Bayesian Graphical Regression

<p>We consider the problem of modeling conditional independence structures in heterogenous data in the presence of additional subject-level covariates—termed graphical regression. We propose a novel specification of a conditional (in)dependence function of covariates—which allows the structure of a directed graph to vary flexibly with the covariates; imposes sparsity in both edge and covariate selection; produces both subject-specific and predictive graphs; and is computationally tractable. We provide theoretical justifications of our modeling endeavor, in terms of graphical model selection consistency. We demonstrate the performance of our method through rigorous simulation studies. We illustrate our approach in a cancer genomics-based precision medicine paradigm, where-in we explore gene regulatory networks in multiple myeloma taking prognostic clinical factors into account to obtain both population-level and subject-level gene regulatory networks. Supplementary materials for this article are available online.</p

Characterizing the chloroplast genome of <i>Mammillaria elongata</i> DC. 1828 in the Cactaceae family and unveiling its phylogenetic affinities within the genus <i>Mammillaria</i>

With its nearly 200 species, the Mammillaria genus is the most species-rich within the Cactaceae family, yet surprisingly, few of its chloroplast genomes have been studied. We focused on the species Mammillaria elongata DC. 1828, a petite cactus native to Mexico and favored by horticulturists, yet whose phylogenetic relationships remain uncertain due to a lack of genomic data. We extracted the DNA from a sample obtained in China, sequenced it using the NovaSeq 6000 platform, and assembled the chloroplast genome using GetOrganelle software. Our assembly resulted in a chloroplast genome of 110,981 base pairs with an overall GC content of 36.28%, which included 100 genes (95 unique). Notably, several protein-coding genes were absent. Phylogenetic analysis using 59 shared genes across nine Mammillaria species and one Obregonia species revealed that M. elongata and M. gracilis are closely related, suggesting a recent common ancestor and possible shared evolutionary pressures or ecological niches. This study provides crucial genomic data for M. elongata and hints at intriguing phylogenetic relationships within the Mammillaria genus.</p

PKD1 Mediates Negative Feedback of PI3K/Akt Activation in Response to G Protein-Coupled Receptors

<div><p>We examined whether protein kinase D1 (PKD1) mediates negative feeback of PI3K/Akt signaling in intestinal epithelial cells stimulated with G protein-coupled receptor (GPCR) agonists. Exposure of intestinal epithelial IEC-18 cells to increasing concentrations of the PKD family inhibitor kb NB 142­70, at concentrations that inhibited PKD1 activation, strikingly potentiated Akt phosphorylation at Thr³⁰⁸ and Ser⁴⁷³ in response to the mitogenic GPCR agonist angiotensin II (ANG II). Enhancement of Akt activation by kb NB 142-70 was also evident in cells with other GPCR agonists, including vasopressin and lysophosphatidic acid. Cell treatment rovincial Hospital Affiliated to Shandong University, Jinan, China with the structurally unrelated PKD family inhibitor CRT0066101 increased Akt phosphorylation as potently as kb NB 142–70. Knockdown of PKD1 with two different siRNAs strikingly enhanced Akt phosphorylation in response to ANG II stimulation in IEC-18 cells. To determine whether treatment with kb NB 142–70 enhances accumulation of phosphatidylinositol (3,4,5)-trisphosphate (PIP₃) in the plasma membrane, we monitored the redistribution of Akt-pleckstrin homology domain-green fluorescent protein (Akt-PH-GFP) in single IEC-18 cells. Exposure to kb NB 142–70 strikingly increased membrane accumulation of Akt-PH-GFP in response to ANG II. The translocation of the PIP₃ sensor to the plasma membrane and the phosphorylation of Akt was completed prevented by prior exposure to the class I p110α specific inhibitor A66. ANG II markedly increased the phosphorylation of p85α detected by a PKD motif-specific antibody and enhanced the association of p85α with PTEN. Transgenic mice overexpressing PKD1 showed a reduced phosphorylation of Akt at Ser⁴⁷³ in intestinal epithelial cells compared to wild type littermates. Collectively these results indicate that PKD1 activation mediates feedback inhibition of PI3K/Akt signaling in intestinal epithelial cells <i>in vitro</i> and <i>in vivo</i>.</p></div

PKD1 inhibition potentiates PI3K-mediated production of PIP₃ in response to angiotensin II stimulation.

