2002a; Häder et al. 2006) while forskolin activates the adenylyl cyclase (Schwer
et al. 2013) and increases the precision of gravitaxis in Euglena (Häder et al. 2006).
Phosphodiesterase quenches the cAMP after stimulation (Cai et al. 2015). Caffeine,
theophylline and IBMX are known inhibitors of this enzyme (Cameron and Baillie
2012; Steck et al. 2014), so that the cAMP signal stays high after application and the
gravitactic activity increases (Lebert et al. 1997; Häder and Lebert 2002; Streb et al.
2002). Application of 8-bromo-cAMP, incorporated into the cells, functions as a
cAMP analog, but is not quenched by the phosphodiesterase. As a result the
precision of gravitaxis increased (Lebert et al. 1997). cAMP seems to be a universal
component of sensory transduction chains since its involvement has been found in
the control of movement and development in the slime mold Dictyostelium (Renart
et al. 1981; Sultana et al. 2012) and in the gravitactic sensory transduction chain of
Paramecium (Bräucker et al. 2001; Hemmersbach and Braun 2006).
The next question is how the cAMP signal controls the flagellar activity which is
instrumental in the steering response of the cell. Favaro et al. (2012) had shown that
cAMP can activate a protein kinase A. Staurosporine, which is known to be an
inhibitor of protein kinases (Chang and Kaufman 2000) impairs gravitaxis in Euglena
(Häder et al. 2010). But it is interesting that prolonged exposure (225 min) to the drug
reversed the negative gravitaxis into a positive one as did strong visible and UV
radiation or salt stress (see above). This drug also impairs negative phototaxis which
could indicate that both sensory transduction chains share the same final step being the
activation of a phosphokinase A (Häder and Iseki 2017).
Using the same molecular biological tools involving degenerate primers as
described above showed that Euglena possesses at least five isoforms of protein
kinase A (PK.1–PK.5). The full range of the RNA sequence was revealed by RACEPCR. RNAi against the different isoforms showed that PK.1–PK.3 and PK.5 are
not involved in the gravitactic sensory transduction chain (Daiker et al. 2011). In
contrast, inhibition of PK.4 effectively blocked gravitaxis for several weeks. Three
weeks after the RNAi treatment a positive gravitaxis was observed. This finding
parallels the result by staurosporine inhibition. In addition, RNAi of PK.4 blocked
phototaxis which further confirms that gravitaxis and phototaxis share the same final
step, the activation of a protein kinase A by cAMP.
3.7 Conclusions and Open Questions
Gravitactic unicellular organisms are specialized for gravity sensing. They use this
information for their spatial orientation (gravitaxis). Summarizing the findings
allows us to construct a complete gravitaxis signal transduction chain, such as in
Euglena (Fig. 3.6). During forward locomotion the cell rotates around its horizontal
axis at about 1 Hz. When the flagellum points downwards the content of the cell
presses onto the lower membrane activating the TRP channels, thought to be located
at the front end adjacent to the flagellum, which allows Ca
2+ to enter the cell along
a previously established gradient by a Ca-ATPase. In fact, several rotations are
3.7 Conclusions and Open Questions
39
et al. 2013) and increases the precision of gravitaxis in Euglena (Häder et al. 2006).
Phosphodiesterase quenches the cAMP after stimulation (Cai et al. 2015). Caffeine,
theophylline and IBMX are known inhibitors of this enzyme (Cameron and Baillie
2012; Steck et al. 2014), so that the cAMP signal stays high after application and the
gravitactic activity increases (Lebert et al. 1997; Häder and Lebert 2002; Streb et al.
2002). Application of 8-bromo-cAMP, incorporated into the cells, functions as a
cAMP analog, but is not quenched by the phosphodiesterase. As a result the
precision of gravitaxis increased (Lebert et al. 1997). cAMP seems to be a universal
component of sensory transduction chains since its involvement has been found in
the control of movement and development in the slime mold Dictyostelium (Renart
et al. 1981; Sultana et al. 2012) and in the gravitactic sensory transduction chain of
Paramecium (Bräucker et al. 2001; Hemmersbach and Braun 2006).
The next question is how the cAMP signal controls the flagellar activity which is
instrumental in the steering response of the cell. Favaro et al. (2012) had shown that
cAMP can activate a protein kinase A. Staurosporine, which is known to be an
inhibitor of protein kinases (Chang and Kaufman 2000) impairs gravitaxis in Euglena
(Häder et al. 2010). But it is interesting that prolonged exposure (225 min) to the drug
reversed the negative gravitaxis into a positive one as did strong visible and UV
radiation or salt stress (see above). This drug also impairs negative phototaxis which
could indicate that both sensory transduction chains share the same final step being the
activation of a phosphokinase A (Häder and Iseki 2017).
Using the same molecular biological tools involving degenerate primers as
described above showed that Euglena possesses at least five isoforms of protein
kinase A (PK.1–PK.5). The full range of the RNA sequence was revealed by RACEPCR. RNAi against the different isoforms showed that PK.1–PK.3 and PK.5 are
not involved in the gravitactic sensory transduction chain (Daiker et al. 2011). In
contrast, inhibition of PK.4 effectively blocked gravitaxis for several weeks. Three
weeks after the RNAi treatment a positive gravitaxis was observed. This finding
parallels the result by staurosporine inhibition. In addition, RNAi of PK.4 blocked
phototaxis which further confirms that gravitaxis and phototaxis share the same final
step, the activation of a protein kinase A by cAMP.
3.7 Conclusions and Open Questions
Gravitactic unicellular organisms are specialized for gravity sensing. They use this
information for their spatial orientation (gravitaxis). Summarizing the findings
allows us to construct a complete gravitaxis signal transduction chain, such as in
Euglena (Fig. 3.6). During forward locomotion the cell rotates around its horizontal
axis at about 1 Hz. When the flagellum points downwards the content of the cell
presses onto the lower membrane activating the TRP channels, thought to be located
at the front end adjacent to the flagellum, which allows Ca
2+ to enter the cell along
a previously established gradient by a Ca-ATPase. In fact, several rotations are
3.7 Conclusions and Open Questions
39