Mechanisms Underlying Stage-1 TRPL Channel Translocation in Drosophila Photoreceptors

Background: TRP channels function as key mediators of sensory transduction and other cellular signaling pathways. In Drosophila, TRP and TRPL are the light-activated channels in photoreceptors. While TRP is statically localized in the signaling compartment of the cell (the rhabdomere), TRPL localization is regulated by light. TRPL channels translocate out of the rhabdomere in two distinct stages, returning to the rhabdomere with dark-incubation. Translocation of TRPL channels regulates their availability, and thereby the gain of the signal. Little, however, is known about the mechanisms underlying this trafficking of TRPL channels. Methodology/Principal Findings: We first examine the involvement of de novo protein synthesis in TRPL translocation. We feed flies cycloheximide, verify inhibition of protein synthesis, and test for TRPL translocation in photoreceptors. We find that protein synthesis is not involved in either stage of TRPL translocation out of the rhabdomere, but that re-localization to the rhabdomere from stage-1, but not stage-2, depends on protein synthesis. We also characterize an ex vivo eye preparation that is amenable to biochemical and genetic manipulation. We use this preparation to examine mechanisms of stage-1 TRPL translocation. We find that stage-1 translocation is: induced with ATP depletion, unaltered with perturbation of the actin cytoskeleton or inhibition of endocytosis, and slowed with increased membrane sterol content. Conclusions/Significance: Our results indicate that translocation of TRPL out of the rhabdomere is likely due to protei

Calmodulin interacts with MLO protein to regulate defence against mildew in barley

In plants, defence against specific isolates of a pathogen can be triggered by the presence of a corresponding race-specific resistance gene, whereas resistance of a more broad-spectrum nature can result from recessive, presumably loss-of-regulatory-function, mutations. An example of the latter are mlo mutations in barley, which have been successful in agriculture for the control of powdery mildew fungus (Blumeria graminis f. sp. hordei; Bgh). MLO protein resides in the plasma membrane, has seven transmembrane domains, and is the prototype of a sequence-diversified family unique to plants, reminiscent of the seven-transmembrane receptors in fungi and animals. In animals, these are known as G-protein- coupled receptors and exist in three main families, lacking sequence similarity, that are thought to be an example of molecular convergence. MLO seems to function independently of heterotrimeric G proteins. We have identified a domain in MLO that mediates a Ca2+-dependent interaction with calmodulin in vitro. Loss of calmodulin binding halves the ability of MLO to negatively regulate defence against powdery mildew in vivo. We propose a sensor role for MLO in the modulation of defence reactions.close17017

UNC119 is required for G protein trafficking in sensory neurons

UNC119 is widely expressed among vertebrates and invertebrates. Here we report that UNC119 recognized the acylated N-terminus of the rod photoreceptor transducin α-subunit (Tα) as well as C. elegans G proteins Odr-3 and Gpa-13. The crystal structure of human UNC119 at 1.95 Å resolution revealed an immunoglobulin-like β-sandwich fold. Pulldowns and isothermal titration calorimetry revealed a tight interaction between UNC119 and acylated Gα peptides. Co-crystallization of UNC119 with an acylated Tα N-terminal peptide at 2.0 Å revealed that the lipid chain is buried deeply into UNC119's hydrophobic cavity. UNC119 bound Tα(GTP) inhibiting its GTPase activity, thereby providing a stable UNC119-Tα(GTP) complex that is capable of diffusing from the inner segment back to the outer segment following light-induced translocation. UNC119 deletion in both mouse and C. elegans lead to G protein mislocalization. These results establish UNC119 as a novel Gα-subunit cofactor that is essential for G-protein trafficking in sensory cilia