Transforming growth factor β-induced phosphorylation of Smad3 is required for growth inhibition and transcriptional induction in epithelial cells

Drosophila Mad proteins are intracellular signal transducers of decapentaplegic (dpp), the Drosophila transforming growth factor β (TGF-β)/bone morphogenic protein (BMP) homolog. Studies in which the mammalian Smad homologs were transiently overexpressed in cultured cells have implicated Smad2 in TGF-β signaling, but the physiological relevance of the Smad3 protein in signaling by TGF-β receptors has not been established. Here we stably expressed Smad proteins at controlled levels in epithelial cells using a novel approach that combines highly efficient retroviral gene transfer and quantitative cell sorting. We show that upon TGF-β treatment Smad3 becomes rapidly phosphorylated at the SSVS motif at its very C terminus. Either attachment of an epitope tag to the C terminus or replacement of these three serine residues with alanine abolishes TGF-β-induced Smad3 phosphorylation; these proteins act in a dominant-negative fashion to block the antiproliferative effect of TGF-β in mink lung epithelial cells. A Smad3 protein in which the three C-terminal serines have been replaced by aspartic acids is also a dominant inhibitor of TGF-β signaling, but can activate plasminogen activator inhibitor 1 (PAI-1) transcription in a ligand-independent fashion when its nuclear localization is forced by transient overexpression. Phosphorylation of the three C-terminal serine residues of Smad3 by an activated TGF-β receptor complex is an essential step in signal transduction by TGF-β for both inhibition of cell proliferation and activation of the PAI-1 promoter.

A common polymorphism associated with antibiotic-induced cardiac arrhythmia

Drug-induced long QT syndrome (LQTS) is a prevalent disorder of uncertain etiology that predisposes to sudden death. KCNE2 encodes MinK-related peptide 1 (MiRP1), a subunit of the cardiac potassium channel IKr that has been associated previously with inherited LQTS. Here, we examine KCNE2 in 98 patients with drug-induced LQTS, identifying three individuals with sporadic mutations and a patient with sulfamethoxazole-associated LQTS who carried a single-nucleotide polymorphism (SNP) found in ≈1.6% of the general population. While mutant channels showed diminished potassium flux at baseline and wild-type drug sensitivity, channels with the SNP were normal at baseline but inhibited by sulfamethoxazole at therapeutic levels that did not affect wild-type channels. We conclude that allelic variants of MiRP1 contribute to a significant fraction of cases of drug-induced LQTS through multiple mechanisms and that common sequence variations that increase the risk of life-threatening drug reactions can be clinically silent before drug exposure.

A novel function for Myc: inhibition of C/EBP-dependent gene activation.

We have investigated the effect of the v-Myc oncoprotein on gene expression in myelomonocytic cells. We find that v-Myc dramatically down-regulates the expression of myelomonocytic-specific genes, such as the chicken mim-1 and lysozyme genes, both of which are known targets for C/EBP transcription factors. We present evidence that Myc downregulates these genes by inhibiting the function of C/EBP transcription factors. Detailed examination of the inhibitory mechanism shows that amino-terminal sequences of v-Myc, but not its DNA-binding domain, are required for the suppression of C/EBP-dependent transactivation. Our findings identify a new function for Myc and reveal a novel mechanism by which Myc affects the expression of other genes.

Complementation of the beige mutation in cultured cells by episomally replicating murine yeast artificial chromosomes.

Chédiak-Higashi syndrome in man and the beige mutation of mice are phenotypically similar disorders that have profound effects upon lysosome and melanosome morphology and function. We isolated two murine yeast artificial chromosomes (YACs) that, when introduced into beige mouse fibroblasts, complement the beige mutation. The complementing YACs exist as extrachromosomal elements that are amplified in high concentrations of G418. When YAC-complemented beige cells were fused to human Chédiak-Higashi syndrome or Aleutian mink fibroblasts, complementation of the mutant phenotype also occurred. These results localize the beige gene to a 500-kb interval and demonstrate that the same or homologous genes are defective in mice, minks, and humans.

A corticosteroid-induced gene expressing an "IsK-like" K+ channel activity in Xenopus oocytes.

