Dev Biol. 2023 Oct 23:S0012-1606(23)00168-9. doi: 10.1016/j.ydbio.2023.09.014. Online ahead of print.

ABSTRACT

The orphan nuclear receptor Tailless (Tll) exhibits conserved roles in brain formation and maintenance that are shared, for example, with vertebrate orthologous forms (Tlx). However, the early expression of tll in two gap domains in the segmentation cascade of Drosophila is unusual even for most other insects. Here we investigate tll regulation on pair-rule stripes. With ectopic misexpression of tll we detected unexpected repression of almost all pair-rule stripes of hairy (h), even-skipped (eve), runt (run), and fushi-tarazu (ftz). Examining Tll embryonic ChIP-chip data with regions mapped as Cis-Regulatory Modules (CRMs) of pair-rule stripes we verified Tll interactions to these regions. With the ChIP-chip data we also verified Tll interactions to the CRMs of gap domains and in the misexpression assay, Tll-mediated repression on Kruppel (Kr), kni (kni) and giant (gt) according to their differential sensitivity to Tll. These results with gap genes confirmed previous data from the literature and argue against indirect repression roles of Tll in the striped pattern. Moreover, the prediction of Tll binding sites in the CRMs of eve stripes and the mathematical modeling of their removal using an experimentally validated theoretical framework shows effects on eve stripes compatible with the absence of a repressor binding to the CRMs. In addition, modeling increased tll levels in the embryo results in the differential repression of eve stripes, agreeing well with the results of the misexpression assay. In genetic assays we investigated eve 5, that is strongly repressed by the ectopic domain and representative of more central stripes not previously implied to be under direct regulation of tll. While this stripe is little affected in tll-, its posterior border is expanded in gt- but detected with even greater expansion in gt-;tll-. We end up by discussing tll with key roles in combinatorial repression mechanisms to contain the expression of medial patterns of the segmentation cascade in the extremities of the embryo.

PMID:37879494 | DOI:10.1016/j.ydbio.2023.09.014

Res Sq. 2023 Sep 28:rs.3.rs-3294408. doi: 10.21203/rs.3.rs-3294408/v1. Preprint.

ABSTRACT

Genome mapping studies have generated a nearly complete collection of genes for the human genome, but we still lack an equivalently vetted inventory of human regulatory sequences. Cis-regulatory modules (CRMs) play important roles in controlling when, where, and how much a gene is expressed. We developed a training data-free CRM-prediction algorithm, the Mammalian Regulatory MOdule Detector (MrMOD) for accurate CRM prediction in mammalian genomes. MrMOD provides genome position-fixed CRM models similar to the fixed gene models for the mouse and human genomes using only genomic sequences as the inputs with one adjustable parameter - the significance p-value. Importantly, MrMOD predicts a comprehensive set of high-resolution CRMs in the mouse and human genomes including all types of regulatory modules not limited to any tissue, cell type, developmental stage, or condition. We computationally validated MrMOD predictions used a compendium of 21 orthogonal experimental data sets including thousands of experimentally defined CRMs and millions of putative regulatory elements derived from hundreds of different tissues, cell types, and stimulus conditions obtained from multiple databases. In ovo transgenic reporter assay demonstrates the power of our prediction in guiding experimental design. We analyzed CRMs located in the chromosome 17 using unsupervised machine learning and identified groups of CRMs with multiple lines of evidence supporting their functionality, linking CRMs with upstream binding transcription factors and downstream target genes. Our work provides a comprehensive base pair resolution annotation of the functional regulatory elements and non-functional regions in the mammalian genomes.

PMID:37841836 | PMC:PMC10571623 | DOI:10.21203/rs.3.rs-3294408/v1

J Virol. 2023 Oct 12:e0130523. doi: 10.1128/jvi.01305-23. Online ahead of print.

