Overexpression of Partner of Numb Induces Asymmetric Distribution of the PI4P 5-Kinase Skittles in Mitotic Sensory Organ Precursor Cells in Drosophila

Unequal segregation of cell fate determinants at mitosis is a conserved mechanism whereby cell fate diversity can be generated during development. In Drosophila, each sensory organ precursor cell (SOP) divides asymmetrically to produce an anterior pIIb and a posterior pIIa cell. The Par6-aPKC complex localizes at the posterior pole of dividing SOPs and directs the actin-dependent localization of the cell fate determinants Numb, Partner of Numb (Pon) and Neuralized at the opposite pole. The plasma membrane lipid phosphatidylinositol (4,5)-bisphosphate (PIP2) regulates the plasma membrane localization and activity of various proteins, including several actin regulators, thereby modulating actin-based processes. Here, we have examined the distribution of PIP2 and of the PIP2-producing kinase Skittles (Sktl) in mitotic SOPs. Our analysis indicates that both Sktl and PIP2 reporters are uniformly distributed in mitotic SOPs. In the course of this study, we have observed that overexpression of full-length Pon or its localization domain (LD) fused to the Red Fluorescent Protein (RFP::PonLD) results in asymmetric distribution of Sktl and PIP2 reporters in dividing SOPs. Our observation that Pon overexpression alters polar protein distribution is relevant because RFP::PonLD is often used as a polarity marker in dividing progenitors

Gastrokine-1, an anti-amyloidogenic protein secreted by the stomach, regulates diet-induced obesity

Obesity and its sequelae have a major impact on human health. The stomach contributes to obesity in ways that extend beyond its role in digestion, including through effects on the microbiome. Gastrokine-1 (GKN1) is an anti-amyloidogenic protein abundantly and specifically secreted into the stomach lumen. We examined whether GKN1 plays a role in the development of obesity and regulation of the gut microbiome. Gkn1−/− mice were resistant to diet-induced obesity and hepatic steatosis (high fat diet (HFD) fat mass (g) = 10.4 ± 3.0 (WT) versus 2.9 ± 2.3 (Gkn1−/−) p < 0.005; HFD liver mass (g) = 1.3 ± 0.11 (WT) versus 1.1 ± 0.07 (Gkn1−/−) p < 0.05). Gkn1−/− mice also exhibited increased expression of the lipid-regulating hormone ANGPTL4 in the small bowel. The microbiome of Gkn1−/− mice exhibited reduced populations of microbes implicated in obesity, namely Firmicutes of the class Erysipelotrichia. Altered metabolism consistent with use of fat as an energy source was evident in Gkn1−/− mice during the sleep period. GKN1 may contribute to the effects of the stomach on the microbiome and obesity. Inhibition of GKN1 may be a means to prevent obesity

The role of epsins in Drosophila eye development

textThe goal of my doctoral work is to understand how proteins involved in vesicle trafficking contribute to proper animal development. To understand aspects of this process, I studied how two vesicle trafficking proteins, Liquid facets(Lqf)/epsin1 and D-Epsin-Related, affect Drosophila eye development. I determined that Lqf, an endocytosis protein, together with Fat facets (Faf), a deubiquitinating enzyme, regulate the Notch and Delta signaling in the developing Drosophila eye. Notch signaling pathway is used in most developmental processes and is dependent on its ligand Delta. Faf deubiquitinates Lqf in the signaling cells, thereby increasing Lqf protein levels and also levels of Delta endocytosis. This event is necessary for Notch activation in neighboring cells. Lqf probably works in concert with the E3 ubiquitin ligase Neuralized (Neur), which ubiquitinates Delta. These conclusions are consistent with a relatively new model describing an obligate role for endocytosis in the signaling cells to effect activation in neighboring cells. To understand how Lqf functions mechanistically in this process, I performed a structure/function analysis of the Lqf protein. Lqf proteins with strategic deletions of certain functional domains were tested for their ability to function in vivo. The major result of these experiments is that the N-terminal ENTH domain of Lqf and a protein without the ENTH domain each retain significant activity. This suggests that Lqf has two functions: the ENTH domain function and the ENTH-less function. These data are in contrast with the most popular model suggesting that ENTH-less epsins are non-functional proteins. I present possible models for how ENTH-less epsins may retain function. The final part of my thesis focuses on D-Epsin-Related (D-Epsin-R) protein. I showed that D-Epsin-R is a Golgi protein, like its homologs. Surprisingly, D-Epsin-R ENTH domain is not required for function because an ENTH-less D-Epsin-R can substitute for endogenous D-Epsin-R. Also, D-Epsin-R has essential and probably specific developmental roles in the eye as D-Epsin-R mutants exhibit impaired cell growth. This work suggests that epsins are specific components of certain developmental pathways.Institute for Cellular and Molecular Biolog

