Protein kinase D1 deficiency promotes differentiation in epidermal keratinocytes

Vivek Choudhary, Lawrence O. Olala, Ismail Kaddour-Djebbar, Inas Helwa, Wendy B Bollag

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

•Effect of protein kinase D1 (PKD1) deficiency was examined in keratinocytes in vitro.•PKD1 was deleted by infection of floxed keratinocytes with Cre-recombinase adenovirus.•PKD1 gene deletion decreased PKD1 mRNA and protein levels in keratinocytes.•PKD1 deletion stimulated keratinocyte differentiation marker expression.•PKD1 deletion inhibited keratinocyte proliferation. Background: Protein kinase D (PKD or PKD1) is a serine/threonine protein kinase that has been shown to play a role in a variety of cellular processes; however, the function of PKD1 in the skin has not been fully investigated. The balance between proliferation and differentiation processes in the predominant cells of the epidermis, the keratinocytes, is essential for normal skin function. Objective: To investigate the effect of PKD1 deficiency on proliferation and differentiation of epidermal keratinocytes. Methods: We utilized a floxed PKD1 mouse model such that infecting epidermal keratinocytes derived from these mice with an adenovirus expressing Cre-recombinase allowed us to determine the effect of PKD1 gene loss in vitro. Proliferation and differentiation were monitored using qRT-PCR, Western blot, transglutaminase activity assays, [3H]thymidine incorporation into DNA and cell cycle analysis. Results: A significant decrease in PKD1 mRNA and protein levels was achieved in adenoviral Cre-recombinase-infected cells. Deficiency of PKD1 resulted in significant increases in the mRNA and protein expression of various differentiation markers such as loricrin, involucrin, and keratin 10 either basally and/or upon stimulation of differentiation. PKD1-deficient keratinocytes also showed an increase in transglutaminase expression and activity, indicating an anti-differentiative role of PKD1. Furthermore, the PKD1-deficient keratinocytes exhibited decreased proliferation. However, PKD1 loss had no effect on stem cell marker expression. Conclusions: Cre-recombinase-mediated knockdown represents an additional approach demonstrating that PKD1 is an anti-differentiative, pro-proliferative signal in mouse keratinocytes.

Original languageEnglish (US)
Pages (from-to)186-195
Number of pages10
JournalJournal of Dermatological Science
Volume76
Issue number3
DOIs
StatePublished - Dec 1 2014

Fingerprint

Keratinocytes
Protein Kinases
Transglutaminases
Differentiation Antigens
Adenoviridae
Messenger RNA
Skin
Keratin-10
Genes
Proteins
Protein-Serine-Threonine Kinases
Gene Deletion
Stem cells
Epidermis
Thymidine
Assays

Keywords

  • Differentiation
  • Keratinocyte
  • Proliferation
  • Protein kinase D
  • Transglutaminase

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Dermatology

Cite this

Protein kinase D1 deficiency promotes differentiation in epidermal keratinocytes. / Choudhary, Vivek; Olala, Lawrence O.; Kaddour-Djebbar, Ismail; Helwa, Inas; Bollag, Wendy B.

In: Journal of Dermatological Science, Vol. 76, No. 3, 01.12.2014, p. 186-195.

Research output: Contribution to journalArticle

Choudhary, Vivek ; Olala, Lawrence O. ; Kaddour-Djebbar, Ismail ; Helwa, Inas ; Bollag, Wendy B. / Protein kinase D1 deficiency promotes differentiation in epidermal keratinocytes. In: Journal of Dermatological Science. 2014 ; Vol. 76, No. 3. pp. 186-195.
@article{ae817566275a49e1955fd117b431cc05,
title = "Protein kinase D1 deficiency promotes differentiation in epidermal keratinocytes",
abstract = "•Effect of protein kinase D1 (PKD1) deficiency was examined in keratinocytes in vitro.•PKD1 was deleted by infection of floxed keratinocytes with Cre-recombinase adenovirus.•PKD1 gene deletion decreased PKD1 mRNA and protein levels in keratinocytes.•PKD1 deletion stimulated keratinocyte differentiation marker expression.•PKD1 deletion inhibited keratinocyte proliferation. Background: Protein kinase D (PKD or PKD1) is a serine/threonine protein kinase that has been shown to play a role in a variety of cellular processes; however, the function of PKD1 in the skin has not been fully investigated. The balance between proliferation and differentiation processes in the predominant cells of the epidermis, the keratinocytes, is essential for normal skin function. Objective: To investigate the effect of PKD1 deficiency on proliferation and differentiation of epidermal keratinocytes. Methods: We utilized a floxed PKD1 mouse model such that infecting epidermal keratinocytes derived from these mice with an adenovirus expressing Cre-recombinase allowed us to determine the effect of PKD1 gene loss in vitro. Proliferation and differentiation were monitored using qRT-PCR, Western blot, transglutaminase activity assays, [3H]thymidine incorporation into DNA and cell cycle analysis. Results: A significant decrease in PKD1 mRNA and protein levels was achieved in adenoviral Cre-recombinase-infected cells. Deficiency of PKD1 resulted in significant increases in the mRNA and protein expression of various differentiation markers such as loricrin, involucrin, and keratin 10 either basally and/or upon stimulation of differentiation. PKD1-deficient keratinocytes also showed an increase in transglutaminase expression and activity, indicating an anti-differentiative role of PKD1. Furthermore, the PKD1-deficient keratinocytes exhibited decreased proliferation. However, PKD1 loss had no effect on stem cell marker expression. Conclusions: Cre-recombinase-mediated knockdown represents an additional approach demonstrating that PKD1 is an anti-differentiative, pro-proliferative signal in mouse keratinocytes.",
keywords = "Differentiation, Keratinocyte, Proliferation, Protein kinase D, Transglutaminase",
author = "Vivek Choudhary and Olala, {Lawrence O.} and Ismail Kaddour-Djebbar and Inas Helwa and Bollag, {Wendy B}",
year = "2014",
month = "12",
day = "1",
doi = "10.1016/j.jdermsci.2014.09.007",
language = "English (US)",
volume = "76",
pages = "186--195",
journal = "Journal of Dermatological Science",
issn = "0923-1811",
publisher = "Elsevier Ireland Ltd",
number = "3",

