Functional Analysis of Yeast Pheromone Receptors in ER Exit, Ligand-Induced Endocytosis and Oligomerization: A Dissertation

This study investigates endocytosis and ER export signals of the yeast α-factor receptor and the role that receptor oligomerization plays in these processes. The α-factor receptor contains signal sequences in the cytoplasmic C-terminal domain that are essential for ligand-mediated endocytosis. In an endocytosis complementation assay, I found that oligomeric complexes of the receptor undergo ligand-mediated endocytosis when the α-factor binding site and the endocytosis signal sequences are located in different receptors. Both in vitro and in vivo assays strongly suggested that ligand-induced conformational changes in one Ste2 subunit do not affect neighboring subunits. Therefore, the recognition of endocytosis signal sequence and the recognition of the ligand-induced conformational change are likely to be two independent events, where the signal sequence plays only a passive role in the ligand-induced endocytosis. Four amino acid substitutions (C59R, H94P, S141P and S145P) in TM domains I, II and III were identified that resulted in the accumulation of truncated receptors in the ER but did not block ER export of full-length receptors. The two DXE motifs in the C-terminal tail were required for export of the mutant receptors from the ER; however DXE was not essential for proper cell surface expression of wild-type receptors apparently because the receptors contain redundant ER export signals. An assay for oligomerization of receptors in the ER was developed based on the ability of truncated mutant receptors to exit the ER. The four substitutions (C59R, H94P, S141P and S145P) that caused DXE-dependent ER export failed to form homo-oligomers, suggesting that the DXE motifs and receptor oligomerization serve as independent ER export signals. Consistent with this view, two of the substitutions (S141P and S145P), when coexpressed, with wild-type receptors, formed hetero-oligomers that exited the ER. Finally, the full-length oligomer-defective mutant Ste2-S141P was sensitive to α-factor, suggesting that receptor monomers that reach the cell surface are able to activate the heterotrimeric G protein. The potential roles that TM1, 2 and 3 play in receptor oligomerization are discussed

Shape restricted regression with random Bernstein polynomials

Shape restricted regressions, including isotonic regression and concave regression as special cases, are studied using priors on Bernstein polynomials and Markov chain Monte Carlo methods. These priors have large supports, select only smooth functions, can easily incorporate geometric information into the prior, and can be generated without computational difficulty. Algorithms generating priors and posteriors are proposed, and simulation studies are conducted to illustrate the performance of this approach. Comparisons with the density-regression method of Dette et al. (2006) are included.Comment: Published at http://dx.doi.org/10.1214/074921707000000157 in the IMS Lecture Notes Monograph Series (http://www.imstat.org/publications/lecnotes.htm) by the Institute of Mathematical Statistics (http://www.imstat.org

Missing link in community psychiatry: When a patient with schizophrenia was expelled from her home

Treatment and disposition of homeless patients with schizophrenia represent a great challenge in clinical practice. We report a case of this special population, and discuss the development of homelessness, the difficulty in disposition, their utilization of health services, and possible applications of mandatory community treatment in this group of patients. A 51-year-old homeless female was brought to an emergency department for left femur fracture caused by an assault. She was diagnosed with schizophrenia about 20 years ago but received little help from mental health services over the decades. During hospitalization, her psychotic symptoms were only partially responsive to treatment. Her family refused to handle caretaking duties. The social welfare system was mobilized for long-term disposition. Homeless patients with schizophrenia are characterized by family disruption, poor adherence to health care, and multiple emergency visits and hospitalization. We hope this article can provide information about the current mental health policy to medical personnel. It is possible that earlier intervention and better outcome can be achieved by utilizing mandatory community treatment in the future, as well as preventing patients with schizophrenia from losing shelters

Profiling time course expression of virus genes---an illustration of Bayesian inference under shape restrictions

There have been several studies of the genome-wide temporal transcriptional program of viruses, based on microarray experiments, which are generally useful in the construction of gene regulation network. It seems that biological interpretations in these studies are directly based on the normalized data and some crude statistics, which provide rough estimates of limited features of the profile and may incur biases. This paper introduces a hierarchical Bayesian shape restricted regression method for making inference on the time course expression of virus genes. Estimates of many salient features of the expression profile like onset time, inflection point, maximum value, time to maximum value, area under curve, etc. can be obtained immediately by this method. Applying this method to a baculovirus microarray time course expression data set, we indicate that many biological questions can be formulated quantitatively and we are able to offer insights into the baculovirus biology.Comment: Published in at http://dx.doi.org/10.1214/09-AOAS258 the Annals of Applied Statistics (http://www.imstat.org/aoas/) by the Institute of Mathematical Statistics (http://www.imstat.org

