The Border Between the Ultrabithorax and abdominal-A Regulatory Domains in the Drosophila Bithorax Complex

The bithorax complex in Drosophila melanogaster includes three homeobox-containing genes—Ultrabithorax (Ubx), abdominal-A (abd-A), and Abdominal-B (Abd-B)—which are required for the proper differentiation of the posterior 10 segments of the body. Each of these genes has multiple distinct regulatory regions; there is one for each segmental unit of the body plan where the genes are expressed. One additional protein- coding gene in the bithorax complex, Glut3, a sugar-transporter homolog, can be deleted without phenotype. We focus here on the upstream regulatory region for Ubx, the bithoraxoid (bxd) domain, and its border with the adjacent infraabdominal-2 (iab-2) domain, which controls abdA. These two domains can be defined by the phenotypes of rearrangement breakpoints, and by the expression patterns of enhancer traps. In D. virilis, the homeotic cluster is split between Ubx and abd-A, and so the border can also be located by a sequence comparison between species. When the border region is deleted in melanogaster, the flies show a dominant phenotype called Front-ultraabdominal (Fub); the first abdominal segment is transformed into a copy of the second abdominal segment. Thus, the border blocks the spread of activation from the bxd domain into the iab-2 domain.     

Architectural and Functional Diversity of Polycomb Group Response Elements in Drosophila

Polycomb group response elements (PREs) play an essential role in gene regulation by the Polycomb group (PcG) repressor proteins in Drosophila. PREs are required for the recruitment and maintenance of repression by the PcG proteins. PREs are made up of binding sites for multiple DNA-binding proteins, but it is still unclear what combination(s) of binding sites is required for PRE activity. Here we compare the binding sites and activities of two closely linked yet separable PREs of the Drosophila engrailed (en) gene, PRE1 and PRE2. Both PRE1 and PRE2 contain binding sites for multiple PRE–DNA-binding proteins, but the number, arrangement, and spacing of the sites differs between the two PREs. These differences have functional consequences. Both PRE1 and PRE2 mediate pairing-sensitive silencing of mini-white, a functional assay for PcG repression; however, PRE1 requires two binding sites for Pleiohomeotic (Pho), whereas PRE2 requires only one Pho-binding site for this activity. Furthermore, for full pairing-sensitive silencing activity, PRE1 requires an AT-rich region not found in PRE2. These two PREs behave differently in a PRE embryonic and larval reporter construct inserted at an identical location in the genome. Our data illustrate the diversity of architecture and function of PREs.     

Network Properties of the Ensemble of RNA Structures

We describe the first dynamic programming algorithm that computes the expected degree for the network, or graph G = (V, E) of all secondary structures of a given RNA sequence a = a ₁, …, a _n. Here, the nodes V correspond to all secondary structures of a, while an edge exists between nodes s, t if the secondary structure t can be obtained from s by adding, removing or shifting a base pair. Since secondary structure kinetics programs implement the Gillespie algorithm, which simulates a random walk on the network of secondary structures, the expected network degree may provide a better understanding of kinetics of RNA folding when allowing defect diffusion, helix zippering, and related conformation transformations. We determine the correlation between expected network degree, contact order, conformational entropy, and expected number of native contacts for a benchmarking dataset of RNAs. Source code is available at http://bioinformatics.bc.edu/clotelab/RNAexpNumNbors.     

SPOROS: A pipeline to analyze DISE/6mer seed toxicity

microRNAs (miRNAs) are (18-22nt long) noncoding short (s)RNAs that suppress gene expression by targeting the 3’ untranslated region of target mRNAs. This occurs through the seed sequence located in position 2-7/8 of the miRNA guide strand, once it is loaded into the RNA induced silencing complex (RISC). G-rich 6mer seed sequences can kill cells by targeting C-rich 6mer seed matches located in genes that are critical for cell survival. This results in induction of Death Induced by Survival gene Elimination (DISE), through a mechanism we have called 6mer seed toxicity. miRNAs are often quantified in cells by aligning the reads from small (sm)RNA sequencing to the genome. However, the analysis of any smRNA Seq data set for predicted 6mer seed toxicity requires an alternative workflow, solely based on the exact position 2–7 of any short (s)RNA that can enter the RISC. Therefore, we developed SPOROS, a semi-automated pipeline that produces multiple useful outputs to predict and compare 6mer seed toxicity of cellular sRNAs, regardless of their nature, between different samples. We provide two examples to illustrate the capabilities of SPOROS: Example one involves the analysis of RISC-bound sRNAs in a cancer cell line (either wild-type or two mutant lines unable to produce most miRNAs). Example two is based on a publicly available smRNA Seq data set from postmortem brains (either from normal or Alzheimer’s patients). Our methods (found at https://github.com/ebartom/SPOROS and at Code Ocean: https://doi.org/10.24433/CO.1732496.v1) are designed to be used to analyze a variety of smRNA Seq data in various normal and disease settings.     