ABSTRACT

Following acute infection, sensory neurons in trigeminal ganglia (TG) are important sites for the life-long latency of human alpha-herpes virus 1 (HSV-1). Acute or chronic stress in humans correlates with increased reactivation from latency, which can lead to recurrent HSV-1 disease, for example, herpes labialis, herpes stromal keratitis, and encephalitis. The glucocorticoid receptor (GR) and the synthetic corticosteroid dexamethasone stimulate key viral transcriptional cis-regulatory modules, viral replication, and explant-induced reactivation from latency. Conversely, a GR-specific antagonist impairs explant-induced reactivation and viral replication. Based on these observations, we hypothesize that GR transcriptional activity enhances reactivation from latency. To test this hypothesis, the HSV-1 latency-reactivation cycle was examined in mice containing a serine 229 to alanine mutation in GR (GRS229A) because phosphorylation of GR serine 229 is crucial for GR-mediated transcription. Virus yields from cornea and conjunctiva of infected GRS229A mice ceased before wild-type (wt) mice, consistent with reduced viral replication in kidney cells from GRS229A mice. However, viral DNA levels in TG were not significantly different during latency and similar numbers of TG neurons express GR in GRS229A and WT mice. Strikingly, HSV-1 viral titers during explant-induced reactivation were significantly reduced in female GRS229A mice versus male GRS229A mice or wt mice. The number of VP16 + TG neurons in female GRS229A mice was significantly lower than in male GRS229A or wt mice (males and females) during the early stages of explant-induced reactivation. Collectively, these studies revealed that GR phosphorylation of serine 229 is more important in GRS229A female mice versus males during explant-induced reactivation from latency. IMPORTANCE A correlation exists between stress and increased episodes of human alpha-herpes virus 1 reactivation from latency. Stress increases corticosteroid levels; consequently, the glucocorticoid receptor (GR) is activated. Recent studies concluded that a GR agonist, but not an antagonist, accelerates productive infection and reactivation from latency. Furthermore, GR and certain stress-induced transcription factors cooperatively transactivate promoters that drive the expression of infected cell protein 0 (ICP0), ICP4, and VP16. This study revealed female mice expressing a GR containing a serine to alanine mutation at position 229 (GRS229A) shed significantly lower levels of infectious virus during explant-induced reactivation compared to male GRS229A or wild-type parental C57BL/6 mice. Furthermore, female GRS229A mice contained fewer VP16 + TG neurons compared to male GRS229A mice or wild-type mice during the early stages of explant-induced reactivation from latency. Collectively, these studies revealed that GR transcriptional activity has female-specific effects, whereas male mice can compensate for the loss of GR transcriptional activation.

PMID:37823644 | DOI:10.1128/jvi.01305-23

NAR Genom Bioinform. 2023 Sep 22;5(3):lqad085. doi: 10.1093/nargab/lqad085. eCollection 2023 Sep.

ABSTRACT

Self-transcribing active regulatory region sequencing (STARR-seq) and its variants have been widely used to characterize enhancers. However, it has been reported that up to 87% of STARR-seq peaks are located in repressive chromatin and are not functional in the tested cells. While some of the STARR-seq peaks in repressive chromatin might be active in other cell/tissue types, some others might be false positives. Meanwhile, many active enhancers may not be identified by the current STARR-seq methods. Although methods have been proposed to mitigate systematic errors caused by the use of plasmid vectors, the artifacts due to the intrinsic limitations of current STARR-seq methods are still prevalent and the underlying causes are not fully understood. Based on predicted cis-regulatory modules (CRMs) and non-CRMs in the human genome as well as predicted active CRMs and non-active CRMs in a few human cell lines/tissues with STARR-seq data available, we reveal prevalent false positives and false negatives in STARR-seq peaks generated by major variants of STARR-seq methods and possible underlying causes. Our results will help design strategies to improve STARR-seq methods and interpret the results.

PMID:37745976 | PMC:PMC10516709 | DOI:10.1093/nargab/lqad085

PLoS One. 2023 Aug 17;18(8):e0290035. doi: 10.1371/journal.pone.0290035. eCollection 2023.

ABSTRACT

The cis-regulatory data that help in transcriptional regulation is arranged into modular pieces of a few hundred base pairs called CRMs (cis-regulatory modules) and numerous binding sites for multiple transcription factors are prominent characteristics of these cis-regulatory modules. The present study was designed to localize transcription factor binding site (TFBS) clusters on twelve Anterior-posterior (A-P) genes in Tribolium castaneum and compare them to their orthologous gene enhancers in Drosophila melanogaster. Out of the twelve A-P patterning genes, six were gap genes (Kruppel, Knirps, Tailless, Hunchback, Giant, and Caudal) and six were pair rule genes (Hairy, Runt, Even-skipped, Fushi-tarazu, Paired, and Odd-skipped). The genes along with 20 kb upstream and downstream regions were scanned for TFBS clusters using the Motif Cluster Alignment Search Tool (MCAST), a bioinformatics tool that looks for set of nucleotide sequences for statistically significant clusters of non-overlapping occurrence of a given set of motifs. The motifs used in the current study were Hunchback, Caudal, Giant, Kruppel, Knirps, and Even-skipped. The results of the MCAST analysis revealed the maximum number of TFBS for Hunchback, Knirps, Caudal, and Kruppel in both D. melanogaster and T. castaneum, while Bicoid TFBS clusters were found only in D. melanogaster. The size of all the predicted TFBS clusters was less than 1kb in both insect species. These sequences revealed more transversional sites (Tv) than transitional sites (Ti) and the average Ti/Tv ratio was 0.75.

