The exocyst complex regulates the last steps of exocytosis, which is essential to organisms across kingdoms. In humans, its dysfunction is correlated with several significant diseases, such as diabetes and cancer progression. Investigation of the dynamic regulation of the evolutionarily conserved exocyst-related processes using mutants in genetically tractable organisms such as Arabidopsis thaliana is limited by the lethality or the severity of phenotypes. We discovered that the small molecule Endosidin2 (ES2) binds to the EXO70 (exocyst component of 70 kDa) subunit of the exocyst complex, resulting in inhibition of exocytosis and endosomal recycling in both plant and human cells and enhancement of plant vacuolar trafficking. An EXO70 protein with a C-terminal truncation results in dominant ES2 resistance, uncovering possible distinct regulatory roles for the N terminus of the protein. This study not only provides a valuable tool in studying exocytosis regulation but also offers a potentially new target for drugs aimed at addressing human disease.T he EXO70 (exocyst component of 70 kDa) protein is a component of the evolutionarily conserved octameric exocyst complex that tethers post-Golgi vesicles to the plasma membrane before SNARE-mediated membrane fusion (1). As an important component of the exocyst complex that mediates exocytosis, EXO70 regulates, for example, neurite outgrowth, epithelial cell polarity establishment, cell motility, and cell morphogenesis in animal cells (2-6). In plants, EXO70 proteins participate in polarized pollen tube growth, root hair growth, deposition of cell wall material, cell plate initiation and maturation, defense, and autophagy (7-12). In humans, EXO70 mediates the trafficking of the glucose transporter Glut4 to the plasma membrane that is stimulated by insulin and involved in the development of diabetes (13). A specific isoform of human EXO70 is also involved in cancer cell invasion (13-15). Endosidin2 (ES2) was identified from a plantbased chemical screen as an inhibitor of trafficking. We demonstrate that the target of ES2 is the EXO70 subunit of the exocyst and that ES2 is active in plants and mammalian systems. Significantly, no inhibitor of the exocyst complex has been reported, yet such compounds could be important for understanding the basic mechanisms of exocyst-mediated processes, for modifying secretion in biotechnological applications, and for the development of potential new drugs with higher affinity and more potent activity to control exocyst-related diseases. Results ES2 InhibitsTrafficking to the Plasma Membrane. ES2 is a previously identified plant endomembrane trafficking disruptor (Fig. 1A) that inhibits polarized growth of pollen tubes in a dose-dependent manner (Fig. S1 A and B) (16). Arabidopsis seedlings grown on media containing ES2 have shorter roots and fewer and shorter root hairs and are less sensitive to gravity stimulation (Fig. S1 C-G). ES2 disrupted the trafficking of proteins that are actively recycled between the plasma membrane and endosome...

Dysfunction of the striatum is frequently associated with sleep disturbances. However, its role in sleep-wake regulation has been paid little attention even though the striatum densely expresses adenosine A2A receptors (A2ARs), which are essential for adenosine-induced sleep. Here we showed that chemogenetic activation of A2AR neurons in specific subregions of the striatum induced a remarkable increase in non-rapid eye movement (NREM) sleep. Anatomical mapping and immunoelectron microscopy revealed that striatal A2AR neurons innervated the external globus pallidus (GPe) in a topographically organized manner and preferentially formed inhibitory synapses with GPe parvalbumin (PV) neurons. Moreover, lesions of GPe PV neurons abolished the sleep-promoting effect of striatal A2AR neurons. In addition, chemogenetic inhibition of striatal A2AR neurons led to a significant decrease of NREM sleep at active period, but not inactive period of mice. These findings reveal a prominent contribution of striatal A2AR neuron/GPe PV neuron circuit in sleep control.

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Upstream ORFs are elements found in the 5′-leader sequences of specific mRNAs that modulate the translation of downstream ORFs encoding major gene products. In Arabidopsis, the translational control of auxin response factors (ARFs) by upstream ORFs has been proposed as a regulatory mechanism required to respond properly to complex auxin-signaling inputs. In this study, we identify and characterize the aberrant auxin responses in specific ribosomal protein mutants in which multiple ARF transcription factors are simultaneously repressed at the translational level. This characteristic lends itself to the use of these mutants as genetic tools to bypass the genetic redundancy among members of the ARF family in Arabidopsis. Using this approach, we were able to assign unique functions for ARF2, ARF3, and ARF6 in plant development.ribosomal mutants | uORFs | auxin regulation