The last decade has seen an enormous expansion in the genetic toolbox of model organisms. This has been particularly apparent in the fruitfly, Drosophila melanogaster, in which the development of these tools and their ongoing extension, along with the completion of the genome sequence has allowed for the analysis of most any process. One particularly elegant example of tool development was the creation of the GAL4/UAS system for targeted gene expression in Drosophila. In addition to helping make Drosophila one of the most genetically tractable metazoans, this system has also helped Drosophila attract attention from the biotechnology industry as a viable means to investigate the function of genes implicated in a wide variety of medically and economically important processes. In this article, we review the GAL4/UAS system in Drosophila and the numerous extensions that have morphed it into a veritable Swiss army knife for the analysis of gene function.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/gene.10150

GAL4 system in Drosophila: a fly geneticist's Swiss army knife.

Transposable elements are mobile genetic units that exhibit broad diversity in their structure and transposition mechanisms. Transposable elements occupy a large fraction of many eukaryotic genomes and their movement and accumulation represent a major force shaping the genes and genomes of almost all organisms. This review focuses on DNA-mediated or class 2 transposons and emphasizes how this class of elements is distinguished from other types of mobile elements in terms of their structure, amplification dynamics, and genomic effect. We provide an up-to-date outlook on the diversity and taxonomic distribution of all major types of DNA transposons in eukaryotes, including Helitrons and Mavericks. We discuss some of the evolutionary forces that influence their maintenance and diversification in various genomic environments. Finally, we highlight how the distinctive biological features of DNA transposons have contributed to shape genome architecture and led to the emergence of genetic innovations in different eukaryotic lineages.

/pdf/dna-transposons-and-the-evolution-of-eukaryotic-genomes-4rq0mdhrsc.pdf

DNA Transposons and the Evolution of Eukaryotic Genomes

The Berkeley Drosophila Genome Project (BDGP) strives to disrupt each Drosophila gene by the insertion of a single transposable element. As part of this effort, transposons in >30,000 fly strains were localized and analyzed relative to predicted Drosophila gene structures. Approximately 6300 lines that maximize genomic coverage were selected to be sent to the Bloomington Stock Center for public distribution, bringing the size of the BDGP gene disruption collection to 7140 lines. It now includes individual lines predicted to disrupt 5362 of the 13,666 currently annotated Drosophila genes (39%). Other lines contain an insertion at least 2 kb from others in the collection and likely mutate additional incompletely annotated or uncharacterized genes and chromosomal regulatory elements. The remaining strains contain insertions likely to disrupt alternative gene promoters or to allow gene misexpression. The expanded BDGP gene disruption collection provides a public resource that will facilitate the application of Drosophila genetics to diverse biological problems. Finally, the project reveals new insight into how transposons interact with a eukaryotic genome and helps define optimal strategies for using insertional mutagenesis as a genomic tool.

/pdf/the-bdgp-gene-disruption-project-single-transposon-4j3rm9e1ix.pdf

The BDGP gene disruption project: single transposon insertions associated with 40% of Drosophila genes.

C/EBP homologous protein (CHOP) is a stress-inducible nuclear protein that is crucial for the development of programmed cell death and regeneration; however, the regulation of its function has not been well characterized. Slbo, a Drosophila homolog of C/EBP (CCAAT/enhancer binding protein), was shown to be unstabilized by tribbles. Here, we identified TRB3 as a tribbles ortholog in humans, which associated with CHOP to suppress the CHOP-dependent transactivation. TRB3 is induced by various forms endoplasmic reticulum (ER) stress later than CHOP. Tunicamycin treatment enhanced the TRB3 promoter activity, while dominant-negative forms of CHOP suppressed the tunicamycin-induced activation. In addition, the tunicamycin response region in the TRB3 promoter contains amino-acid response elements overlapping the CHOP-binding site, and CHOP and ATF4 cooperated to activate this promoter activity. Knockdown of endogenous ATF4 or CHOP expression dramatically repressed tunicamycin-induced TRB3 induction. Furthermore, knockdown of TRB3 expression decreased ER stress-dependent cell death. These results indicate that TRB3 is a novel target of CHOP/ATF4 and downregulates its own induction by repression of CHOP/ATF4 functions, and that it is involved in CHOP-dependent cell death during ER stress.

/pdf/trb3-a-novel-er-stress-inducible-gene-is-induced-via-atf4-3i3u6aq79c.pdf

TRB3, a novel ER stress-inducible gene, is induced via ATF4–CHOP pathway and is involved in cell death

Insulin resistance is a major hallmark in the development of type II diabetes, which is characterized by the failure of insulin to promote glucose uptake in muscle and to suppress glucose production in liver. The serine-threonine kinase Akt (PKB) is a principal target of insulin signaling that inhibits hepatic glucose output when glucose is available from food. Here we show that TRB3, a mammalian homolog of Drosophila tribbles, functions as a negative modulator of Akt. TRB3 expression is induced in liver under fasting conditions, and TRB3 disrupts insulin signaling by binding directly to Akt and blocking activation of the kinase. Amounts of TRB3 RNA and protein were increased in livers of db/db diabetic mice compared with those in wild-type mice. Hepatic overexpression of TRB3 in amounts comparable to those in db/db mice promoted hyperglycemia and glucose intolerance. Our results suggest that, by interfering with Akt activation, TRB3 contributes to insulin resistance in individuals with susceptibility to type II diabetes.

