SETD2 haploinsufficiency for microtubule methylation is an early driver of genomic instability in renal cell carcinoma

Loss of the short arm of chromosome 3 (3p) occurs early in >95% of clear cell renal cell carcinoma (ccRCC). Nearly ubiquitous 3p loss in ccRCC suggests haploinsufficiency for 3p tumor suppressors as early drivers of tumorigenesis. We previously reported methyltransferase SETD2, which trimethylates H3 histones on lysine 36 (H3K36me3) and is located in the 3p deletion, to also trimethylate microtubules on lysine 40 (aTubK40me3) during mitosis, with aTubK40me3 required for genomic stability. We now show that monoallelic, Setd2-deficient cells retaining H3K36me3, but not aTubK40me3, exhibit a dramatic increase in mitotic defects and micronuclei count, with increased viability compared with biallelic loss. In SETD2-inactivated human kidney cells, rescue with a pathogenic SETD2 mutant deficient for microtubule (aTubK40me3), but not histone (H3K36me3) methylation, replicated this phenotype. Genomic instability (micronuclei) was also a hallmark of patient-derived cells from ccRCC. These data show that the SETD2 tumor suppressor displays a haploinsufficiency phenotype disproportionately impacting microtubule methylation and serves as an early driver of genomic instability. Significance: Loss of a single allele of a chromatin modifier plays a role in promoting oncogenesis, underscoring the growing relevance of tumor suppressor haploinsufficiency in tumorigenesis

Small, Dense Quark Stars from Perturbative QCD

As a model for nonideal behavior in the equation of state of QCD at high density, we consider cold quark matter in perturbation theory. To second order in the strong coupling constant, $\alpha_s$, the results depend sensitively on the choice of the renormalization mass scale. Certain choices of this scale correspond to a strongly first order chiral transition, and generate quark stars with maximum masses and radii approximately half that of ordinary neutron stars. At the center of these stars, quarks are essentially massless.Comment: ReVTeX, 5 pages, 3 figure

Direct Evidence for a Critical Role of Myosin II in Budding Yeast Cytokinesis and the Evolvability of New Cytokinetic Mechanisms in the Absence of Myosin II

In the budding yeast Saccharomyces cerevisiae, an actomyosin-based contractile ring is present during cytokinesis, as occurs in animal cells. However, the precise requirement for this structure during budding yeast cytokinesis has been controversial. Here we show that deletion of MYO1, the single myosin II gene, is lethal in a commonly used strain background. The terminal phenotype of myo1Δ is interconnected chains of cells, suggestive of a cytokinesis defect. To further investigate the role of Myo1p in cytokinesis, we conditionally disrupted Myo1 function by using either a dominant negative Myo1p construct or a strain where expression of Myo1p can be shut-off. Both ways of disruption of Myo1 function result in a failure in cytokinesis. Additionally, we show that a myo1Δ strain previously reported to grow nearly as well as the wild type contains a single genetic suppressor that alleviates the severe cytokinesis defects of myo1Δ. Using fluorescence time-lapse imaging and electron microscopy techniques, we show that cytokinesis in this strain is achieved through formation of multiple aberrant septa. Taken together, these results strongly suggest that the actomyosin ring is crucial for successful cytokinesis in budding yeast, but new cytokinetic mechanisms can evolve through genetic changes when myosin II function is impaired

Anticancer properties of an important drug lead podophyllotoxin can be efficiently mimicked by diverse heterocyclic scaffolds accessible via one-step synthesis

Structural simplification of an antimitotic natural product podophyllotoxin with mimetic heterocyclic scaffolds constructed using multicomponent reactions led to the identification of compounds exhibiting low nanomolar antiproliferative and apoptosis-inducing properties. The most potent compounds were found in the dihydropyridopyrazole, dihydropyridonaphthalene, dihydropyridoindole, and dihydropyridopyrimidine scaffold series. Biochemical mechanistic studies performed with dihydropyridopyrazole compounds showed that these heterocycles inhibit in vitro tubulin polymerization and disrupt the formation of mitotic spindles in dividing cells at low nanomolar concentrations, in a manner similar to podophyllotoxin itself. Separation of a racemic dihydropyridonaphthalene into individual enantiomers demonstrated that only the optical antipode matching the absolute configuration of podophyllotoxin possessed potent anticancer activity. Computer modeling, performed using the podophyllotoxin binding site on β-tubulin, provided a theoretical understanding of these successful experimental findings. © 2011 American Chemical Society.Articl

