Abstract
Connexins (Cxs) are believed to play a significant role in the control of cellular events such as cell proliferation and differentiation. The overall aim of the present work was to determine if modification of Cx expression and/or function altered the commitment of cells to either enter the differentiation pathway or stay in a proliferative state. Two Cx modification agents, GAP27 and AAP10, and a novel murine epidermal stem cell model were employed as investigative tools in this study. The development of the novel mouse model was examined under control condition and following modification of Cxs to determine the role of Cxs in controlling cell proliferation and differentiation events.The Cx specificity and potential mechanism of action of the modification agents, GAP27 and AAP10, were characterised using HeLa cells transfected and selected to express individual Cx subtypes. GAP27 was found to selectively down-regulate Cx43-mediated gap junction intercellular communication (GJIC). AAP10 selectively up-regulated Cx43 and Cx40 expression and function via a PKC-dependent pathway. These results suggested that these versatile tools could be valuable agents to investigate the impact Cx modification has on cell fate.
An epidermal stem cell model was characterised by morphology and real-time PCR and shown to be composed of three distinct cell types - ‘stem-like’, early and late amplifying cells. The epidermal stem cell model was shown to express Cx43 in a differential manner. Cx43 expression was not uniform throughout the epidermal model with discrete compartments of Cx43 expression observed. Discrete communication compartments were identified within the epidermal model. Modification of Cx43 by AAP10 prevented cell maturation from early to late amplifying cells, although no effect was observed in the maturation of ‘stem-like’ to early amplifying cells. No effect in colony growth and maturation was observed following GAP27 exposure.
Exposure to AAP10 resulted in enhanced Cx43 expression and also in enhanced E- cadherin expression. This suggests a further mechanism for AAP10 in the stability of the junctional nexus. Such an action could occur through alterations to P-catenin and possibly the Wnt signalling pathway.
Putative epidermal stem cell colonies were isolated from D66E1 mice to determine the effect mutation of Cx26 had on cell proliferation and differentiation. D66H epidermal colonies were observed to have a faster growth and proliferation rate in comparison to wt colonies.
We conclude that data presented in this study suggests that Cxs do have a role to play in the control of cellular events such as cell proliferation and differentiation. The data presented here indicates that if you enhance Cx43 protein expression by AAP10, this results in a reduction in cell proliferation. Alternatively a change in Cx26 protein expression by genetic mutation increases cell proliferation.
| Date of Award | 2009 |
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| Original language | English |
| Awarding Institution |
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| Supervisor | Patricia Martin (Supervisor) |