}

TY - JOUR

T1 - Protein kinase D1 deficiency promotes differentiation in epidermal keratinocytes

AU - Choudhary, Vivek

AU - Olala, Lawrence O.

AU - Kaddour-Djebbar, Ismail

AU - Helwa, Inas

AU - Bollag, Wendy B

PY - 2014/12/1

Y1 - 2014/12/1

N2 - •Effect of protein kinase D1 (PKD1) deficiency was examined in keratinocytes in vitro.•PKD1 was deleted by infection of floxed keratinocytes with Cre-recombinase adenovirus.•PKD1 gene deletion decreased PKD1 mRNA and protein levels in keratinocytes.•PKD1 deletion stimulated keratinocyte differentiation marker expression.•PKD1 deletion inhibited keratinocyte proliferation. Background: Protein kinase D (PKD or PKD1) is a serine/threonine protein kinase that has been shown to play a role in a variety of cellular processes; however, the function of PKD1 in the skin has not been fully investigated. The balance between proliferation and differentiation processes in the predominant cells of the epidermis, the keratinocytes, is essential for normal skin function. Objective: To investigate the effect of PKD1 deficiency on proliferation and differentiation of epidermal keratinocytes. Methods: We utilized a floxed PKD1 mouse model such that infecting epidermal keratinocytes derived from these mice with an adenovirus expressing Cre-recombinase allowed us to determine the effect of PKD1 gene loss in vitro. Proliferation and differentiation were monitored using qRT-PCR, Western blot, transglutaminase activity assays, [3H]thymidine incorporation into DNA and cell cycle analysis. Results: A significant decrease in PKD1 mRNA and protein levels was achieved in adenoviral Cre-recombinase-infected cells. Deficiency of PKD1 resulted in significant increases in the mRNA and protein expression of various differentiation markers such as loricrin, involucrin, and keratin 10 either basally and/or upon stimulation of differentiation. PKD1-deficient keratinocytes also showed an increase in transglutaminase expression and activity, indicating an anti-differentiative role of PKD1. Furthermore, the PKD1-deficient keratinocytes exhibited decreased proliferation. However, PKD1 loss had no effect on stem cell marker expression. Conclusions: Cre-recombinase-mediated knockdown represents an additional approach demonstrating that PKD1 is an anti-differentiative, pro-proliferative signal in mouse keratinocytes.

AB - •Effect of protein kinase D1 (PKD1) deficiency was examined in keratinocytes in vitro.•PKD1 was deleted by infection of floxed keratinocytes with Cre-recombinase adenovirus.•PKD1 gene deletion decreased PKD1 mRNA and protein levels in keratinocytes.•PKD1 deletion stimulated keratinocyte differentiation marker expression.•PKD1 deletion inhibited keratinocyte proliferation. Background: Protein kinase D (PKD or PKD1) is a serine/threonine protein kinase that has been shown to play a role in a variety of cellular processes; however, the function of PKD1 in the skin has not been fully investigated. The balance between proliferation and differentiation processes in the predominant cells of the epidermis, the keratinocytes, is essential for normal skin function. Objective: To investigate the effect of PKD1 deficiency on proliferation and differentiation of epidermal keratinocytes. Methods: We utilized a floxed PKD1 mouse model such that infecting epidermal keratinocytes derived from these mice with an adenovirus expressing Cre-recombinase allowed us to determine the effect of PKD1 gene loss in vitro. Proliferation and differentiation were monitored using qRT-PCR, Western blot, transglutaminase activity assays, [3H]thymidine incorporation into DNA and cell cycle analysis. Results: A significant decrease in PKD1 mRNA and protein levels was achieved in adenoviral Cre-recombinase-infected cells. Deficiency of PKD1 resulted in significant increases in the mRNA and protein expression of various differentiation markers such as loricrin, involucrin, and keratin 10 either basally and/or upon stimulation of differentiation. PKD1-deficient keratinocytes also showed an increase in transglutaminase expression and activity, indicating an anti-differentiative role of PKD1. Furthermore, the PKD1-deficient keratinocytes exhibited decreased proliferation. However, PKD1 loss had no effect on stem cell marker expression. Conclusions: Cre-recombinase-mediated knockdown represents an additional approach demonstrating that PKD1 is an anti-differentiative, pro-proliferative signal in mouse keratinocytes.

KW - Differentiation

KW - Keratinocyte

KW - Proliferation

KW - Protein kinase D

KW - Transglutaminase

UR - http://www.scopus.com/inward/record.url?scp=84919467811&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84919467811&partnerID=8YFLogxK

U2 - 10.1016/j.jdermsci.2014.09.007

DO - 10.1016/j.jdermsci.2014.09.007

M3 - Article

VL - 76

SP - 186

EP - 195

JO - Journal of Dermatological Science

JF - Journal of Dermatological Science

SN - 0923-1811

IS - 3

ER -