A novel function for the IκB inhibitor Cactus in promoting Dorsal nuclear localization and activity in the Drosophila embryo

Abstract

The evolutionarily conserved Toll signaling pathway controls innate immunity across phyla and embryonic patterning in insects. In the embryo, Toll is required to establish gene expression domains along the dorsal-ventral axis. Pathway activation induces degradation of the IκB inhibitor Cactus, resulting in a ventral-to-dorsal nuclear gradient of the NFκB effector Dorsal. Here, we investigate how modulates Toll signals through its effects on the Dorsal gradient and on Dorsal target genes. Quantitative analysis using a series of loss- and gain-of-function conditions shows that the ventral and lateral aspects of the Dorsal gradient can behave differently with respect to Cactus fluctuations. In lateral and dorsal embryo domains, loss of Cactus allows more Dorsal to translocate to the nucleus. Unexpectedly, loss-of-function alleles decrease Dorsal nuclear localization ventrally, where Toll signals are high. Overexpression analysis suggests that this ability of Cactus to enhance Toll stems from the mobilization of a free Cactus pool induced by the Calpain A protease. These results indicate that Cactus acts to bolster Dorsal activation, in addition to its role as a NFκB inhibitor, ensuring a correct response to Toll signals.

A Reaction-Diffusion Network model predicts a dual role of Cactus/IκB to regulate Dorsal/NFκB nuclear translocation inDrosophila

“…One fundamental characteristic of the embryonic Dl gradient is that it is robust to variations in Dl and Cact levels, tolerating a Dl/Cact ratio of 0.3 to 2.0 without a significant effect on embryo viability (Govind et al, 1993). In order to test how the amount of total Dl protein impacts its nuclear localization, we simulated nDl levels in embryos generated by mothers heterozygous for a loss-of-function dl allele (dl[6], Cardoso et al, 2017). These embryos (from dl[6]/+ mothers) produce less Dl protein than embryos generated from wildtype mothers (Cardoso et al, 2017).…”

“…In order to test how the amount of total Dl protein impacts its nuclear localization, we simulated nDl levels in embryos generated by mothers heterozygous for a loss-of-function dl allele (dl[6], Cardoso et al, 2017). These embryos (from dl[6]/+ mothers) produce less Dl protein than embryos generated from wildtype mothers (Cardoso et al, 2017). Even though embryos from dl[6]/+ mothers show a reduction in peak nDl in the ventral domain, they do not show a significant change in nDl levels in either the lateral or dorsal regions ( Fig.…”

“…Even though embryos from dl[6]/+ mothers show a reduction in peak nDl in the ventral domain, they do not show a significant change in nDl levels in either the lateral or dorsal regions ( Fig. 3A, Cardoso et al, 2017;Carrell et al, 2017).…”

“…We have previously shown that Cact displays not only a role as inhibitor of Dl nuclear translocation, but also acts to favor nDl localization in ventral nuclei ( Fig. 1 in Cardoso et al, 2017). In order to use our model to unveil the molecular mechanism behind this dual role exhibited by Cact, we simulated the pattern of hypomorphic cact allelic combinations.…”

“…Recently, by analyzing the effects of a series of cact and dorsal (dl) loss-of-function alleles, we detected a novel function of Cact to promote Dl nuclear translocation. In addition to inhibiting Dl nuclear import, Cact also acts to favor Dl nuclear translocation where Toll signals are high (Cardoso et al, 2017). However, the mechanism behind this effect is unclear.…”