<p>IEC-18 cells were transiently transfected with a plasmid encoding a fusion protein between GFP and the PH domain of Akt (Akt-PH-GFP). The cultures were incubated in the in the absence (−) or presence of either 3.5 µM kb NB 142–70 (kb) or 3.5 µM kb NB 142–70 and 10 µM A66 (kb + A66) in DMEM containing 10 mM HEPES for 1 h prior to stimulation with 50 nM angiotensin II (ANG II). The intracellular distribution of Akt-PH-GFP was monitored under a fluorescence microscope, as described in <i>Materials and Methods</i>. The selected cells, after 1 h of stimulation with angiotensin II, displayed in the figures were representative of 90% of the population of positive cells. The bars represent the fluorescence ratio (membrane/cytosol) mean ± S.E, n =  at least 20, *p<05.</p

Mechanism of PKD1-mediated negative feedback of PIP₃/Akt in IEC-18 cells and role of PKD1 in the regulation of Akt phosphorylation <i>in vivo</i>.

<p><b>Panel A, <i>PKD1-mediated p85α phosphorylation</i></b> Confluent cultures of IEC-18 cells were incubated in the absence (−) or presence (kb) of either 3.5 µM kb NB 142–70 or 5 μM CRT0066101 (CRT) for 1 h prior to stimulation of the cells with 50 nM angiotensin II (ANGII) for 1 h. Cells were lysed and p85α immunoprecipitated as described in Materials and Methods. Immunoblotting was peformed using the PKD substrate motif antibody (Cell Signling Technology). Fold increases in p85α phosphorylation was quantified using Multi Gauge V3.0 and plotted as bars, mean ± S.E; n = 4. <b>Panel B, </b><b><i>p85α complex formation with EGFR</i></b><i>.</i> Confluent cultures of IEC-18 cells were incubated in the absence (−) or presence (kb) of either 3.5 µM kb NB 142–70 or 1 μM AG1478 for 1 h prior to stimulation of the cells with 50 nM angiotensin II (ANGII) for either 30 or 60 min, as indicated. Cells were lysed and p85α immunoprecipitates were analyzed by antiphosphotyrosine immunoblotting. <b>Panel C, </b><b><i>p85α complex formation with PTEN</i></b> Confluent cultures of IEC-18 cells were incubated in the absence (−) or presence (kb) of 3.5 µM kb NB 142–70 for 1 h prior to stimulation of the cells with 50 nM angiotensin II (ANGII) for either 30 or 60 min, as indicated. Cells were lysed and p85α immunoprecipitates were analyzed by immunoblotting with PTEN antibodies. Bars stand for the mean ± S.E n = 4. Individual values are the ratio of PTEN band intensity to the corresponding p85α band intensity in each experiment. *p<0.05 as compared with values at time 0 or values obtained in cells treated with kb NB 142–70 and stimulated with ANG II and at each time point. <b>Panel D, </b><b><i>Overexpression of PKD1 reduces Akt phosphorylation in intestinal epithelial cells</i></b><i>.</i> Epithelial cells from the ileum of transgenic (Tg) mice and nontransgenic (NTg) littermates mice were isolated sequentially by timed incubations in a EDTA-PBS solution. Western blot was used to analyze lysates of these cells for Akt phosphorylated at Ser⁴⁷³, total Akt, PKD1 autophosphorylated at Ser⁹¹⁶ and total PKD1 (PKD-C20). Equivalent loading was verified by immunoblotting for tubulin. Results are shown for 2 transgenic mice and 2 nontransgenic littermates. Bars: represent Akt phosphorylated at Ser⁴⁷³ (means ± SE; n = 4). *p<0.05.</p

PKD1 inhibition by increasing concentrations of kb NB 142-70 potentiates PI3K/Akt activation in angiotensin II-stimulated IEC-18 cells.

