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N-terminal tagging of RNA Polymerase II shapes transcriptomes more than C-terminal alterations

Adam Callan-Sidat, Emmanuel Zewdu, Massimo Cavallaro, Juntai Liu, Daniel Hebenstreit

RNA polymerase II (Pol2) has a C-terminal domain (CTD) that is unstructured, consists of a large number of heptad repeats, and whose precise function remains unclear. Here, we investigate how altering the CTD’s length and fusing it with protein tags affects transcriptional output on genome-wide scale in mammalian cells, at single cell resolution. While transcription generally appears to occur in burst-like fashion, where RNA is predominantly made during short bursts of activity that are interspersed with periods of transcriptional silence, the CTD’s role in shaping these dynamics seems gene-dependent; global patterns of bursting appear mostly robust to CTD alterations

iScience. May 2024

Tue 28 May 2024, 08:03 | Tags: Quantitative, Systems & Engineering Biology

RecGraph: recombination-aware alignment of sequences to variation graphs

Jorge Avila Cartes, Paola Bonizzoni, Simone Ciccolella, Gianluca Della Vedova, Luca Denti, Xavier Didelot, Davide Monti, Yuri Pirola

In this paper, we present the extension of the notion of sequence-to-graph alignment to a variation graph that incorporates a recombination, so that the latter are explicitly represented and evaluated in an alignment. Moreover, we present a dynamic programming approach for the special case where there is at most a recombination—we implement this case as RecGraph. From a modelling point of view, a recombination corresponds to identifying a new path of the variation graph, where the new arc is composed of two halves, each extracted from an original path, possibly joined by a new arc. Our experiments show that RecGraph accurately aligns simulated recombinant bacterial sequences that have at most a recombination, providing evidence for the presence of recombination events. 

Bioinformatics. April 2024

Thu 23 May 2024, 08:31 | Tags: Microbiology & Infectious Disease

Multiple toxins and a protease contribute to the aphid-killing ability of Pseudomonas fluorescens PpR24

Deepa Paliwal, Mojgan Rabiey, Tim H. Mauchline

Aphids are globally important pests causing damage to a broad range of crops. Due to insecticide resistance, there is an urgent need to develop alternative control strategies. In our previous work, we found Pseudomonas fluorescens PpR24 can orally infect and kill the insecticide-resistant green-peach aphid (Myzus persicae). However, the genetic basis of the insecticidal capability of PpR24 remains unclear. This comprehensive host–pathogen transcriptomic analysis provides novel insight into the molecular basis of bacteria-mediated aphid mortality and the potential of PpR24 as an effective biocontrol agent.

Environmental Biology. April 2024

Thu 23 May 2024, 08:23 | Tags: Plant & Agricultural Bioscience

Imaging Single-Cell Ca2+ Dynamics of Brainstem Neurons and Glia in Freely Behaving Mice

Amol M. Bhandare, Nicholas Dale, Robert T. R. Huckstepp

In vivo brain imaging, using a combination of genetically encoded Ca2+ indicators and gradient refractive index (GRIN) lens, is a transformative technology that has become an increasingly potent research tool over the last decade. We have refined the intravital imaging technique to image deep brain nuclei in the ventral medulla oblongata, one of the most difficult brain structures to image due to the movement of brainstem structures outside the cranial cavity during free behaviour (head and neck movement), whose targeting requires GRIN lens insertion through the cerebellum—a key structure for balance and movement. Our protocol refines the implantation method of GRIN lenses, giving the best possible approach to image deep extracranial brainstem structures in awake rodents with improved cell rejection/acceptance criteria during analysis. This revised method paves the way to image challenging brainstem structures to investigate their role in complex behaviours such as breathing, circulation, sleep, digestion, and swallowing, and could be extended to image and study the role of cerebellum in balance, movement, motor learning, and beyond.

