Notch Cooperates with Survivin to Maintain Stemness and to Stimulate Proliferation in Human Keratinocytes during Ageing

Abstract

The Notch signaling pathway orchestrates cell fate by either inducing cell differentiation or maintaining cells in an undifferentiated state. This study aims to evaluate Notch expression and function in normal human keratinocytes. Notch1 is expressed in all epidermal layers, though to a different degree of intensity, with a dramatic decrease during ageing. Notch1 intracellular domain (N1ICD) levels are decreased during transit from keratinocyte stem cells (KSC) to transit amplifying (TA) cells, mimicking survivin expression in samples from donors of all ages. Calcium markedly reduces N1ICD levels in keratinocytes. N1ICD overexpression induces the up-regulation of survivin and the down-regulation of keratin 10 and involucrin, while increasing the S phase of the cell cycle. On the other hand, Notch1 inhibition (DAPT) dose-dependently decreases survivin, stimulates differentiation, and reduces keratinocyte proliferation in samples from donors of all ages. Silencing Notch downgrades survivin and increases keratin 10. In addition, Notch1 inhibition decreases survivin levels and proliferation both in KSC and TA cells. Finally, while survivin overexpression decreases keratinocyte differentiation and increases N1ICD expression both in KSC and TA cells, silencing survivin results in N1ICD down-regulation and an increase in differentiation markers. These results suggest that the Notch1/survivin crosstalk contributes to the maintenance of stemness in human keratinocytes.

A novel DLX3–PKC integrated signaling network drives keratinocyte differentiation

“…ChIP-seq analysis of DLX3 and H3K4me3 DNA binding domains identified DLX3-bound promoter regions of genes involved in cell cycle regulation during differentiation, such as Ccnd1, Ccne1; 41, 42 and epidermal structure, such as Clic4, Dsp, Evpl, Fabp5, 43, 44, 45, 46 and transcription factors such as Grhl3, Klf4, Msx2 and Notch1 47, 48, 49, 50 (Figure 8). Together with the transcriptomic analysis, the ChIP-seq analysis of DLX3 and H4K4m3 binding supports a potential DLX3-dependent transcriptional regulation of these genes during epidermal differentiation.…”

Insights into the Regulation of Survivin Expression in Tumors

“…Additionally, it has been shown that SOX2 regulates self-renewal and tumorigenicity of human melanoma-initiating cells [46]. Of note, Notch1 cooperates with survivin to maintain stemness and stimulates proliferation of human keratinocytes [47]. Taken together, survivin, in a large part through the Hedgehog/GLI pathway, substantially contribute to the pro-oncogenic activities including stemness in human cancer.…”

Survivin Modulates Squamous Cell Carcinoma-Derived Stem-Like Cell Proliferation, Viability and Tumor Formation in Vivo

“…While scramble transfected cells grow exponentially with time, survivin depleted cells display a slow rate of proliferation, as shown by vital cell analysis ( Figure 1 A), and give rise to a significantly lower amount of colonies with bigger size ( Figure 1 B,C). In association with the short and long term proliferative potential, survivin expression knocking-down in cSCC primary cells correlates with the decreased expression of stemness-associated markers, such as OCT-4, NOTCH1, CD133 and β 1 -integrin [ 22 , 28 , 29 , 30 , 31 ] ( Figure 1 D). Interestingly, NOTCH1-survivin axis in the maintenance of human keratinocyte stemness [ 30 ] appears to be preserved also in the context of tumor cells.…”

“…In association with the short and long term proliferative potential, survivin expression knocking-down in cSCC primary cells correlates with the decreased expression of stemness-associated markers, such as OCT-4, NOTCH1, CD133 and β 1 -integrin [ 22 , 28 , 29 , 30 , 31 ] ( Figure 1 D). Interestingly, NOTCH1-survivin axis in the maintenance of human keratinocyte stemness [ 30 ] appears to be preserved also in the context of tumor cells. On the other hand, the expression of early epidermal differentiation markers, such as keratin 10 (K10) and involucrin, was increased ( Figure 1 D).…”