Biology subjects

Andersen, J. B.

Publications and source records attributed to Andersen, J. B..

5 recordsLinked to original sources

bioRxiv2023

Inhibition of Pseudomonas aeruginosa quorum sensing by chemical induction of the MexEF-oprN efflux pump

The cell-to-cell communication system quorum sensing (QS), used by various pathogenic bacteria to synchronize gene expression and increase host invasion potentials, is studied as a potential target for persistent infection control. To search for novel molecules targeting the QS system in the Gram-negative opportunistic pathogen Pseudomonas aeruginosa a chemical library consisting of 3280 small compounds from LifeArc was screened. A series of ten conjugated phenones that have not previously been reported to target bacteria were identified as inhibitors of QS in P. aeruginosa. Two lead compounds (ethylthio enynone and propylthio enynone) were resynthesized for verification of activity and further elucidation of the mode of action. The isomeric pure Z-ethylthio enynone was used for RNA sequencing revealing a strong inhibitor of QS-regulated genes and the QS-regulated virulence factors rhamnolipid and pyocyanin were significantly decreased by treatment with the compounds. A transposon mutagenesis screen performed in a newly constructed lasB-gfp monitor strain identified the target of Z-ethylthio enynone in P. aeruginosa to be the MexEF-OprN efflux pump, which was further established using defined mex knockout mutants. Our data indicate that the QS inhibitory capabilities of Z-ethylthio enynone were caused by the drainage of intracellular signal molecules as a response to chemical-induced stimulation of the MexEF-oprN efflux pump thereby inhibiting the auto-generated positive feedback and its enhanced signal-molecule synthesis.

microbiology↗

bioRxiv2022

Alternative promoters in CpG depleted regions pervasively account for epigenetic misregulation of cancer transcriptomes

Eukaryotic genes are regulated by multiple alternative promoters with distinct expression patterns. In cancer, alternative promoters are pervasively utilized, but our understanding of the mechanism of activation and how their regulatory architecture differs from reference promoters remains elusive. We analyzed 100 CAGE-seq libraries from HCC patients and annotated 4083 alternative promoters in 2926 multi-promoter genes that are known genes involved in hepatocarcinogenesis. Many alternative promoters are undetected in the normal liver. We find that multi-promoter genes are enriched among genes downregulated in the tumor, highlighting alternative promoters impact in global transcription changes in cancer. Alternative promoters are depleted for CpG islands, have narrow nucleosome depleted regions, and are enriched for sharp promoters as well as tissue-specific transcription factors. Alternative promoters have high DNA methylation levels around transcription start sites. Tumor cells globally lose DNA methylation, but there exists a hierarchical retention of intragenic DNA methylation, which is dictated by the genomic CG content. As such, intragenic CG-poor regions lose methylation, while CG-rich regions retain it, a phenomenon caused by differential binding of H3K36me3, DNMT3B, TET1 and SETD2. Thus, the selective loss of DNA methylation in CG-poor regions opens the chromatin and makes these regions accessible for transcription. Upon transcription factors availability, alternative transcription can pervasively occur in cancer. These results provide a framework for understanding the importance of alternative promoters in controlling the tumor transcriptomes, highlighting their architecture and role in regulatory mechanism(s).

genomics↗

bioRxiv2020

Determination of primary microRNA processing in clinicalsamples by targeted pri-miR-sequencing

MicroRNA expression is important for gene regulation and deregulated microRNA expression is often observed in disease such as cancer. The processing of primary microRNA transcripts is an important regulatory step in microRNA biogenesis. Due to low expression level and association with chromatin primary microRNAs are challenging to study in clinical samples where input material is limited. Here, we present a high-sensitivity targeted method to determine processing efficiency of several hundred primary microRNAs from total RNA using as little as 500 thousand Illumina HiSeq sequencing reads. We validate the method using RNA from HeLa cells and show the applicability to clinical samples by analyzing RNA from normal liver and hepatocellular carcinoma. We identify 24 primary microRNAs with significant changes in processing efficiency from normal liver to hepatocellular carcinoma, among those the highly expressed miRNA-122 and miRNA-21, demonstrating that differential processing of primary microRNAs is occurring and could be involved in disease. With our method presented here we provide means to study pri-miRNA processing in disease from clinical samples.

molecular biology↗

bioRxiv2018

Intertwined canonical and non-canonical initiation in dual promoters are pervasive and differentially regulate Polymerase II transcription

The diversity and complexity of transcription start site (TSS) selection reflects variation of preinitiation complexes, divergent function of promoter-binding proteins and underlies not only transcriptional dynamics but may also impact on post-transcriptional fates of RNAs. The majority of metazoan genes are transcribed by RNA polymerase II from a canonical initiation motif having an YR dinucleotide at their TSSs. In contrast, translation machinery-associated genes carry promoters with polypyrimidine initiator (known as 5-TOP or TCT) with cytosine replacing the R nucleotide. The functional significance of start site choice in promoter architectures is little understood. To get insight into the developmental regulation of start site selection we profiled 5 ends of transcripts during zebrafish embryogenesis. We uncovered a novel class of dual-initiation (DI) promoters utilized by thousands of genes. In DI promoters non-canonical YC-initiation representing 5-TOP/TCT initiators is intertwined with canonical YR-initiation. During maternal to zygotic transition, the two initiation types are divergently used in hundreds of DI promoters, demonstrating that the two initiation systems are distinctly regulated. We show via the example of snoRNA host genes and translation interference experiments that dual-initiation from shared promoters can lead to divergent spatio-temporal expression dynamics generating distinct sets of RNAs with different post-transcriptional fates. Thus utilization of DI promoters in large number of genes suggests two transcription initiation mechanisms targeting these promoters. DI promoters are conserved within human and fruit fly and reflect an evolutionary conserved mechanism for switching transcription initiation to adapt to the changing developmental context. Thus, our findings highlight a novel level of complexity of core promoter regulation in metazoans and broaden the scope for identification and characterization of alternative RNA products generated at shared core promoters.

genomics↗

bioRxiv2018

HOTAIR ancient sequence suggests regulatory roles both in cis and trans

HOTAIR is a long noncoding RNA transcribed between HOXC11 and HOXC12 in mammals. The proposed function(s) of HOTAIR lacks consensus as to whether it regulates HoxD cluster genes in trans or HoxC cluster genes in cis. We have identified a 32-nucleotide long conserved noncoding element (CNE) as HOTAIR ancient sequence which has a paralogous copy embedded in HOXD11 noncoding transcript. All vertebrates except teleosts have two copies of CNE and the paralogous CNEs exhibit sequence complementarity in the transcribed orientation. Moreover, paralogous CNEs underwent compensatory mutations suggesting they co-evolved and might hybridize. In both human and mouse, HOTAIR CNE exhibits characteristic features of a poised enhancer in HOTAIR-unexpressed stem cells and of an active enhancer in HOTAIR-expressed cells. Tight correlation between the transcriptional activity of the CNE and HOTAIR promoter suggests HOTAIR transcription is crucial for enhancer activity. In HOTAIR-expressed cells, HOTAIR expression is positively correlated with HOXC11 in cis and negatively correlated with HOXD11 in trans, suggesting a dual modality of HOTAIR ancient sequence.

genomics↗