<p>A, Confluent cultures of IEC-18 cells were incubated in the in the absence (−) or presence (kb) of increasing concentrations of kb NB 142–70 for 1 h prior to stimulation of the cells without (−) or with 50 nM angiotensin II for 1 h, as indicated. All cultures were then lysed with 2×SDS–PAGE sample buffer. The samples were analyzed by SDS-PAGE and immunoblotting with antibodies that detect the phosphorylated state of Akt at Ser⁴⁷³, Akt at Thr³⁰⁸ and total Akt to verify equal gel loading and PKD1 at Ser⁹¹⁶ and Ser⁷⁴⁸<b>B and C</b>, Quantifications were performed by using Multi Gauge V3.0 and plotted as a percentage of the maximum response, mean ± S.E n = 3, induced by angiotensin II and kb NB 142–70 <b>(B)</b> or as percentage of the maximum response, mean ± S.E n = 3, induced by angiotensin II <b>(C)</b>.</p

PKD1 inhibition potentiates PI3K/Akt activation in IEC-18 cells stimulated with vasopressin and LPA.

<p>Confluent cultures of IEC-18 cells were incubated in the in the absence (−) or presence 3.5 µM kb NB 142–70 (kb) for 1 h prior to stimulation of the cells with either 50 nM angiotensin II (ANGII) or 50 nM vasopressin (VP) for 1 h (<b>panel A</b>)<b>,</b> 10 µM LPA for 2.5 or 10 min (<b>panel B</b>)<b>,</b> 50 nM angiotensin II (ANGII) or 50 ng/ml EGF for 1 h (<b>panel C</b>)<b>.</b> All cultures were then lysed with 2× SDS–PAGE sample buffer. The samples were analyzed by SDS-PAGE and immunoblotting with antibodies that detect the phosphorylated state of Akt at Ser⁴⁷³ and Thr³⁰⁸, total Akt to verify equal gel loading and PKD1 at Ser⁹¹⁶ (p-PKD). <b>Panel D,</b> Fold increases following PKD1 inhibition for angiotensin II (ANG, n = 20), vasopressin (VP, n = 4), LPA (n = 4) and EGF (n = 3) in Akt at Thr³⁰⁸ and Ser⁴⁷³ phosphorylation were quantified using Multi Gauge V3.0 and plotted as bars, mean ± S.E.</p

PKD1 mediates feedback inhibition of PI3K/Akt activation in IEC-18 cells stimulated with angiotensin II.

<p><b>Panel A,</b> Confluent cultures of IEC-18 cells were incubated in the absence (−) or presence (kb) of either 3.5 µM kb NB 142–70 or 5 μM CRT0066101 (CRT) for 1 h prior to stimulation of the cells with 50 nM angiotensin II (ANGII) for 1 h. All cultures were then lysed with 2× SDS–PAGE sample buffer. The samples were analyzed by SDS-PAGE and immunoblotting with antibodies that detect the phosphorylated state of Akt at Ser⁴⁷³ and Thr³⁰⁸ and total Akt to verify equal gel loading. PKD1 phosphorylated at Ser⁹¹⁶ was also determined. <b>Panels </b><b>B,</b> Cultures of IEC-18 cells were transfected with non-targeting siRNA (N Targ) or with two different siRNAs targeting PKD1, siPKD1 (1) and siPKD1 (2). Other cultures were not subjected to transfection (Cont). Then, the cultures were stimulated with 50 nM angiotensin II (ANGII) for 10 min and lysed with 2× SDS–PAGE sample buffer. The samples were analyzed by SDS-PAGE and immunoblotting with antibodies that detect the phosphorylated state of Akt at Ser⁴⁷³ and Thr³⁰⁸, total Akt to verify equal gel loading and total PKD1 (PKD-C20) to evaluate siRNA-mediated knockdown of PKD1 expression. Similar results were obtained in at least 2 independent experiments in each case.</p

Copper-Catalyzed Regioselective Intramolecular Electrophilic Sulfenoamination via Lewis Acid Activation of Disulfides under Aerobic Conditions

The activation of disulfides by Cu­(II) salts has been realized, which triggers a highly efficient electrophilic sulfenoamination of alkenes under aerobic conditions. Various sulfenyl N-heterocycles and their Selena counterparts were produced regioselectively, with no competing disulfidation products detected. Mechanistic studies suggest a profound influence of the counterions on the Lewis acidic copper center, and the important roles of oxygen and DMSO as co-oxidants for these cyclization processes