Identification of different isoforms of 14-3-3 protein family in human dermal and epidermal layers

Abstract

We have previously demonstrated a high level of stratifin, also known as 14-3-3 sigma in differentiated keratinocyte cell lysate and conditioned medium (CM). In this study, we asked the question of whether other 14-3-3 isoforms are expressed in human dermal fibroblasts, keratinocytes, intact dermal and epidermal layers of skin. In order to address this question, total proteins extracted from cultured cells or skin layers were subjected to western blot analysis using seven different primary antibodies specific to well-known mammalian isoforms, beta, gamma, epsilon, eta, sigma, tau, and zeta of 14-3-3 protein family. The autoradiograms corresponding to each isoform were then quantified and compared. The results revealed the presence of very high levels of all seven isoforms in cultured keratinocyte and conditioned medium. With the exception of tau isoform, other 14-3-3 isoforms were also present in intact epidermal layer of normal skin. The profile of 14-3-3 proteins in whole skin was similar to that of epidermis. In contrast, only gamma 14-3-3 isoform, was present in dermal layer obtained from the same skin sample. On the other hand, cultured fibroblasts express a high level of beta, epsilon, gamma and eta and a low level of zeta and tau, but not sigma isoform. However, the levels of 14-3-3 epsilon, gamma and eta were barely detectable in fibroblast conditioned medium. Further, we also used immunohistochemical staining to identify the 14-3-3 isoform expressing cells in human skin sections. The finding revealed different expression profile for each of these isoforms mainly in differentiated keratinocytes located within the layer of lucidum. However, fibroblasts located within the dermal layer did not show any detectable levels of these proteins. In conclusion, all members of 14-3-3 proteins are expressed by cells of epidermal but not dermal layer of skins and that these proteins are mainly expressed by differentiated keratinocytes.

In vitro study of the impact of mechanical tension on the dermal fibroblast phenotype in the context of skin wound healing

“…Indeed, keratinocytes are known to release stratifin, also called MMP-14, which can stimulate the expression of MMP-1 and matrix breakdown by fibroblasts (Kilani et al, 2008(Kilani et al, , 2013Lam et al, 2004).…”

Systematic evaluations of skin damage irradiated by an erbium:YAG laser: Histopathologic analysis, proteomic profiles, and cellular response

“…They are a ubiquitous family of acidic eukaryotic proteins [32]. Kilani et al [33] suggested that all 14-3-3 proteins are expressed by cells of the epidermal but not the dermal layer, and these proteins are mainly expressed by differentiated keratinocytes. The Western blot results showed that the level of total 14-3-3 expression was higher in skin treated with the laser relative to that without treatment.…”

14-3-3 Proteins are Regulators of Autophagy

“…A large number of 14-3-3 binding motifs have been established [5], and these have recently been reviewed [6]. Seven mammalian 14-3-3 isoforms have been reported (α/β, ε, γ, σ, ζ/δ, τ/θ and η) that are encoded by seven different genes and vary in their expression levels between tissues [7,8,9]. Moreover, highly redundant functions exist between the different 14-3-3 isoforms.…”