Effects of Label-Dose Permethrin Administration on Reproductive Function and Embryo Quality on Superovulated Beef Heifers

Commercial pyrethroid pour-ons are commonly appliedin cow-calf operations to eliminate the potential for insectborne diseases and to improve productivity. However,recent literature has focused on potential negativereproductive effects in the bull after exposure to pyrethroids.While the female bovine has been primarily neglected fromthe debated pyrethroid concern on reproduction, literature inmice and rats have reported potential endocrine disruptionof sex steroids resulting from pyrethroid exposure, withpotential detrimental effects on female fertility. Theobjective was to study the effects of a commercialpyrethroid-based pour-on product, permethrin, onreproductive performance in superovulated beef heifers byassessing steroid biosynthesis and embryo quality. It washypothesized that exposure to pyrethroid pour-on at labeldose would cause minimal effects on embryoquantity/quality and steroidogenesis in the female bovine.Results from this study revealed pyrethroid-treated heifershad a tendency for reduced progesterone, but embryoquantity and quality were not affected compared to control

Illuminating G␤ 5 Signaling

ABSTRACT G proteins are key intermediates in cellular signaling and act in response to a variety of extracellular stimuli. The prevailing paradigm is that G protein subunits form a heterotrimeric complex and function principally at the plasma membrane. However, there is growing evidence for localization at, and signaling by, G proteins at intracellular compartments. Moreover, different cellular pools of G proteins may be composed of distinct subunit subtypes, including some binding partners that function in the place of G protein ␥ subunits. An article in this issue of Molecular Pharmacology (Yost et al., p. 812) describes the use of an innovative fluorescent cell imaging technique to study interactions of the G protein ␤ 5 subunit with a panel of G␥ subunits as well as regulator of G protein signaling (RGS) proteins that contain a G␥-like subdomain. The approach used here provides a new strategy to elucidate the spatial and temporal properties of G proteins, including a growing number of atypical G␤␥ pairings. Heterotrimeric G proteins normally consist of ␣, ␤, and ␥ subunits and are coupled to seven transmembrane receptors at the plasma membrane. Agonist binding to the receptor induces a conformational change of the G␣ subunit promoting the release of GDP and binding to GTP. This exchange triggers G␤␥ disassociation from the G␣, freeing both components to modulate downstream signals. Hydrolysis of GTP to GDP by the G␣ results in reassociation of the heterotrimer and termination of the signal (Sprang, 1997). So far, 23 G␣, 5 G␤, and 12 G␥ subunits have been identified in the mammalian genome. Of the G␤ isoforms, types 1 to 4 are highly conserved, sharing 80% sequence identity, but G␤ 5 is divergent, sharing only 50% identity. Like other ␤ isoforms, G␤ 5 interacts with G␥ subunits; unlike the others, G␤ 5 can also interact with RGS proteins from the R7 family (RGS6, RGS7, RGS9, and RGS11) The RGS/G␤ 5 complex could be thought of as a highly atypical G␤␥ pair. Others are likely to exist (see below). With the identification of such atypical subunit complexes, new techniques are needed to ascertain their function within the cell. Bimolecular fluorescence complementation (BiFC) is one promising technique In this issue of Molecular Pharmacology

Effects of Label Dose Permethrin Use in Yearling Angus Beef Bulls on Reproductive Function and Testicular Histopathology

To eliminate the potential for insect borne diseases and improve productivity in cow-calf operations, many producers use pour-on pyrethroids. However, popular press literature has identified potential links between use of pyrethroids and reproductive function failures in bulls. While literature in mice and rats have reported potential endocrine disruption of sex steroids resulting from pyrethroid exposure, effects of pyrethroid use on bull fertility is still questionable. The objective of this study was to measure reproductive parameters in peripubertal Angus bulls using a commonly used pyrethroid pour-on. We hypothesized that use of a pyrethroid pour-on at labeled dose would have limited effects on semen and testicular characteristics. Results from this study revealed pyrethroid-treated bulls had greater spermatozoa head and midpiece abnormalities compared to controls, resulting in higher primary morphological abnormalities. Although some morphological semen parameters appear to be negatively affected by use of pyrethroid administration in bulls, biological relevance of this result needs to be further elucidated as the ability to pass a breeding soundness exam was not affected. Moreover, further research should be conducted to determine the effects of pyrethroid pour-ons on fertility when used with other pyrethroids such fly tags or perimeter sprays. Based on these results, pour-on pyrethroid use in yearling bulls is still recommended for label dose administration

The RACK1 Ortholog Asc1 Functions as a G-protein β Subunit Coupled to Glucose Responsiveness in Yeast