PMID:37590227 | DOI:10.1371/journal.pone.0290035

BMC Biol. 2023 Jul 31;21(1):165. doi: 10.1186/s12915-023-01665-4.

ABSTRACT

BACKGROUND: The development of cotton fiber is regulated by the orchestrated binding of regulatory proteins to cis-regulatory elements associated with developmental genes. The cis-trans regulatory dynamics occurred throughout the course of cotton fiber development are elusive. Here we generated genome-wide high-resolution DNase I hypersensitive sites (DHSs) maps to understand the regulatory mechanisms of cotton ovule and fiber development.

RESULTS: We generated DNase I hypersensitive site (DHS) profiles from cotton ovules at 0 and 3 days post anthesis (DPA) and fibers at 8, 12, 15, and 18 DPA. We obtained a total of 1185 million reads and identified a total of 199,351 DHSs through ~ 30% unique mapping reads. It should be noted that more than half of DNase-seq reads mapped multiple genome locations and were not analyzed in order to achieve a high specificity of peak profile and to avoid bias from repetitive genomic regions. Distinct chromatin accessibilities were observed in the ovules (0 and 3 DPA) compared to the fiber elongation stages (8, 12, 15, and 18 DPA). Besides, the chromatin accessibility during ovules was particularly elevated in genomic regions enriched with transposable elements (TEs) and genes in TE-enriched regions were involved in ovule cell division. We analyzed cis-regulatory modules and revealed the influence of hormones on fiber development from the regulatory divergence of transcription factor (TF) motifs. Finally, we constructed a reliable regulatory network of TFs related to ovule and fiber development based on chromatin accessibility and gene co-expression network. From this network, we discovered a novel TF, WRKY46, which may shape fiber development by regulating the lignin content.

CONCLUSIONS: Our results not only reveal the contribution of TEs in fiber development, but also predict and validate the TFs related to fiber development, which will benefit the research of cotton fiber molecular breeding.

PMID:37525156 | DOI:10.1186/s12915-023-01665-4

Front Cell Dev Biol. 2023 Jul 10;11:1215406. doi: 10.3389/fcell.2023.1215406. eCollection 2023.

ABSTRACT

Introduction: The articulating ends of limb bones have precise morphology and asymmetry that ensures proper joint function. Growth differentiation factor 5 (Gdf5) is a secreted morphogen involved in cartilage and bone development that contributes to the architecture of developing joints. Dysregulation of Gdf5 results in joint dysmorphogenesis often leading to progressive joint degeneration or osteoarthritis (OA). The transcription factors and cis-regulatory modules (CRMs) that regulate Gdf5 expression are not well characterized. We previously identified a Gdf5-associated regulatory region (GARR) that contains predicted binding sites for Lmx1b, Osr2, Fox, and the Sox transcription factors. These transcription factors are recognized factors involved in joint morphogenesis and skeletal development. Methods: We used in situ hybridization to Gdf5, Col2A1, and the transcription factors of interest in developing chicken limbs to determine potential overlap in expression. We further analyzed scRNA-seq data derived from limbs and knees in published mouse and chicken datasets, identifying cells with coexpression of Gdf5 and the transcription factors of interest. We also performed site-directed mutatgenesis of the predicted transcription factor binding sites in a GARR-reporter construct and determined any change in activity using targeted regional electroporation (TREP) in micromass and embryonic chicken wing bioassays. Results: Gdf5 expression overlapped the expression of these transcription factors during joint development both by in situ hybridization (ISH) and scRNA-seq analyses. Within the GARR CRM, mutation of two binding sites common to Fox and Sox transcripstion factors reduced enhancer activity to background levels in micromass cultures and in ovo embryonic chicken wing bioassays, whereas mutation of two Sox-only binding sites caused a significant increase in activity. These results indicate that the Fox/Sox binding sites are required for activity, while the Sox-only sites are involved in repression of activity. Mutation of Lmx1b binding sites in GARR caused an overall reduction in enhancer activity in vitro and a dorsal reduction in ovo. Despite a recognized role for Osr2 in joint development, disruption of the predicted Osr2 site did not alter GARR activity. Conclusion: Taken together, our data indicates that GARR integrates positive, repressive, and asymmetrical inputs to fine-tune the expression of Gdf5 during elbow joint development.