Bio-protocol. April 2024

Xavier Didelot publications

Epistasis, core-genome disharmony, and adaptation in recombining bacteria

Aidan J. Taylor, Koji Yahara, Ben Pascoe, Seungwon Ko, Leonardos Mageiros, Evangelos Mourkas, Jessica K. Calland, Santeri Puranen, Matthew D. Hitchings, Keith A. Jolley, Carolin M. Kobras, Sion Bayliss, Nicola J. Williams, Arnoud H. M. van Vliet, Julian Parkhill, Martin C. J. Maiden, Jukka Corander, Laurence D. Hurst, Daniel Falush, Paul Keim, Xavier Didelot, David J. Kelly, Samuel K. Sheppard

Recombination of short DNA fragments via horizontal gene transfer (HGT) can introduce beneficial alleles, create genomic disharmony through negative epistasis, and create adaptive gene combinations through positive epistasis. For non-core (accessory) genes, the negative epistatic cost is likely to be minimal because the incoming genes have not co-evolved with the recipient genome and are frequently observed as tightly linked cassettes with major effects. By contrast, interspecific recombination in the core genome is expected to be rare because disruptive allelic replacement is likely to introduce negative epistasis. Why then is homologous recombination common in the core of bacterial genomes? To understand this enigma, we take advantage of an exceptional model system, the common enteric pathogens Campylobacter jejuni and C. coli that are known for very high magnitude interspecies gene flow in the core genome. As expected, HGT does indeed disrupt co-adapted allele pairings, indirect evidence of negative epistasis. However, multiple HGT events enable recovery of the genome's co-adaption between introgressing alleles, even in core metabolism genes (e.g., formate dehydrogenase). These findings demonstrate that, even for complex traits, genetic coalitions can be decoupled, transferred, and independently reinstated in a new genetic background-facilitating transition between fitness peaks. mBio. April 2024

KmerAperture: Retaining k-mer synteny for alignment-free extraction of core and accessory differences between bacterial genomes

Matthew P. Moore, Mirjam Laager,Paolo Ribeca, Xavier Didelot

By decomposing genome sequences into k-mers, it is possible to estimate genome differences without alignment. Techniques such as k-mer minimisers, for example MinHash, have been developed and are often accurate approximations of distances based on full k-mer sets. These and other alignment-free methods avoid the large temporal and computational expense of alignment. However, these k-mer set comparisons are not entirely accurate within-species and can be completely inaccurate within-lineage. This is due, in part, to their inability to distinguish core polymorphism from accessory differences. Here we present a new approach, KmerAperture, which uses information on the k-mer relative genomic positions to determine the type of polymorphism causing differences in k-mer presence and absence between pairs of genomes. We show that KmerAperture can accurately distinguish both core and accessory sequence diversity without alignment, outperforming other k-mer based tools.

PLos Genetics.; April 2024

Thu 16 May 2024, 09:04 | Tags: Microbiology & Infectious Disease

Genetic Variation of Turnip Yellows Virus in Arable and Vegetable Brassica Crops, Perennial Wild Brassicas, and Aphid Vectors Collected from the Plants

Ricardo J G Pimenta, Kyle Macleod, Robyn Babb, Kaitlyn Coleman, Joni MacDonald, Elvis Asare-Bediako, Max J Newbert, Carol E Jenner, John A Walsh

Turnip yellows virus (TuYV; Polerovirus, Solemoviridae) infects and causes yield losses in a range of economically important crop species, particularly the Brassicaceae. Although the incidence and genetic diversity of TuYV has been extensively investigated in recent years, little is known about how the diversity within host plants relates to that in its vectors. Arable oilseed rape (Brassica napus) and vegetable brassica plants (Brassica oleracea), wild cabbage (B. oleracea), and aphids present on these plants were sampled in the field in three regions of the United Kingdom. High levels of TuYV (82 to 97%) were detected in plants in all three regions following enzyme-linked immunosorbent assaysComparative analyses between TuYV sequences from host plants and B. brassicae collected from respective plants revealed differences between some ORF0 sequences, which possibly indicated that at least two of the aphids might not have been carrying the same TuYV isolates as those present in their host plants. Maximum likelihood phylogenetic analyses revealed three distinct major clades for ORF0 and one for ORF3, with some distinct subclades. Some clustering was related to geographic origin. Explanations for TuYV sequence differences between plants and the aphids present on respective plants and implications for the epidemiology and control of TuYV are discussed.

Plant Disease. March 2024

Sun 05 May 2024, 17:41 | Tags: Plant & Agricultural Bioscience

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