoter. In fact, we show that C/EBPb-LIP is the only isoform capable to significantly induce P-cadherin protein expression, confirming in a way the results obtained in our previous study, where a significant activation of the promoter was only revealed for LIP, although LAP1 and LAP2 were also able to activate the promoter. However, in this study, we found that CDH3 gene is also significantly responsive to LAP1 and slightly to LAP2 isoform at the promoter level. These significant results were probably due to improved transfection efficiencies; however, although LAP1 and LAP2 are activating the gene promoter, supporting the classical knowledge described for these isoforms as transcriptional activators, this might not imply that these isoforms induce functional activity through protein synthesis. In fact, it has been largely discussed that the functionally transactivation potential of each C/ EBPb isoform can be highly modulated, since this ability strongly depends not only on dimer composition formed by C/EBPs, but specially on the partner proteins and responsive elements found in target gene promoters [5]. The fact that LIP activates CDH3 promoter, leading to protein synthesis, reinforces the emerging evidence that LIP acts as a transcriptional activator of gene expression, challenging the long-standing concept that LIPfashionably functions as a dominant-negative isoform [5]. We also observed that LAP2 was the C/EBPb isoform that activated CDH3 promoter in a less extent, which is apparently surprising in light that LAP2 isoform is considered to be the most transcriptionally ac.Logy, P-cadherin is involved in homeostatic processes, such as cell differentiation, development and embryogenesis [32]. We have recently found that P-cadherin enriched cell populations show enhanced mammosphere forming efficiency (MFE), as well as increased expression of CD24, CD44 and CD49f, already described as normal or cancer stem cell markers. These results allowed to link P-cadherin expression to the luminal progenitor phenotype of the normal breast hierarchy and established an indirect effect of P-cadherin in stem cell biology [33]. Interestingly, these findings come along with observations that C/EBPb regulates stem cell activity and specifies luminal cell fate in the mammary gland, categorizing C/EBPb as one of the several critical transcription factors that specifies mammary stem cells fate during mammary gland development [34]. In a breast cancer biology setting, another interesting finding is related to the fact that P-cadherin, like C/EBPb, is not mutated in breast tumours, but its overexpression has been widely described in a subset of aggressive breast cancers [5]. Importantly, at a clinicopathological level, some C/EBPb isoforms, especially C/EBPb-LIP, correlates with an ER-negative breast cancer phenotype, highly proliferative and high grade lesions and poor patient outcome [8,35]. All these characteristics overlap with the ones observed in highly malignant breast tumours overexpressing P-cadherin. The present work demonstrates for the first time that Pcadherin and C/EBPb co-localize in the same breast cancer cells, and that there is a physical interaction between this transcription factor and CDH3 gene promoter. Herein, in addition to the identification of the promoter binding sites that are relevant for the transcriptional modulation of CDH3 gene activity by C/EBPb, we still tested the relevance of the different C/EBPb isoforms along the CDH3 promoter. In fact, we show that C/EBPb-LIP is the only isoform capable to significantly induce P-cadherin protein expression, confirming in a way the results obtained in our previous study, where a significant activation of the promoter was only revealed for LIP, although LAP1 and LAP2 were also able to activate the promoter. However, in this study, we found that CDH3 gene is also significantly responsive to LAP1 and slightly to LAP2 isoform at the promoter level. These significant results were probably due to improved transfection efficiencies; however, although LAP1 and LAP2 are activating the gene promoter, supporting the classical knowledge described for these isoforms as transcriptional activators, this might not imply that these isoforms induce functional activity through protein synthesis. In fact, it has been largely discussed that the functionally transactivation potential of each C/ EBPb isoform can be highly modulated, since this ability strongly depends not only on dimer composition formed by C/EBPs, but specially on the partner proteins and responsive elements found in target gene promoters [5]. The fact that LIP activates CDH3 promoter, leading to protein synthesis, reinforces the emerging evidence that LIP acts as a transcriptional activator of gene expression, challenging the long-standing concept that LIPfashionably functions as a dominant-negative isoform [5]. We also observed that LAP2 was the C/EBPb isoform that activated CDH3 promoter in a less extent, which is apparently surprising in light that LAP2 isoform is considered to be the most transcriptionally ac.
