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dc.contributor.authorDodd, M. Ernest
dc.date.accessioned2015-02-20T16:16:30Z
dc.date.available2015-02-20T16:16:30Z
dc.date.issued2004-08en
dc.identifier.urihttp://hdl.handle.net/10675.2/344626
dc.descriptionThe file you are attempting to access is currently restricted to Augusta University. Please log in with your NetID if off campus.
dc.description.abstractThe epidermis is important for the body's maintenance of water homeostasis and resistance to environmental stress, and the m ajor cell type of the epidermis is the keratinocyte. Keratinocyte maturation requires proliferation, followed by terminal differentiation, and diseases of the skin often exhibit deregulated epidermal maturation. Protein kinase D (PKD) expression correlates with proliferation in keratinocytes, and PKD activation occurs in response to mitogen stimulation in other cell types. W e have hypothesized that PKD functions as a pro-proliferative and/or anti-differentiative signal in primary mouse keratinocytes and have predicted that agents that stimulate differentiation might also initiate a reduction in PKD expression and/or activation to allow differentiation to proceed. Thus, changes in PKD levels, autophosphorylation and activity were analyzed upon treatment with differentiating agents and with 1 2 -0 - tetradecanoylphorbol-13-acetate, TPA, which stimulates differentiation acutely and proliferation chronically. 1,25-dihydroxyvitamin D3 -, elevated extracellular calcium-, and acute TPA-induced differentiation down-modulated PKD levels and autophosphorylation at serine 916. In addition, elevated extracellular calcium- and acute TPA-induced differentiation down-modulated PKD activity. Chronic TPA treatment stimulated proliferation and caused a recovery o f PKD levels, autophosphorylation and activity. In co-transfection experiments in keratinocytes, co-expression of PKD increased and decreased the promoter activities of keratin 5, a marker of proliferation, and involucrin, a marker of differentiation, respectively, and opposed the effects of elevated extracellular calcium on the expression of these markers. W hile cloning PKD for expression studies, we identified a splice variant of PKD, PKD{3, which is differentially spliced in a region important in activation and subcellular localization. Therefore, we hypothesized that this splice variant may have dissimilar activation properties and/or alternate roles in keratinocyte maturation. However, in vitro activation studies demonstrated equal activation of PK D a (full length) and PKDj3 by TPA and DAG. Co-transfection experiments showed that P K D a and PKDp affected marker expression to the same degree and similarly opposed the effects of elevated extracellular calcium-induced differentiation on marker expression. Our work represents the first demonstration of: 1) down-modulation o f PKD during differentiation, 2) pro-proliferative/anti-differentiative effects of PKD on keratinocyte marker expression and 3) existence of a splice variant of PKD.
dc.relation.urlhttp://search.proquest.com/docview/305097688?accountid=12365en
dc.rightsCopyright protected. Unauthorized reproduction or use beyond the exceptions granted by the Fair Use clause of U.S. Copyright law may violate federal law.en
dc.subjectPKDen
dc.subjectKeratinocyteen
dc.subjectCalciumen
dc.subjectTPAen
dc.subject1,25-dihydroxyvitamin D3en
dc.titleProtein Kinase D In Keratinocyte Maturationen
dc.typeDissertationen
dc.contributor.departmentDepartment of Physiologyen
dc.description.advisorBollag, Wendy B.en
dc.description.committeeChew, Catherine; Johnson, John; Mivechi, Nahid; Hsu, Stephenen
dc.description.degreeDoctor of Philosophy (Ph.D.)en
refterms.dateFOA2021-01-15T20:24:51Z
html.description.abstractThe epidermis is important for the body's maintenance of water homeostasis and resistance to environmental stress, and the m ajor cell type of the epidermis is the keratinocyte. Keratinocyte maturation requires proliferation, followed by terminal differentiation, and diseases of the skin often exhibit deregulated epidermal maturation. Protein kinase D (PKD) expression correlates with proliferation in keratinocytes, and PKD activation occurs in response to mitogen stimulation in other cell types. W e have hypothesized that PKD functions as a pro-proliferative and/or anti-differentiative signal in primary mouse keratinocytes and have predicted that agents that stimulate differentiation might also initiate a reduction in PKD expression and/or activation to allow differentiation to proceed. Thus, changes in PKD levels, autophosphorylation and activity were analyzed upon treatment with differentiating agents and with 1 2 -0 - tetradecanoylphorbol-13-acetate, TPA, which stimulates differentiation acutely and proliferation chronically. 1,25-dihydroxyvitamin D3 -, elevated extracellular calcium-, and acute TPA-induced differentiation down-modulated PKD levels and autophosphorylation at serine 916. In addition, elevated extracellular calcium- and acute TPA-induced differentiation down-modulated PKD activity. Chronic TPA treatment stimulated proliferation and caused a recovery o f PKD levels, autophosphorylation and activity. In co-transfection experiments in keratinocytes, co-expression of PKD increased and decreased the promoter activities of keratin 5, a marker of proliferation, and involucrin, a marker of differentiation, respectively, and opposed the effects of elevated extracellular calcium on the expression of these markers. W hile cloning PKD for expression studies, we identified a splice variant of PKD, PKD{3, which is differentially spliced in a region important in activation and subcellular localization. Therefore, we hypothesized that this splice variant may have dissimilar activation properties and/or alternate roles in keratinocyte maturation. However, in vitro activation studies demonstrated equal activation of PK D a (full length) and PKDj3 by TPA and DAG. Co-transfection experiments showed that P K D a and PKDp affected marker expression to the same degree and similarly opposed the effects of elevated extracellular calcium-induced differentiation on marker expression. Our work represents the first demonstration of: 1) down-modulation o f PKD during differentiation, 2) pro-proliferative/anti-differentiative effects of PKD on keratinocyte marker expression and 3) existence of a splice variant of PKD.


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