Influence of RARα gene on MDR1 expression and P-glycoprotein function in human leukemic cells

BACKGROUND: Multidrug resistance (MDR) phenotype of malignant cells is the major problem in the chemotherapy of neoplasia. The treatment of leukemia with retinoids is aimed on the induction of leukemic cells differentiation. However the interconnections between retinoid regulated differentiation of leukemic cells and regulation of MDR remains unclear. METHODS: Four lines of cultured leukemic cells of diverse types of differentiation were infected with RARα gene and stable transfectants were isolated. We investigated the differentiation of these cells as well as the expression of RARα and MDR1 genes and P-glycoprotein (Pgp, MDR protein) functional activity in these cells. RESULTS: All RARα transfected sublines demonstrated the increase in the quantity of RARα mRNA. All these sublines became more differentiated. Intrinsic activity of MDR1 gene (but not Pgp functional activity) was increased in one of the transfectants. All-trans-retinoic acid (ATRA) induced Pgp activity in two of three infectants to a larger extent than in parental cells. CONCLUSION: The data show that RARα regulates MDR1/ Pgp activity in human leukemic cells, in the first place, Pgp activity induced by ATRA. These results show that RARα overexpression in leukemic cells could result in MDR

Evolutionary analysis across mammals reveals distinct classes of long non-coding RNAs

Background: Recent advances in transcriptome sequencing have enabled the discovery of thousands of long non-coding RNAs (lncRNAs) across many species. Though several lncRNAs have been shown to play important roles in diverse biological processes, the functions and mechanisms of most lncRNAs remain unknown. Two significant obstacles lie between transcriptome sequencing and functional characterization of lncRNAs: identifying truly non-coding genes from de novo reconstructed transcriptomes, and prioritizing the hundreds of resulting putative lncRNAs for downstream experimental interrogation. Results: We present slncky, a lncRNA discovery tool that produces a high-quality set of lncRNAs from RNA-sequencing data and further uses evolutionary constraint to prioritize lncRNAs that are likely to be functionally important. Our automated filtering pipeline is comparable to manual curation efforts and more sensitive than previously published computational approaches. Furthermore, we developed a sensitive alignment pipeline for aligning lncRNA loci and propose new evolutionary metrics relevant for analyzing sequence and transcript evolution. Our analysis reveals that evolutionary selection acts in several distinct patterns, and uncovers two notable classes of intergenic lncRNAs: one showing strong purifying selection on RNA sequence and another where constraint is restricted to the regulation but not the sequence of the transcript. Conclusion: Our results highlight that lncRNAs are not a homogenous class of molecules but rather a mixture of multiple functional classes with distinct biological mechanism and/or roles. Our novel comparative methods for lncRNAs reveals 233 constrained lncRNAs out of tens of thousands of currently annotated transcripts, which we make available through the slncky Evolution Browser

Defining the 5 and 3 landscape of the Drosophila transcriptome with Exo-seq and RNaseH-seq

Cells regulate biological responses in part through changes in transcription start sites (TSS) or cleavage and polyadenylation sites (PAS). To fully understand gene regulatory networks, it is therefore critical to accurately annotate cell type-specific TSS and PAS. Here we present a simple and straightforward approach for genome-wide annotation of 5- and 3-RNA ends. Our approach reliably discerns bona fide PAS from false PAS that arise due to internal poly(A) tracts, a common problem with current PAS annotation methods. We applied our methodology to study the impact of temperature on the Drosophila melanogaster head transcriptome. We found hundreds of previously unidentified TSS and PAS which revealed two interesting phenomena: first, genes with multiple PASs tend to harbor a motif near the most proximal PAS, which likely represents a new cleavage and polyadenylation signal. Second, motif analysis of promoters of genes affected by temperature suggested that boundary element association factor of 32 kDa (BEAF-32) and DREF mediates a transcriptional program at warm temperatures, a result we validated in a fly line where beaf-32 is downregulated. These results demonstrate the utility of a high-throughput platform for complete experimental and computational analysis of mRNA-ends to improve gene annotation

