Lineage specific composition of cyclin D-CDK4/CDK6-p27 complexes reveals distinct functions of CDK4, CDK6 and individual D-type cyclins in differentiating cells of embryonic origin

Cell Prolif. 2008 Dec;41(6):875-893. doi: 10.1111/j.1365-2184.2008.00556.x.

Abstract

Objectives: This article is to study the role of G(1)/S regulators in differentiation of pluripotent embryonic cells.

Materials and methods: We established a P19 embryonal carcinoma cell-based experimental system, which profits from two similar differentiation protocols producing endodermal or neuroectodermal lineages. The levels, mutual interactions, activities, and localization of G(1)/S regulators were analysed with respect to growth and differentiation parameters of the cells.

Results and conclusions: We demonstrate that proliferation parameters of differentiating cells correlate with the activity and structure of cyclin A/E-CDK2 but not of cyclin D-CDK4/6-p27 complexes. In an exponentially growing P19 cell population, the cyclin D1-CDK4 complex is detected, which is replaced by cyclin D2/3-CDK4/6-p27 complex following density arrest. During endodermal differentiation kinase-inactive cyclin D2/D3-CDK4-p27 complexes are formed. Neural differentiation specifically induces cyclin D1 at the expense of cyclin D3 and results in predominant formation of cyclin D1/D2-CDK4-p27 complexes. Differentiation is accompanied by cytoplasmic accumulation of cyclin Ds and CDK4/6, which in neural cells are associated with neural outgrowths. Most phenomena found here can be reproduced in mouse embryonic stem cells. In summary, our data demonstrate (i) that individual cyclin D isoforms are utilized in cells lineage specifically, (ii) that fundamental difference in the function of CDK4 and CDK6 exists, and (iii) that cyclin D-CDK4/6 complexes function in the cytoplasm of differentiated cells. Our study unravels another level of complexity in G(1)/S transition-regulating machinery in early embryonic cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Differentiation*
  • Cell Line, Tumor
  • Cell Lineage*
  • Cell Proliferation
  • Cyclin A / metabolism
  • Cyclin D
  • Cyclin E / metabolism
  • Cyclin-Dependent Kinase 4 / metabolism*
  • Cyclin-Dependent Kinase 6 / metabolism*
  • Cyclin-Dependent Kinase Inhibitor p27 / metabolism*
  • Cyclins / metabolism*
  • Embryo, Mammalian / cytology*
  • Embryo, Mammalian / metabolism
  • Embryonic Stem Cells / metabolism
  • G1 Phase
  • Humans
  • Intracellular Space / metabolism
  • Mice
  • Models, Biological
  • Protein Binding
  • Protein Transport
  • S Phase

Substances

  • Cyclin A
  • Cyclin D
  • Cyclin E
  • Cyclins
  • Cyclin-Dependent Kinase Inhibitor p27
  • CDK4 protein, human
  • Cyclin-Dependent Kinase 4
  • Cyclin-Dependent Kinase 6