ABSTRACT
The relationship between the two coexpressed differentiation markers, profilaggrin and loricrin, is not clear right now. In this study, we explored the interaction of profilaggrin N-terminal domain (PND) with loricrin in keratinocytes and epidermis. Confocal immunofluorescence microscopic analysis of human epidermis showed that PND colocalized with loricrin. Loricrin nucleofected into HaCaT cells colocalized with PND in the nucleus and cytoplasm. The PND localizes to both the nucleus and cytoplasm of epidermal granular layer cells. Nucleofected PND also colocalized with keratin 10 (K10) in the nucleus and cytoplasm. Immunoelectron microscopic analysis of human epidermis confirmed the findings in nucleofected keratinocytes. Yeast two-hybrid assays showed that the B domain of human and mouse PND interacted with loricrin. The glutathione S-transferase (GST) pull-down analysis using recombinant GST-PND revealed that PND interacted with loricrin and K10. Knockdown of PND in an organotypic skin culture model caused loss of filaggrin expression and a reduction in both the size and number of keratohyalin granules, as well as markedly reduced expression of loricrin. Considering that expression of PND is closely linked to keratinocyte terminal differentiation, we conclude that PND interacts with loricrin and K10 in vivo and that these interactions are likely to be relevant for cornified envelope assembly and subsequent epidermal barrier formation.
Subject(s)
Epidermis/metabolism , Intermediate Filament Proteins/metabolism , Keratinocytes/metabolism , Membrane Proteins/metabolism , Protein Interaction Domains and Motifs/physiology , Animals , Cell Nucleus/metabolism , Cells, Cultured , Cytoplasm/metabolism , Epidermal Cells , Filaggrin Proteins , Gene Expression Regulation/drug effects , Humans , Intermediate Filament Proteins/genetics , Keratin-10/metabolism , Keratinocytes/cytology , Membrane Proteins/genetics , Mice , Mice, Knockout , Protein Interaction Domains and Motifs/genetics , RNA, Small Interfering/pharmacologyABSTRACT
Animal and clinical studies supporting the kindling hypothesis of alcohol withdrawal suggest the need to revisit current treatment concepts. While traditional approaches have emphasized symptom reduction and prevention of complications, novel approaches include slowing progress of clinical severity associated with multiple withdrawals. Currently, it is unclear if medications can halt cumulative neurotoxicity associated with multiple withdrawals. However, the ability of anticonvulsants to improve the course of alcohol withdrawal and their neuroprotective effects may be of interest. The use of anticonvulsants as probes in animal models of kindling and controlled trials examining the efficacy of newer anticonvulsants in the treatment of alcohol withdrawal may improve understanding of alcohol withdrawal kindling and its treatment.