PMID:37492222 | PMC:PMC10364121 | DOI:10.3389/fcell.2023.1215406

Plant Biotechnol J. 2023 Jun 28. doi: 10.1111/pbi.14103. Online ahead of print.

ABSTRACT

High-molecular-weight glutenin subunits (HMW-GS), a major component of seed storage proteins (SSP) in wheat, largely determine processing quality. HMW-GS encoded by GLU-1 loci are mainly controlled at the transcriptional level by interactions between cis-elements and transcription factors (TFs). We previously identified a conserved cis-regulatory module CCRM1-1 as the most essential cis-element for Glu-1 endosperm-specific high expression. However, the TFs targeting CCRM1-1 remained unknown. Here, we built the first DNA pull-down plus liquid chromatography-mass spectrometry platform in wheat and identified 31 TFs interacting with CCRM1-1. TaB3-2A1 as proof of concept was confirmed to bind to CCRM1-1 by yeast one hybrid and electrophoretic mobility shift assays. Transactivation experiments demonstrated that TaB3-2A1 repressed CCRM1-1-driven transcription activity. TaB3-2A1 overexpression significantly reduced HMW-GS and other SSP, but enhanced starch content. Transcriptome analyses confirmed that enhanced expression of TaB3-2A1 down-regulated SSP genes and up-regulated starch synthesis-related genes, such as TaAGPL3, TaAGPS2, TaGBSSI, TaSUS1 and TaSUS5, suggesting that it is an integrator modulating the balance of carbon and nitrogen metabolism. TaB3-2A1 also had significant effects on agronomic traits, including heading date, plant height and grain weight. We identified two major haplotypes of TaB3-2A1 and found that TaB3-2A1-Hap1 conferred lower seed protein content, but higher starch content, plant height and grain weight than TaB3-2A1-Hap2 and was subjected to positive selection in a panel of elite wheat cultivars. These findings provide a high-efficiency tool to detect TFs binding to targeted promoters, considerable gene resources for dissecting regulatory mechanisms underlying Glu-1 expression, and a useful gene for wheat improvement.

PMID:37381172 | DOI:10.1111/pbi.14103

J Virol. 2023 Apr 6:e0007323. doi: 10.1128/jvi.00073-23. Online ahead of print.

ABSTRACT

Stress-mediated activation of the glucocorticoid receptor (GR) and specific stress-induced transcription factors stimulate herpes simplex virus 1 (HSV-1) productive infection, explant-induced reactivation, and immediate early (IE) promoters that drive expression of infected cell protein 0 (ICP0), ICP4, and ICP27. Several published studies concluded the virion tegument protein VP16, ICP0, and/or ICP4 drives early steps of reactivation from latency. Notably, VP16 protein expression was induced in trigeminal ganglionic neurons of Swiss Webster or C57BL/6J mice during early stages of stress-induced reactivation. If VP16 mediates reactivation, we hypothesized stress-induced cellular transcription factors would stimulate its expression. To address this hypothesis, we tested whether stress-induced transcription factors transactivate a VP16 cis-regulatory module (CRM) located upstream of the VP16 TATA box (-249 to -30). Initial studies revealed the VP16 CRM cis-activated a minimal promoter more efficiently in mouse neuroblastoma cells (Neuro-2A) than mouse fibroblasts (NIH-3T3). GR and Slug, a stress-induced transcription factor that binds enhancer boxes (E-boxes), were the only stress-induced transcription factors examined that transactivated the VP16 CRM construct. GR- and Slug-mediated transactivation was reduced to basal levels when the E-box, two 1/2 GR response elements (GREs), or NF-κB binding site was mutated. Previous studies revealed GR and Slug cooperatively transactivated the ICP4 CRM, but not ICP0 or ICP27. Silencing of Slug expression in Neuro-2A cells significantly reduced viral replication, indicating Slug-mediated transactivation of ICP4 and VP16 CRM activity correlates with enhanced viral replication and reactivation from latency. IMPORTANCE Herpes simplex virus 1 (HSV-1) establishes lifelong latency in several types of neurons. Periodically cellular stressors trigger reactivation from latency. Viral regulatory proteins are not abundantly expressed during latency, indicating cellular transcription factors mediate early stages of reactivation. Notably, the glucocorticoid receptor (GR) and certain stress-induced transcription factors transactivate cis-regulatory modules (CRMs) essential for expression of infected cell protein 0 (ICP0) and ICP4, key viral transcriptional regulatory proteins linked to triggering reactivation from latency. Virion protein 16 (VP16) specifically transactivates IE promoter and was also reported to mediate early stages of reactivation from latency. GR and Slug, a stress-induced enhancer box (E-box) binding protein, transactivate a minimal promoter downstream of VP16 CRM, and these transcription factors occupy VP16 CRM sequences in transfected cells. Notably, Slug stimulates viral replication in mouse neuroblastoma cells suggesting Slug, by virtue of transactivating VP16 and ICP4 CRM sequences, can trigger reactivation in certain neurons.