https://science.sciencemag.org/content/sci/300/5625/1574.full.pdf

TRB3: A tribbles Homolog That Inhibits Akt/PKB Activation by Insulin in Liver

A modular misexpression system was used to carry out systematic gain-of-function genetic screens in Drosophila. The system is based on inducible expression of genes tagged by insertion of a P-element vector carrying a GAL4-regulated promoter oriented to transcribe flanking genomic sequences. To identify genes involved in eye and wing development, the 2300 independent lines were screened for dominant phenotypes. Among many novel genes, the screen identified known genes, including hedgehog and decapentaplegic, implicated in these processes. A genetic interaction screen for suppressors of a cell migration defect in a hypomorphic slow border cells mutant identified known genes with likely roles in tyrosine kinase signaling and control of actin cytoskeleton, among many novel genes. These studies demonstrate the ability of the modular misexpression system to identify developmentally important genes and suggest that it will be generally useful for genetic interaction screens.

http://www.cscb.northwestern.edu/jcpdfs/rorth98.pdf

Systematic gain-of-function genetics in Drosophila

https://www.cell.com/molecular-cell/pdf/S1097-2765(05)00008-0.pdf

The Level of C/EBP Protein Is Critical for Cell Migration during Drosophila Oogenesis and Is Tightly Controlled by Regulated Degradation

The existence of a gut-skin axis is supported by increasing evidence, but its translational potential is not widely recognized. Studies linked inflammatory skin diseases to an imbalanced gut microbiome; hence, the modulation of the gut microbiota to improve skin condition seems to be a feasible approach. Today, there is a growing interest in natural products as alternatives to synthetic drugs. In this respect, oral probiotics could be a simple, safe and cheap modality in the therapeutic management of skin inflammation. Unfortunately, very few studies have looked into how probiotic supplementation influences inflammatory skin disorders. The result, though promising, are difficult to implement in clinical practice due to the heterogeneity of the applied supplemental regimen in the different studies. In this Viewpoint, we aim to encourage the conduction of more research in that direction to explore unambiguously the therapeutic potential of oral probiotics in dermatology. We focus on the most common inflammatory skin diseases (atopic dermatitis, psoriasis, rosacea, acne vulgaris) with an associated gut dysbiosis, but we also discuss some less common, but very serious skin pathologies (eg erythema nodosum, pyoderma gangrenosum, hidradenitis suppurativa) that are possibly linked to a disturbed gut flora composition. We dissect the possible mechanisms along the gut-skin axis and highlight novel points where probiotics could interfere in this communication in the diseased state.

https://onlinelibrary.wiley.com/doi/epdf/10.1111/exd.14016

Targeting the gut-skin axis—Probiotics as new tools for skin disorder management?

We used the Sleeping Beauty (SB) transposable element as a tool to probe transposon-host cell interactions in vertebrates. The Miz-1 transcription factor was identified as an interactor of the SB transposase in a yeast two-hybrid screen. Through its association with Miz-1, the SB transposase down-regulates cyclin D1 expression in human cells, as evidenced by differential gene expression analysis using microarray hybridization. Down-regulation of cyclin D1 results in a prolonged G(1) phase of the cell cycle and retarded growth of transposase-expressing cells. G(1) slowdown is associated with a decrease of cyclin D1/cdk4-specific phosphorylation of the retinoblastoma protein. Both cyclin D1 down-regulation and the G(1) slowdown induced by the transposase require Miz-1. A temporary G(1) arrest enhances transposition, suggesting that SB transposition is favored in the G(1) phase of the cell cycle, where the nonhomologous end-joining pathway of DNA repair is preferentially active. Because nonhomologous end-joining is required for efficient SB transposition, the transposase-induced G(1) slowdown is probably a selfish act on the transposon's part to maximize the chance for a successful transposition event.

https://www.pnas.org/content/pnas/103/11/4062.full.pdf

Sleeping Beauty transposase modulates cell-cycle progression through interaction with Miz-1.

Summary

Background Merkel cell carcinoma (MCC) is a rare, aggressive tumour for which an increasing incidence has been reported. A new human polyomavirus, Merkel cell polyomavirus (MCV), was recently isolated from these tumours by applying digital transcriptome subtraction methodology.



Objectives  To detect the presence or absence of MCV in MCCs and other, randomly selected neoplasms.



Methods  Nine primary or recurrent MCCs from seven patients were examined; 29 other tumours (squamous cell, basal cell and basosquamous carcinomas and malignant melanomas) were examined for comparative purposes. Viral large T protein (LT1 and LT3), and viral capsid protein (VP1) were detected by primer-directed amplification, using a polymerase chain reaction (PCR)-based method, and the amplified PCR products were analysed by agarose gel electrophoresis and subsequent sequence analysis.



Results  The presence of viral T antigen and/or viral capsid DNA sequences was demonstrated in seven of the eight MCC lesions. None of the comparative samples contained MCV DNA.



Conclusions  Our findings strongly support the hypothesis that MCV infection may well be specific for MCC, and MCV may play a role in the pathogenesis of MCC.

Kornélia Szabó

Papers

Systematic gain-of-function genetics in Drosophila

The Level of C/EBP Protein Is Critical for Cell Migration during Drosophila Oogenesis and Is Tightly Controlled by Regulated Degradation

Targeting the gut-skin axis—Probiotics as new tools for skin disorder management?

Sleeping Beauty transposase modulates cell-cycle progression through interaction with Miz-1.

Detection of Merkel cell polyomavirus DNA in Merkel cell carcinomas.