Drosophila liquid facets-Related encodes Golgi epsin and is an essential gene required for cell proliferation, growth, and patterning

AbstractEpsin and epsin-Related (epsinR) are multi-modular proteins that stimulate clathrin-coated vesicle formation. Epsin promotes endocytosis at the plasma membrane, and epsinR functions at the Golgi and early endosomes for trans-Golgi network/endosome vesicle trafficking. In Drosophila, endocytic epsin is known as Liquid facets, and it is essential specifically for Notch signaling. Here, by generating and analyzing loss-of-function mutants in the liquid facets-Related (lqfR) gene of Drosophila, we investigated the function of Golgi epsin in a multicellular context. We found that LqfR is indeed a Golgi protein, and that like liquid facets, lqfR is essential for Drosophila viability. In addition, primarily by analyzing mutant eye discs, we found that lqfR is required for cell proliferation, insulin-independent cell growth, and cell patterning, consistent with a role in one or several signaling pathways. Epsins in all organisms share an ENTH (epsin N-terminal homology) domain, which binds phosphoinositides enriched at the plasma membrane or the Golgi membrane. The epsinR ENTH domain is also the recognition element for particular cargos. By generating wild-type and mutant lqfR transgenes, we found that all apparent LqfR functions are independent of its ENTH domain. These results suggest that LqfR transports specific cargo critical to one or more signaling pathways, and lays the foundation for identifying those proteins

On the conservation of function of the Drosophila Fat facets deubiquitinating enzyme and Fam, its mouse homolog

The original publication can be found at www.springerlink.comFat facets is a Drosophila deubiquitinating enzyme required for eye development and early embryogenesis. Genetic evidence suggests that Fat facets deubiquitinates and thereby prevents the proteasomal degradation of specific substrates. The Drosophila Liquid facets protein is implicated as the critical substrate of Fat facets in the eye. A mouse homolog of Fat facets, called Fam, has been identified. The results of biochemical experiments implicate two different proteins, Af-6 and β-catenin, as substrates for Fam. Here, the functional relationship between Fat facets and Fam is explored. We show that Fam can substitute for Fat facets in all of its essential functions in Drosophila. In addition, we tested the hypothesis that Canoe and Armadillo, the Drosophila homologs of Af-6 and β-catenin, respectively, are important substrates for Fat facets in the Drosophila eye. We found no genetic evidence to support a role for either Canoe or Armadillo in the essential Fat facets pathways in Drosophila eye development. The significance of these results is discussed in light of the biochemical experiments that suggest that Af-6 and β-catenin are substrates of Fam

Abortion

Over time abortion has moved from a medical issue to a political one. This presentation covers the issue of abortion as it pertains to American politics. Its arguments are based on the pro-choice platform supporting a womens rights to choose. The presentation covers a variety of modern arguments including bodily autonomy and more effective methods to decreasing abortion rates, as well as historical precedents set by past laws and the Supreme Court

PIP2 Reporters and GFP::Sktl localized uniformly in SOPs in the absence of RFP::Pon^LD.

<p>The distribution of PH::GFP (A), ENTH::GFP (B) and GFP::Sktl (C–D″) was examined by live imaging (A–C) or by antibody staining (anti-GFP in green and anti-Senseless in red in D–D″) in dividing SOPs at prometaphase. When RFP::Pon^LD was not co-expressed in SOPs, the PIP2 reporters and GFP::Sktl (panel C; 100%; n = 20) localized uniformly. All transgenes were expressed under the control of neur^P72Gal4Gal80^ts.</p