Differentiation-dependent gene expression is a hallmark of biological systems/cell types that undergo differentiation. Thus, understanding normal and abnormal keratinocyte differentiation, requires an understanding of the mechanisms that regulate gene expression. This includes identifying ligands that initiate the regulation, understanding events that transfer the regulation from the cell surface to the nucleus, and describe nuclear mechanisms that regulate target genes. We have developed the human involucrin (hINV) gene as an advanced model for the study of differentiation-dependent gene expression. As part of this effort, we have identified transcription factors (junB, junD, Fra-1) that regulate hINV gene expression by binding to specific DNA binding sites within the hINV gene upstream regulatory region. Knowledge of these nuclear events has made it possible to begin examining the signal transduction cascade that transfers the gene regulatory signal from the cell surface to these transcription factors in the nucleus (Effimova et al., J. Biol. Chem. 273:24387, 1998). We initiated this study because in spite of the importance of these signaling cascades in other systems, little is known regarding the steps that transfer regulatory information to the nucleus for keratinocyte genes that are regulated during differentiation. This published study initiates a comprehensive effort to understand signal transduction for a differentiation-regulated gene, and provides the rationale for the experiments outlined in this proposal. Our studies implicate a mitogen-activated protein kinase (MAPK) signaling cascade that involves protein kinase-c, Ras, MEKK1, MEK3/MEK6, p38, and AP1 family members (junB, junD, Fra-1). This cascade is clearly important, as inhibition of these kinases turn off hINV gene expression. These studies are also innovative, as the p38 MAPKs have not been thought to function as regulators of gene expression during differentiation. However, much needs to be learned, and in the studies outlined in this proposal, we will focus on key points in this cascade for which we have limited knowledge. We will (i) identify the PKC isoforms involved in this regulation, (ii) study the role of MEK3 and MEK6 as regulators of p38 activity and determine which p38 isoforms are involved in this regulation, (iii) perform studies designed to determine how p38 regulates AP1 transcription factor distribution, level and activation state, and (iv) evaluate the role of these kinases, in vivo, using DNA ballistics (gene gun) delivery to mouse epidermis. These studies are designed to provide new insights regarding regulation of differentiation-dependent gene expression in epidermis. As involucrin is one of many genes that are regulated during differentiation, we hope and expect that understanding provided by these studies will be useful to other investigators.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR046494-03
Application #
6512028
Study Section
General Medicine A Subcommittee 2 (GMA)
Program Officer
Moshell, Alan N
Project Start
2000-03-01
Project End
2005-02-28
Budget Start
2002-03-01
Budget End
2003-02-28
Support Year
3
Fiscal Year
2002
Total Cost
$181,452
Indirect Cost
Name
Case Western Reserve University
Department
Physiology
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Rorke, Ellen A; Adhikary, Gautam; Young, Christina A et al. (2015) Suppressing AP1 factor signaling in the suprabasal epidermis produces a keratoderma phenotype. J Invest Dermatol 135:170-180
Saha, Kamalika; Adhikary, Gautam; Kanade, Santosh R et al. (2014) p38? regulates p53 to control p21Cip1 expression in human epidermal keratinocytes. J Biol Chem 289:11443-53
Chew, Yap Ching; Adhikary, Gautam; Xu, Wen et al. (2013) Protein kinase C ? increases Kruppel-like factor 4 protein, which drives involucrin gene transcription in differentiating keratinocytes. J Biol Chem 288:17759-68
Han, Bingshe; Rorke, Ellen A; Adhikary, Gautam et al. (2012) Suppression of AP1 transcription factor function in keratinocyte suppresses differentiation. PLoS One 7:e36941
Chew, Yap Ching; Adhikary, Gautam; Wilson, Gerald M et al. (2012) Sulforaphane induction of p21(Cip1) cyclin-dependent kinase inhibitor expression requires p53 and Sp1 transcription factors and is p53-dependent. J Biol Chem 287:16168-78
Kanade, Santosh R; Eckert, Richard L (2012) Protein arginine methyltransferase 5 (PRMT5) signaling suppresses protein kinase C?- and p38?-dependent signaling and keratinocyte differentiation. J Biol Chem 287:7313-23
Chen, Cheng-Jueng; Wu, Bai-Yao; Tsao, Pai-In et al. (2011) Increased matriptase zymogen activation in inflammatory skin disorders. Am J Physiol Cell Physiol 300:C406-15
Chew, Yap Ching; Adhikary, Gautam; Wilson, Gerald M et al. (2011) Protein kinase C (PKC) delta suppresses keratinocyte proliferation by increasing p21(Cip1) level by a KLF4 transcription factor-dependent mechanism. J Biol Chem 286:28772-82
Adhikary, Gautam; Chew, Yap Ching; Reece, E Albert et al. (2010) PKC-delta and -eta, MEKK-1, MEK-6, MEK-3, and p38-delta are essential mediators of the response of normal human epidermal keratinocytes to differentiating agents. J Invest Dermatol 130:2017-30
Rorke, E A; Adhikary, G; Jans, R et al. (2010) AP1 factor inactivation in the suprabasal epidermis causes increased epidermal hyperproliferation and hyperkeratosis but reduced carcinogen-dependent tumor formation. Oncogene 29:5873-82

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