Characterization of basal and estrogen-regulated antisense transcription in breast cancer cells: Role in regulating sense transcription.

Estrogen-responsive breast cancer cells exhibit both basal and estrogen-regulated transcriptional programs, which lead to the transcription of many different transcription units (i.e., genes), including those that produce coding and non-coding sense (e.g., mRNA, lncRNA) and antisense (i.e., asRNA) transcripts. We have previously characterized the global basal and estrogen-regulated transcriptomes in estrogen receptor alpha (ERα)-positive MCF-7 breast cancer cells. Herein, we have mined genomic data to define three classes of antisense transcription in MCF-7 cells based on where their antisense transcription termination sites reside relative to their cognate sense mRNA and lncRNA genes. These three classes differ in their response to estrogen treatment, the enrichment of a number of genomic features associated with active promoters (H3K4me3, RNA polymerase II, open chromatin architecture), and the biological functions of their cognate sense genes as analyzed by DAVID gene ontology. We further characterized two estrogen-regulated antisense transcripts arising from the MYC gene in MCF-7 cells, showing that these antisense transcripts are 5'-capped, 3'-polyadenylated, and localized to different compartments of the cell. Together, our analyses have revealed distinct classes of antisense transcription correlated to different biological processes and response to estrogen stimulation, uncovering another layer of hormone-regulated gene regulation.

Emerging Therapies for Inflammatory Bowel Diseases.

BACKGROUND: The past decade has seen important advances in the management of chronic inflammatory bowel diseases (IBD), consisting of Crohn's disease (CD) and ulcerative colitis. The development of TNF antagonists, the recognition of interrupting lymphocyte trafficking as an effective treatment strategy, confirmation of the value of combination therapy, and the need, particularly in CD, for the treatment of high-risk patients early in the disease course are all fundamental concepts upon which the next generation of IBD treatment algorithms will be built. Emerging concepts that will continue to evolve and shape the field include an increased emphasis on personalized medicine (right drug, right dose, right time) and the development of new therapeutic classes. In this article, we review the clinical data and provide some insights into recent data regarding IBD therapies. KEY MESSAGES: In this article, we review the mechanism of action and data for novel therapies in IBD with particular focus on the evidence for agents targeting leukocyte trafficking, cytokine signaling, including interleukin-12/23 and the Janus kinase-signal transducers/activators of transcription pathway, and the emergence of antisense therapy for the treatment of IBD. CONCLUSIONS: Multiple new therapies are emerging for IBD; however, the potential positioning of these agents in treatment algorithms is difficult to predict in the absence of comparative effectiveness studies.

Glutathione decreases in dopaminergic PC12 cells interfere with the ubiquitin protein degradation pathway: relevance for Parkinson's disease?

Parkinson's disease (PD) is characterized by the presence of proteinaceous neuronal inclusions called Lewy bodies in susceptible dopaminergic midbrain neurons. Inhibition of the ubiquitin-proteasome protein degradation pathway may contribute to protein build-up and subsequent cell death. Ubiquitin is normally activated for transfer to substrate proteins by interaction with the E1 ubiquitin ligase enzyme via a thiol ester bond. Parkinson's disease is also characterized by decreases in midbrain levels of total glutathione which could impact on E1 enzyme activity via oxidation of the active site sulfhydryl. We have demonstrated that increasing reductions in total glutathione in dopaminergic PC12 cells results in corresponding decreases in ubiquitin-protein conjugate levels suggesting that ubiquitination of proteins is inhibited in a glutathione-dependent fashion. Decreased ubiquitinated protein levels appears to be due to inhibition of E1 activity as demonstrated by reductions in endogenous E1-ubiquitin conjugate levels as well as decreases in the production of de novo E1-ubiquitin conjugates when glutathione is depleted. This is a reversible process as E1 activity increases upon glutathione restoration. Our data suggests that decreases in cellular glutathione in dopaminergic cells results in decreased E1 activity and subsequent disruption of the ubiquitin pathway. This may have implications for neuronal degeneration in PD.

Antisense imaging: where are we now?

The field of antisense targeting is changing rapidly as additional results from in vitro studies and animal and patient trials become available. While these developments apply primarily to antisense chemotherapy, many have implications for antisense imaging and radiotherapy. It may now be profitable to reconsider antisense imaging in the light of these recent developments. With the benefit of further insight, it may be possible to predict which antisense mechanisms will be preferable for antisense imaging. It is also possible to consider the influences of carriers (vectors) on the targeting of antisense DNA and whether this might improve imaging. Furthermore, estimates showing only low mRNA steady-state copy numbers per cell may be reconsidered in refining predictions of tissue counting rates. Finally, recent results suggest that radiolabeling antisense DNAs may not adversely influence the targeting properties of antisense DNAs.

Expression of the 25-kDa heat-shock protein (HSP27) correlates with resistance to the toxicity of cadmium chloride, mercuric chloride, cis-platinum(II)-diammine dichloride, or sodium arsenite in mouse embryonic stem cells transfected with sense or antisense HSP27 cDNA.

Embryonic stem (ES) cells were transfected with the protein-coding region of rat HSP27 cDNA placed in sense or antisense orientation in vector pcDNA3 under the control of the constitutive cytomegalovirus (CMV) promoter. Compared with nontransfected ES cells, five sense HSP27 cDNA-transfected ES cell clones displayed up to fourfold increases in expression of HSP27 mRNA and up to sixfold increases in expression of HSP27 protein, whereas four antisense HSP27 cDNA-transfected ES cell lines exhibited synthesis of antisense HSP27 RNA and a 50-85% decrease in HSP27 protein expression. Compared to the parental ES cell lines or ES cells transfected with the vector lacking any HSP27 sequence, all ES cell lines overexpressing HSP27 were resistant to killing by cadmium chloride (CdCl2), mercuric chloride (HgCl2), cis-platinum(II)-diammine dichloride (cDDP), sodium arsenite (NaAsO2), and heat while ES cell lines expressing reduced HSP27 were more sensitive to metal toxicity and heat. The relative toxicities of the tested metals to ES cells were cDDP > NaAsO2 > HgCl2 > CdCl2. Protection of ES cells against metal or heat toxicity was positively correlated with the level of HSP27. These data confirm in ES cells previous reports of the ability of HSP27 to protect other cell types against heat and demonstrate that HSP27 protects mammalian cells against the toxic effects of diverse metals.