PCR screening and allele frequency estimation of bovine leukocyte adhesion deficiency in Holstein and Gir cattle in Brazil

Bovine leukocyte adhesion deficiency (BLAD) is a disease known to affect the Holstein cattle breed throughout the world. Eighty-eight Holstein dairy cows and 88 Gir dairy bulls were genotyped by PCR for the CD18 BLAD alelle. The frequency of the BLAD mutant allele and the BLAD-carrier prevalence in Brazilian Holstein cows were 2.8 and 5.7%, respectively. No mutant allele was found in any of the 88 Gir animals. We conclude that the CD18 gene mutation is prevalent in Brazilian Holstein cattle and absent or present at a very low frequency in Gir cattle.<br>Oitenta e oito vacas da raça Holandesa e 88 touros da raça Gir foram genotipados através da PCR para o gene CD18 da deficiência de adesão de leucócitos em bovinos (BLAD). As freqüências do alelo mutante BLAD e de vacas heterozigotas da raça Holandesa foram 2,8 e 5,7%, respectivamente. Por outro lado, todos animais Gir foram identificados como homozigotos normais, ou seja, nenhum alelo mutante BLAD foi encontrado. Estes resultados sugerem que a mutação no gene CD18 é prevalente em bovinos brasileiros da raça Holandesa e ausente ou presente numa freqüência muito baixa em animais Gir

Exploring natural product chemistry and biology with multicomponent reactions. 5. Discovery of a novel tubulin-targeting scaffold derived from the rigidin family of marine alkaloids

We developed synthetic chemistry to access the marine alkaloid rigidins and over 40 synthetic analogues based on the 7-deazaxanthine, 7-deazaadenine, 7-deazapurine, and 7-deazahypoxanthine skeletons. Analogues based on the 7-deazahypoxanthine skeleton exhibited nanomolar potencies against cell lines representing cancers with dismal prognoses, tumor metastases, and multidrug resistant cells. Studies aimed at elucidating the mode(s) of action of the 7-deazahypoxanthines in cancer cells revealed that they inhibited in vitro tubulin polymerization and disorganized microtubules in live HeLa cells. Experiments evaluating the effects of the 7-deazahypoxanthines on the binding of [3H]colchicine to tubulin identified the colchicine site on tubulin as the most likely target for these compounds in cancer cells. Because many microtubule-targeting compounds are successfully used to fight cancer in the clinic, we believe the new chemical class of antitubulin agents represented by the 7-deazahypoxanthine rigidin analogues have significant potential as new anticancer agents. © 2013 American Chemical Society.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Interactions of Elongation Factor 1α with F-Actin and β-Actin mRNA: Implications for Anchoring mRNA in Cell Protrusions

The targeting of mRNA and local protein synthesis is important for the generation and maintenance of cell polarity. As part of the translational machinery as well as an actin/microtubule-binding protein, elongation factor 1α (EF1α) is a candidate linker between the protein translation apparatus and the cytoskeleton. We demonstrate in this work that EF1α colocalizes with β-actin mRNA and F-actin in protrusions of chicken embryo fibroblasts and binds directly to F-actin and β-actin mRNA simultaneously in vitro in actin cosedimentation and enzyme-linked immunosorbent assays. To investigate the role of EF1α in mRNA targeting, we mapped the two actin-binding sites on EF1α at high resolution and defined one site at the N-terminal 49 residues of domain I and the other at the C-terminal 54 residues of domain III. In vitro actin-binding assays and localization in vivo of recombinant full-length EF1α and its various truncates demonstrated that the C terminus of domain III was the dominant actin-binding site both in vitro and in vivo. We propose that the EF1α–F-actin complex is the scaffold that is important for β-actin mRNA anchoring. Disruption of this complex would lead to delocalization of the mRNA. This hypothesis was tested by using two dominant negative polypeptides: the actin-binding domain III of EF1α and the EF1α-binding site of yeast Bni1p, a protein that inhibits EF1α binding to F-actin and also is required for yeast mRNA localization. We demonstrate that either domain III of EF1α or the EF1α-binding site of Bni1p inhibits EF1α binding to β-actin mRNA in vitro and causes delocalization of β-actin mRNA in chicken embryo fibroblasts. Taken together, these results implicate EF1α in the anchoring of β-actin mRNA to the protrusion in crawling cells