Screening a rat colon cDNA library for aldosterone-induced genes resulted in the molecular cloning of a cDNA whose corresponding mRNA is strongly induced in the colon by dexamethasone, aldosterone, and a low NaCl diet. A similar mRNA was detected in kidney papilla but not in brain, heart, or skeletal muscle. Xenopus laevis oocytes injected with cRNA synthesized from this clone, designated CHIF (channel-inducing factor), express a K(+)-specific channel activity. The biophysical, pharmacological, and regulatory characteristics of this channel are very similar to those reported before for IsK (minK). These include: slow (tau > 20 s) activation by membrane depolarization with a threshold potential above -50 mV, blockade by clofilium, inhibition by phorbol ester, and activation by 8-bromoadenosine 3',5'-cyclic monophosphate and high cytoplasmic Ca2+. The primary structure of this clone, however, shows no homology to IsK. Instead, CHIF exhibits > 50% similarity to two other short bitopic membrane proteins, phospholemman and the gamma subunit of Na+K(+)-ATPase. The data are consistent with the possibility that CHIF is a member of a family of transmembrane regulators capable of activating endogenous oocyte transport proteins.

On the formation and evolution of the first Be star in a black hole binary MWC 656

We find that the formation of MWC 656 (the first Be binary containing a black hole) involves a common envelope phase and a supernova explosion. This result supports the idea that a rapidly rotating Be star can emerge out of a common envelope phase, which is very intriguing because this evolutionary stage is thought to be too fast to lead to significant accretion and spin up of the B star. We predict ∼10–100 of B-BH binaries to currently reside in the Galactic disc, among which around 1/3 contain a Be star, but there is only a small chance to observe a system with parameters resembling MWC 656. If MWC 656 is representative of intrinsic Galactic Be-BH binary population, it may indicate that standard evolutionary theory needs to be revised. This would pose another evolutionary problem in understanding black hole (BH) binaries, with BH X-ray novae formation issue being the prime example. Future evolution of MWC 656 with an ∼5 M⊙ BH and with an ∼13 M⊙ main-sequence companion on an ∼60 d orbit may lead to the formation of a coalescing BH–NS (neutron star) system. The estimated Advanced LIGO/Virgo detection rate of such systems is up to ∼0.2 yr−1. This empirical estimate is a lower limit as it is obtained with only one particular evolutionary scenario, the MWC 656 binary. This is only a third such estimate available (after Cyg X-1 and Cyg X-3), and it lends additional support to the existence of so far undetected BH–NS binaries.

2000 Ice Hockey Team

Front Row (L-R): Asst. Coach Billy Powers, Josh Blackburn, Krikor Arman, Sean Peach, Head Coach Red Berenson, Andrew Merrick, Kevin Magnuson, Kevin O'Malley, Associate Head Coach Mel Pearson

Second Row (L-R): L.J. Scarpace, Andy Hilbert, Bob Gassoff, Bill Trainor, Scott Matzka, Mark Kosick, Mike Comrie, John Shouneyia, Mike Cammalleri, J.J. Swistak

Third Row (L-R): Mark Mink, Craig Murray, Jay Vancik, Jeff Jillson, Dave Huntzicker, Josh Langfeld, Geoff Koch, Jed Ortmeyer

Fourth Row (L-R): Athletic Trainer Rick Bancroft, Student Equipment Manager Eric Sikkenga, Equipment Manager Ian Hume, Student Equipment Manager Jeff Conrad, Student Equipment Manager Royce Wilkerson, Video Coordinator Josh Richelew

2000 Ice Hockey Team posed with CCHA Trophy

Front Row (L-R): Asst. Coach Billy Powers, Josh Blackburn, Krikor Arman, Sean Peach, Head Coach Red Berenson, Andrew Merrick, Kevin Magnuson, Kevin O'Malley, Associate Head Coach Mel Pearson

Second Row (L-R): L.J. Scarpace, Andy Hilbert, Bob Gassoff, Bill Trainor, Scott Matzka, Mark Kosick, Mike Comrie, John Shouneyia, Mike Cammalleri, J.J. Swistak

Third Row (L-R): Mark Mink, Craig Murray, Jay Vancik, Jeff Jillson, Dave Huntzicker, Josh Langfeld, Geoff Koch, Jed Ortmeyer

Fourth Row (L-R): Athletic Trainer Rick Bancroft, Student Equipment Manager Eric Sikkenga, Equipment Manager Ian Hume, Student Equipment Manager Jeff Conrad, Student Equipment Manager Royce Wilkerson, Video Coordinator Josh Richelew

Birth and mortality statistics of the Virgin Islands of the United States. 1901-1919.

On t.-p., seal of Department of the navy, Bureau of medicine & surgery.

Sefer Liḳuṭe Yosef /

Bound with: Sefer Yesod Yosef / Yosef Yosḳa ben Yitsḥaḳ, ha-Leṿi.

An analytic study of a shared device among independent computing systems /

Item 247.

