Funktion von cCMP und cCMP-spezifischen Signalwegen

Stefanie Wolfertstetter
2015
The cyclic purine nucleotides cAMP and cGMP are established "second messengers" and play an essential role for numerous of (patho)physiological processes. The existence of the cyclic pyrimidine nucleotide cCMP was postulated in the late 1970s, but could not be clearly demonstrated due to the lack of sensitivity of the methods. Furthermore the presence of specific cCMP interacting proteins, a cCMP cleaving PDE or a cCMP forming cyclase could not be confirmed. Years later it was possible to
more » ... s possible to detect cCMP in vitro and in vivo with improved mass spectrometry methods. Also, it was found that in bacteria sAC and sGC have a cytidyl cyclase activity and are able to generate cCMP. Recently, the specific cCMP degrading PDE7A1 was found and also numerous cCMP interacting proteins, as cAK, cGK, MRPs and HCN channels, have been identified. These findings suggest that cCMP is involved in (patho)physiological processes as a "second messenger". In the present work, the main focus was put on finding further evidence of the "second messenger" function of cCMP. It was confirmed that both purified cAK, as well as purified cGKI or cGKII can be activated by cCMP. To investigate the physiological relevance of cCMP endogenous cG kinases in tissue lysates were stimulated concentration-dependently with cCMP. Using cCMP agarose precipitation studies it was confirmed that cGKI, cGKII and cAK Iÿ/Iÿ/IIÿ reg can specifically interact with cCMP in various tissues. Furthermore MAPK was identified as a specific cCMP-binding protein. Using co-immunoprecipitation it was also shown that there is a protein-protein interaction between cGKIÿ or cGKII and p44/42 MAPK. A direct stimulation of the MAP kinase with a cCMP analogue results in an increased MAPK phosphorylation, which was almost completely inhibited when adding a cAK inhibitor and a PKC inhibitor. To investigate the role of cGK in this process, we worked with cGK-KO animals. Especially in cGKII KO tissue the MAPK phosphorylation was increased. Using cGKI KO tissue a reduced MAPK phosphorylation [...]
doi:10.5283/epub.32681 fatcat:ggvuzajoardt5ktix2ju7bfunq