The Characterization of two Oriental Beetle Pheromone Insensitive Mutants obi-1 and obi-3 in Pristionchus pacificus

Although biologists have been aware of nematode associations with insects for some time, the details of these interactions have received little attention. A systematic effort to identify the natural ecology of Pristionchus nematodes revealed species-specific host preferences for several beetle species, including the oriental beetle found in Japan and northeastern U.S. In Pristionchus pacificus, the first and only molecular component identified so far that is involved in odor signaling is the cGMP dependent protein kinase, Ppa-EGL-4. To obtain more upstream factors involved in insect pheromone attraction, we performed a non-saturating X-ray mutagenesis on P. pacificus for mutants that do not show attraction toward their host oriental beetle pheromone (ZTDO) using the conventional plate chemotaxis assay. Of the seven obi candidates isolated, we selected two lines for further analyses due to their strong obi chemotaxis phenotype, viability, as well as linked morphological phenotypes after >3x outcrossing to wildtype. The two promising candidates were named obi-1 and obi-3 for Oriental Beetle pheromone Insensitive mutants. The chemosensory phenotype of obi-1 and obi-3 is specific for ZTDO and does not affect the chemoattraction toward other known P. pacificus attractants such as the plant volatile β-caryophyllene and the moth pheromone ETDA. Neither obi-1 nor obi-3 show dauer formation defect (daf-c) or temperature sensitivity. However, both obi-1 and obi-3 animals show higher frequency and amplitude turns than wildtype on OP50. Adult obi-1 hermaphrodites are ~20%; longer than wildtype, with a body length-to-width ratio of 17 compared to 13 in the wildtype. An increase in egg retention (egl) also accompany this long phenotype (lon). Because the chemotaxis assay with cGMP treatment is extremely labor intensive, we used a linked long body morphological phenotype in obi-1 to conduct subsequent mapping of F2 progeny from crosses between obi-1 and the mapping strain Washington. Using 200 F2 lines and molecular markers (Single Stranded Conformational Polymorphisms and Simple Sequence Length Polymorphisms), we were able to confine obi-1 to a 153 kb region represented by 10 subcontigs of Supercontig 58. No known C. elegans long mutants map near the syntenic chromosome region in P. pacificus. Obi-1’s odor specific phenotype and genetic location suggest that it is not an allele of egl-4 (Chr IV) and thus likely to be a parallel or upstream component of odor signaling in P. pacificus

A host beetle pheromone regulates development and behavior in the nematode Pristionchus pacificus

Nematodes and insects are the two most speciose animal phyla and nematode-insect associations encompass widespread biological interactions. To dissect the chemical signals and the genes mediating this association, we investigated the effect of an oriental beetle sex pheromone on the development and behavior of the nematode Pristionchus pacificus. We found that while the beetle pheromone is attractive to P. pacificus adults, the pheromone arrests embryo development, paralyzes J2 larva, and inhibits exit of dauer larvae. To uncover the mechanism that regulates insect pheromone sensitivity, a newly identified mutant, Ppa-obi-1, is used to reveal the molecular links between altered attraction towards the beetle pheromone, as well as hypersensitivity to its paralyzing effects. Ppa-obi-1 encodes lipid-binding domains and reaches its highest expression in various cell types, including the amphid neuron sheath and excretory cells. Our data suggest that the beetle host pheromone may be a species-specific volatile synomone that co-evolved with necromeny

let-7 miRNAs can act through notch to regulate human gliogenesis.

It is clear that neural differentiation from human pluripotent stem cells generates cells that are developmentally immature. Here, we show that the let-7 plays a functional role in the developmental decision making of human neural progenitors, controlling whether these cells make neurons or glia. Through gain- and loss-of-function studies on both tissue and pluripotent derived cells, our data show that let-7 specifically regulates decision making in this context by regulation of a key chromatin-associated protein, HMGA2. Furthermore, we provide evidence that the let-7/HMGA2 circuit acts on HES5, a NOTCH effector and well-established node that regulates fate decisions in the nervous system. These data link the let-7 circuit to NOTCH signaling and suggest that this interaction serves to regulate human developmental progression

let-7 miRNAs Can Act through Notch to Regulate Human Gliogenesis

It is clear that neural differentiation from human pluripotent stem cells generates cells that are developmentally immature. Here, we show that the let-7 plays a functional role in the developmental decision making of human neural progenitors, controlling whether these cells make neurons or glia. Through gain- and loss-of-function studies on both tissue and pluripotent derived cells, our data show that let-7 specifically regulates decision making in this context by regulation of a key chromatin-associated protein, HMGA2. Furthermore, we provide evidence that the let-7/HMGA2 circuit acts on HES5, a NOTCH effector and well-established node that regulates fate decisions in the nervous system. These data link the let-7 circuit to NOTCH signaling and suggest that this interaction serves to regulate human developmental progression

A small molecule screen to identify regulators of let-7 targets

The let-7 family of miRNAs has been shown to be crucial in many aspects of biology, from the regulation of developmental timing to cancer. The available methods to regulate this family of miRNAs have so far been mostly genetic and therefore not easily performed experimentally. Here, we describe a small molecule screen designed to identify regulators of let-7 targets in human cells. In particular, we focused our efforts on the identification of small molecules that could suppress let-7 targets, as these could serve to potentially intercede in tumors driven by loss of let-7 activity. After screening through roughly 36,000 compounds, we identified a class of phosphodiesterase inhibitors that suppress let-7 targets. These compounds stimulate cAMP levels and raise mature let-7 levels to suppress let-7 target genes in multiple cancer cell lines such as HMGA2 and MYC. As a result, these compounds also show growth inhibitory activity on cancer cells