2001 Ice Hockey Team

Front Row (L-R): Asst. Coach Billy Powers, Josh Blackburn, Josh Langfeld, Scott Matzka, Dave Huntzicker, Bob Gassoff, L.J. Scarpace, Head Coach Red Berenson

Second Row (L-R): Goaltender coach Stan Matwijiw, Kevin O'Malley, John Shouneyia, Mark Mink, Mike Roemensky, Mark Kosick, Geoff Koch, Bill Trainor, Andy Hilbert, Craig Murray, Mike Cammalleri, J.J. Swistak

Third Row (L-R): Rob Kohen, Brad Fraser, Jay Vancik, Jeff Jillson, Mike Komisarek, Jed Ortmeyer, David Wyzgowski, Joe Kautz, Andy Burnes

Fourth Row (L-R): Administrative Assistant Brian Wiseman, Student Equipment Manager Eric Sikkenga, Student Athletic Trainer Jennifer Reynolds, Video Coordinator Ryan Rezmierski, Student Equipment Manager Royce Wilkerson, Equipment Manager Ian Hume, Student Equipment Manager David Brooks, Athletic Trainer Rick Bancroft

2002 Ice Hockey Team

Front Row (L-R): Associate Head Coach Mel Pearson, Josh Blackburn, Jay Vancik, Mike Cammalleri, Head Coach Red Berenson, John Shouneyia, Craig Murray, Kevin O'Malley, Assistant Coach Billy Powers.

Second Row (L-R): Justin Spurlock, Milan Gajic, Eric Werner, David Wyzgowski, Joe Kautz, Brad Fraser, Jed Ortmeyer, Andy Burnes, Mike Roemensky, Mark Mink, J.J. Swistak.

Third Row (L-R): Michael Woodford, Charlie Henderson, Dwight Helminen, Reilly Olson, Jason Ryznar, Brandon Rogers, Eric Nystrom, David Moss, Nick Martens, Mike Komisarek.

Fourth Row (L-R): Video Coordinator Ryan Rezmierski, Administrative Assistant Brian Wiseman, Athletic Trainer Rick Bancroft, Student Athletic Trainer Jesse Johnson, Equipment Manager Ian Hume, Assistant Equipment Manager David Brooks.

Do changes in transglutaminase activity alter latent transforming growth factor beta activation in experimental diabetic nephropathy?

Background. Diabetic nephropathy is the leading cause of end-stage kidney failure worldwide. It is characterized by excessive extracellular matrix accumulation. Transforming growth factor beta 1 (TGF-ß1) is a fibrogenic cytokine playing a major role in the healing process and scarring by regulating extracellular matrix turnover, cell proliferation and epithelial mesanchymal transdifferentiation. Newly synthesized TGF-ß is released as a latent, biologically inactive complex. The cross-linking of the large latent TGF-ß to the extracellular matrix by transglutaminase 2 (TG2) is one of the key mechanisms of recruitment and activation of this cytokine. TG2 is an enzyme catalyzing an acyl transfer reaction leading to the formation of a stable e(?-glutamyl)-lysine cross-link between peptides.Methods. To investigate if changes in TG activity can modulate TGF-ß1 activation, we used the mink lung cell bioassay to assess TGF-ß activity in the streptozotocin model of diabetic nephropathy treated with TG inhibitor NTU281 and in TG2 overexpressing opossum kidney (OK) proximal tubular epithelial cells.Results. Application of the site-directed TG inhibitor NTU281 caused a 25% reduction in kidney levels of active TGF-ß1. Specific upregulation of TG2 in OK proximal tubular epithelial cells increased latent TGF-ß recruitment and activation by 20.7% and 19.7%, respectively, in co-cultures with latent TGF-ß binding protein producing fibroblasts.Conclusions. Regulation of TG2 directly influences the level of active TGF-ß1, and thus, TG inhibition may exert a renoprotective effect by targeting not only a direct extracellular matrix deposition but also TGF-ß1 activation and recruitment.

Potential impact of invasive anuran species in western Newfoundland

Successful dispersal and establishment of invasive anurans (frogs and toads) may be influenced by competitive exclusion and/or niche differentiation with competing species. I investigated the dispersal of anurans in western Newfoundland using anuran calling surveys and pond-edge visual encounter surveys. The Mink Frog, Lithobates septentrionalis, had dispersed ~50 km northeast from the original (2001) discovery location and ~34 km southwest; displaying spatial separation from Green Frogs, Lithobates clamitans, at landscape and local scales. Visual encounter surveys did not reveal any correlation between adult Mink Frogs and odonate competitors. Additionally, I assessed the impact of varying tadpole densities on removal of epilithic periphyton by providing epilithon covered substrates for American Toad, Anaxyrus americanus, tadpoles raised in laboratory or field enclosures. Higher tadpole densities resulted in smaller tadpoles that removed more periphyton from substrates. As anuran population ranges expand, there may be effects on ecological resources for vertebrate and invertebrate competitors.