Evolutionary analysis across mammals reveals distinct classes of long non-coding RNAs

BACKGROUND: Recent advances in transcriptome sequencing have enabled the discovery of thousands of long non-coding RNAs (lncRNAs) across many species. Though several lncRNAs have been shown to play important roles in diverse biological processes, the functions and mechanisms of most lncRNAs remain unknown. Two significant obstacles lie between transcriptome sequencing and functional characterization of lncRNAs: identifying truly non-coding genes from de novo reconstructed transcriptomes, and prioritizing the hundreds of resulting putative lncRNAs for downstream experimental interrogation. RESULTS: We present slncky, a lncRNA discovery tool that produces a high-quality set of lncRNAs from RNA-sequencing data and further uses evolutionary constraint to prioritize lncRNAs that are likely to be functionally important. Our automated filtering pipeline is comparable to manual curation efforts and more sensitive than previously published computational approaches. Furthermore, we developed a sensitive alignment pipeline for aligning lncRNA loci and propose new evolutionary metrics relevant for analyzing sequence and transcript evolution. Our analysis reveals that evolutionary selection acts in several distinct patterns, and uncovers two notable classes of intergenic lncRNAs: one showing strong purifying selection on RNA sequence and another where constraint is restricted to the regulation but not the sequence of the transcript. CONCLUSION: Our results highlight that lncRNAs are not a homogenous class of molecules but rather a mixture of multiple functional classes with distinct biological mechanism and/or roles. Our novel comparative methods for lncRNAs reveals 233 constrained lncRNAs out of tens of thousands of currently annotated transcripts, which we make available through the slncky Evolution Browser

Revealing Josephson vortex dynamics in proximity junctions below critical current

Made of a thin non-superconducting metal (N) sandwiched by two superconductors (S), SNS Josephson junctions enable novel quantum functionalities by mixing up the intrinsic electronic properties of N with the superconducting correlations induced from S by proximity. Electronic properties of these devices are governed by Andreev quasiparticles [1] which are absent in conventional SIS junctions whose insulating barrier (I) between the two S electrodes owns no electronic states. Here we focus on the Josephson vortex (JV) motion inside Nb-Cu-Nb proximity junctions subject to electric currents and magnetic fields. The results of local (Magnetic Force Microscopy) and global (transport) experiments provided simultaneously are compared with our numerical model, revealing the existence of several distinct dynamic regimes of the JV motion. One of them, identified as a fast hysteretic entry/escape below the critical value of Josephson current, is analyzed and suggested for low-dissipative logic and memory elements.Comment: 11 pages, 3 figures, 1 table, 43 reference

A Defective mRNA Cleavage and Polyadenylation Complex Facilitates Expansions of Transcribed (GAA) n Repeats Associated with Friedreich’s Ataxia

Expansions of microsatellite repeats are responsible for numerous hereditary diseases in humans, including myotonic dystrophy and Friedreich's ataxia. Whereas the length of an expandable repeat is the main factor determining disease inheritance, recent data point to genomic trans modifiers that can impact the likelihood of expansions and disease progression. Detection of these modifiers may lead to understanding and treating repeat expansion diseases. Here, we describe a method for the rapid, genome-wide identification of trans modifiers for repeat expansion in a yeast experimental system. Using this method, we found that missense mutations in the endoribonuclease subunit (Ysh1) of the mRNA cleavage and polyadenylation complex dramatically increase the rate of (GAA) n repeat expansions but only when they are actively transcribed. These expansions correlate with slower transcription elongation caused by the ysh1 mutation. These results reveal an interplay between RNA processing and repeat-mediated genome instability, confirming the validity of our approach. Keywords: genome instability; repeat expansion; RNA polyadenylation; RNA processing; transcription-replication conflicts; Friedreich’s ataxia; DNA double-strand breaks; trans-modifiers of repeat expansions; genetic screen; whole-genome sequencin