According to the prevailing paradigm, G-proteins are composed of three subunits, an alpha subunit with GTPase activity and a tightly associated betagamma subunit complex. In the yeast Saccharomyces cerevisiae there are two known Galpha proteins (Gpa1 and Gpa2) but only one Gbetagamma, which binds only to Gpa1. Here we show that the yeast ortholog of RACK1 (receptor for activated protein kinase C1) Asc1 functions as the Gbeta for Gpa2. As with other known Gbeta proteins, Asc1 has a 7-WD domain structure, interacts directly with the Galpha in a guanine nucleotide-dependent manner, and inhibits Galpha guanine nucleotide exchange activity. In addition, Asc1 binds to the effector enzyme adenylyl cyclase (Cyr1), and diminishes the production of cAMP in response to glucose stimulation. Thus, whereas Gpa2 promotes glucose signaling through elevated production of cAMP, Asc1 has opposing effects on these same processes. Our findings reveal the existence of an unusual Gbeta subunit, one having multiple functions within the cell in addition to serving as a signal transducer for cell surface receptors and intracellular effectors

Structure and Function of Vps15 in the Endosomal G Protein Signaling Pathway ,

G protein-coupled receptors mediate cellular responses to a wide variety of stimuli, including taste, light and neurotransmitters. In the yeast Saccharomyces cerevisiae, activation of the pheromone pathway triggers events leading to mating. The view had long been held that the G protein-mediated signal occurs principally at the plasma membrane. Recently, it has been shown that the G protein α subunit Gpa1 can promote signaling at endosomes and requires two components of the sole phosphatidylinositol-3-kinase in yeast, Vps15 and Vps34. Vps15 contains multiple WD repeats and also binds to Gpa1 preferentially in the GDP-bound state; these observations led us to hypothesize that Vps15 may function as a G protein β subunit at the endosome. Here we show an X-ray crystal structure of the Vps15 WD domain that reveals a seven-bladed propeller resembling that of typical Gβ subunits. We show further that the WD domain is sufficient to bind Gpa1 as well as to Atg14, a potential Gγ protein that exists in a complex with Vps15. The Vps15 kinase domain together with the intermediate domain (linking the kinase and WD domains) also contributes to Gpa1 binding, and is necessary for Vps15 to sustain G protein signaling. These findings reveal that the Vps15 Gβ-like domain serves as a scaffold to assemble Gpa1 and Atg14, whereas the kinase and intermediate domains are required for proper signaling at the endosome

Multi-Omics Analysis of Multiple Glucose-Sensing Receptor Systems in Yeast

The yeast Saccharomyces cerevisiae has long been used to produce alcohol from glucose and other sugars. While much is known about glucose metabolism, relatively little is known about the receptors and signaling pathways that indicate glucose availability. Here, we compare the two glucose receptor systems in S. cerevisiae. The first is a heterodimer of transporter-like proteins (transceptors), while the second is a seven-transmembrane receptor coupled to a large G protein (Gpa2) that acts in coordination with two small G proteins (Ras1 and Ras2). Through comprehensive measurements of glucose-dependent transcription and metabolism, we demonstrate that the two receptor systems have distinct roles in glucose signaling: the G-protein-coupled receptor directs carbohydrate and energy metabolism, while the transceptors regulate ancillary processes such as ribosome, amino acids, cofactor and vitamin metabolism. The large G-protein transmits the signal from its cognate receptor, while the small G-protein Ras2 (but not Ras1) integrates responses from both receptor pathways. Collectively, our analysis reveals the molecular basis for glucose detection and the earliest events of glucose-dependent signal transduction in yeast

Synthetic Morphology Using Alternative Inputs

Designing the shape and size of a cell is an interesting challenge for synthetic biology. Prolonged exposure to the mating pheromone α-factor induces an unusual morphology in yeast cells: multiple mating projections. The goal of this work was to reproduce the multiple projections phenotype in the absence of α-factor using a gain-of-function approach termed “Alternative Inputs (AIs)”. An alternative input is defined as any genetic manipulation that can activate the signaling pathway instead of the natural input. Interestingly, none of the alternative inputs were sufficient to produce multiple projections although some produced a single projection. Then, we extended our search by creating all combinations of alternative inputs and deletions that were summarized in an AIs-Deletions matrix. We found a genetic manipulation (AI-Ste5p ste2Δ) that enhanced the formation of multiple projections. Following up this lead, we demonstrated that AI-Ste4p and AI-Ste5p were sufficient to produce multiple projections when combined. Further, we showed that overexpression of a membrane-targeted form of Ste5p alone could also induce multiple projections. Thus, we successfully re-engineered the multiple projections mating morphology using alternative inputs without α-factor