Intramuscular electroporation with the pro-opiomelanocortin gene in rat adjuvant arthritis

Endogenous opioid peptides have an essential role in the intrinsic modulation and control of inflammatory pain, which could be therapeutically useful. In this study, we established a muscular electroporation method for the gene transfer of pro-opiomelanocortin (POMC) in vivo and investigated its effect on inflammatory pain in a rat model of rheumatoid arthritis. The gene encoding human POMC was inserted into a modified pCMV plasmid, and 0–200 μg of the plasmid-POMC DNA construct was transferred into the tibialis anterior muscle of rats treated with complete Freund's adjuvant (CFA) with or without POMC gene transfer by the electroporation method. The safety and efficiency of the gene transfer was assessed with the following parameters: thermal hyperalgesia, serum adrenocorticotropic hormone (ACTH) and endorphin levels, paw swelling and muscle endorphin levels at 1, 2 and 3 weeks after electroporation. Serum ACTH and endorphin levels of the group into which the gene encoding POMC had been transferred were increased to about 13–14-fold those of the normal control. These levels peaked 1 week after electroporation and significantly decreased 2 weeks after electroporation. Rats that had received the gene encoding POMC had less thermal hypersensitivity and paw swelling than the non-gene-transferred group at days 3, 5 and 7 after injection with CFA. Our promising results showed that transfer of the gene encoding POMC by electroporation is a new and effective method for its expression in vivo, and the analgesic effects of POMC cDNA with electroporation in a rat model of rheumatoid arthritis are reversed by naloxone

Aciculatin inhibits lipopolysaccharide-mediated inducible nitric oxide synthase and cyclooxygenase-2 expression via suppressing NF-κB and JNK/p38 MAPK activation pathways

<p>Abstract</p> <p>Objectives</p> <p>Natural products have played a significant role in drug discovery and development. Inflammatory mediators such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) have been suggested to connect with various inflammatory diseases. In this study, we explored the anti-inflammatory potential of aciculatin (8-((2<it>R</it>,4<it>S</it>,5<it>S</it>,6<it>R</it>)-tetrahydro-4,5-dihydroxy-6-methyl-2<it>H</it>-pyran-2-yl)-5-hydroxy-2-(4-hydroxyphenyl)-7-methoxy-4<it>H</it>-chromen-4-one), one of main components of <it>Chrysopogon aciculatis</it>, by examining its effects on the expression and activity of iNOS and COX-2 in lipopolysaccharide (LPS)-activated macrophages.</p> <p>Methods</p> <p>We used nitrate and prostaglandin E₂(PGE₂) assays to examine inhibitory effect of aciculatin on nitric oxide (NO) and PGE₂levels in LPS-activated mouse RAW264.7 macrophages and further investigated the mechanisms of aciculatin suppressed LPS-mediated iNOS/COX-2 expression by western blot, RT-PCR, reporter gene assay and confocal microscope analysis.</p> <p>Results</p> <p>Aciculatin remarkably decreased the LPS (1 μg/mL)-induced mRNA and protein expression of iNOS and COX-2 as well as their downstream products, NO and PGE₂respectively, in a concentration-dependent manner (1-10 μM). Such inhibition was found, via immunoblot analyses, reporter gene assays, and confocal microscope observations that aciculatin not only acts through significant suppression of LPS-induced NF-κB activation, an effect highly correlated with its inhibitory effect on LPS-induced IκB kinase (IKK) activation, IκB degradation, NF-κB phosphorylation, nuclear translocation and binding of NF-κB to the κB motif of the iNOS and COX-2 promoters, but also suppressed phosphorylation of JNK/p38 mitogen-activated protein kinases (MAPKs).</p> <p>Conclusion</p> <p>Our results demonstrated that aciculatin exerts potent anti-inflammatory activity through its dual inhibitory effects on iNOS and COX-2 by regulating NF-κB and JNK/p38 MAPK pathways.</p

Effect of acetic acid on telomerase activity in premalignant and malignant cervical lesions

PubMe