Fig. 6. Osmotic stress-induced activation of Dd-STATc in Dictyostelium strains mutant in the cAMP- and cGMP-mediated stress-response pathways. The activation of Dd-STATc in response to sorbitol in the parental (Ax-2) strain and the indicated mutants was determined essentially as described in Fig. 2. However, after probing with CP22 (tyrP-STATc), the filter was reprobed with 7H3 (Total STATc) as described in Materials and Methods. The latter signal provides a convenient loading control. All the strains are published (see text), with the exception of the sgc-/dokA- strain. This was generated using as a start point the dokA- strain (Schuster et al., 1996), which was created using a G418 resistance cassette. An sgc blasticidin-based disruption cassette was created using a novel in vitro transposition technique (T. Abe and J. G. Williams, unpublished). PCR analysis shows that the mutated Dictyostelium cells contain a blasticidin-resistance cassette in the centre of the sgc gene, in the approximate position of the C1/C2 catalytic domains (Roeloffs et al., 2001). Consistent with this, we analysed osmotic stress induced guanylyl cyclase activation in the sgc-/dokA- strain (Roeloffs et al., 2000) and it is absent (data not shown).