Genome sequence of the free-living aerobic spirochete Turneriella parva type strain (HT), and emendation of the species Turneriella parva

Turneriella parva Levett et al. 2005 is the only species of the genus Turneriella which was established as a result of the reclassification of Leptospira parva Hovind-Hougen et al. 1982. Together with Leptonema and Leptospira, Turneriella constitutes the family Leptospiraceae, within the order Spirochaetales. Here we describe the features of this free-living aerobic spirochete together with the complete genome sequence and annotation. This is the first complete genome sequence of a member of the genus Turneriella and the 13^th member of the family Leptospiraceae for which a complete or draft genome sequence is now available. The 4,409,302 bp long genome with its 4,169 protein-coding and 45 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.     

Kinetochore Biorientation in Saccharomyces cerevisiae Requires a Tightly Folded Conformation of the Ndc80 Complex

Accurate transmission of genetic material relies on the coupling of chromosomes to spindle microtubules by kinetochores. These linkages are regulated by the conserved Aurora B/Ipl1 kinase to ensure that sister chromatids are properly attached to spindle microtubules. Kinetochore–microtubule attachments require the essential Ndc80 complex, which contains two globular ends linked by large coiled-coil domains. In this study, we isolated a novel ndc80 mutant in Saccharomyces cerevisiae that contains mutations in the coiled-coil domain. This ndc80 mutant accumulates erroneous kinetochore–microtubule attachments, resulting in misalignment of kinetochores on the mitotic spindle. Genetic analyses with suppressors of the ndc80 mutant and in vitro cross-linking experiments suggest that the kinetochore misalignment in vivo stems from a defect in the ability of the Ndc80 complex to stably fold at a hinge in the coiled coil. Previous studies proposed that the Ndc80 complex can exist in multiple conformations: elongated during metaphase and bent during anaphase. However, the distinct functions of individual conformations in vivo are unknown. Here, our analysis revealed a tightly folded conformation of the Ndc80 complex that is likely required early in mitosis. This conformation is mediated by a direct, intracomplex interaction and involves a greater degree of folding than the bent form of the complex at anaphase. Furthermore, our results suggest that this conformation is functionally important in vivo for efficient error correction by Aurora B/Ipl1 and, consequently, to ensure proper kinetochore alignment early in mitosis.     

High quality draft genome sequence of Olivibacter sitiensis type strain (AW-6T), a diphenol degrader with genes involved in the catechol pathway

Olivibacter sitiensis Ntougias et al. 2007 is a member of the family Sphingobacteriaceae, phylum Bacteroidetes. Members of the genus Olivibacter are phylogenetically diverse and of significant interest. They occur in diverse habitats, such as rhizosphere and contaminated soils, viscous wastes, composts, biofilter clean-up facilities on contaminated sites and cave environments, and they are involved in the degradation of complex and toxic compounds. Here we describe the features of O. sitiensis AW-6^T, together with the permanent-draft genome sequence and annotation. The organism was sequenced under the Genomic Encyclopedia for Bacteria and Archaea (GEBA) project at the DOE Joint Genome Institute and is the first genome sequence of a species within the genus Olivibacter. The genome is 5,053,571 bp long and is comprised of 110 scaffolds with an average GC content of 44.61%. Of the 4,565 genes predicted, 4,501 were protein-coding genes and 64 were RNA genes. Most protein-coding genes (68.52%) were assigned to a putative function. The identification of 2-keto-4-pentenoate hydratase/2-oxohepta-3-ene-1,7-dioic acid hydratase-coding genes indicates involvement of this organism in the catechol catabolic pathway. In addition, genes encoding for β-1,4-xylanases and β-1,4-xylosidases reveal the xylanolytic action of O. sitiensis.     

The small nucleolar ribonucleoprotein (snoRNP) database

Small nucleolar ribonucleoproteins (snoRNPs) are widely studied and characterized as guide RNAs for sequence-specific 2′-O-ribose methylation and psuedouridylation of ribosomal RNAs. In addition, snoRNAs have also been shown to interact with some tRNAs and direct alternative splicing in mRNA biogenesis. Recent advances in bioinformatics have resulted in new algorithms able to rapidly identify noncoding RNAs generally and snoRNAs specifically in genomic and metagenomic sequences, resulting in a rapid increase in the number and diversity of identified snoRNA sequences. The snoRNP database is a web-based collection of snoRNA and snoRNA-associated protein sequences from a wide range of species. The database currently contains 8994 snoRNA sequences from Bacteria, Archaea, and Eukaryotes and 589 snoRNA-associated protein sequences. The snoRNP database can be found at: http://evolveathome.com/snoRNA/snoRNA.php.