PMID:37022165 | DOI:10.1128/jvi.00073-23

Methods Mol Biol. 2023;2640:397-412. doi: 10.1007/978-1-0716-3036-5_27.

ABSTRACT

Actively transcribed genes harbor cis-regulatory modules with comparatively low nucleosome occupancy and few high-order structures (="open chromatin"), whereas non-transcribed genes are characterized by high nucleosome density and extensive interactions between nucleosomes (="closed chromatin"), preventing transcription factor binding. Knowledge about chromatin accessibility is crucial to understand gene regulatory networks determining cellular decisions. Several techniques are available to map chromatin accessibility, among which the Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) is one of the most popular. ATAC-seq is based on a straightforward and robust protocol but requires adjustments for different cell types. Here, we describe an optimized protocol for ATAC-seq of freshly isolated murine muscle stem cells. We provide details for the isolation of MuSC, tagmentation, library amplification, double-sided SPRI bead cleanup, and library quality assessment and give recommendations for sequencing parameters and downstream analysis. The protocol should facilitate generation of high-quality data sets of chromatin accessibility in MuSCs, even for newcomers to the field.

PMID:36995609 | DOI:10.1007/978-1-0716-3036-5_27

Int J Mol Sci. 2023 Mar 17;24(6):5766. doi: 10.3390/ijms24065766.

ABSTRACT

Long non-coding RNAs (lncRNAs), a class of poorly conserved transcripts without protein-encoding ability, are widely involved in plant organogenesis and stress responses by mediating the transmission and expression of genetic information at the transcriptional, posttranscriptional, and epigenetic levels. Here, we cloned and characterized a novel lncRNA molecule through sequence alignment, Sanger sequencing, transient expression in protoplasts, and genetic transformation in poplar. lncWOX11a is a 215 bp transcript located on poplar chromosome 13, ~50 kbp upstream of PeWOX11a on the reverse strand, and the lncRNA may fold into a series of complex stem-loop structures. Despite the small open reading frame (sORF) of 51 bp within lncWOX11a, bioinformatics analysis and protoplast transfection revealed that lncWOX11a has no protein-coding ability. The overexpression of lncWOX11a led to a decrease in the quantity of adventitious roots on the cuttings of transgenic poplars. Further, cis-regulatory module prediction and CRISPR/Cas9 knockout experiments with poplar protoplasts demonstrated that lncWOX11a acts as a negative regulator of adventitious rooting by downregulating the WUSCHEL-related homeobox gene WOX11, which is supposed to activate adventitious root development in plants. Collectively, our findings imply that lncWOX11a is essential for modulating the formation and development of adventitious roots.

PMID:36982841 | DOI:10.3390/ijms24065766

Elife. 2023 Mar 27;12:e79380. doi: 10.7554/eLife.79380. Online ahead of print.

ABSTRACT

Histone acetylation is a pivotal epigenetic modification that controls chromatin structure and regulates gene expression. It plays an essential role in modulating zygotic transcription and cell lineage specification of developing embryos. While the outcomes of many inductive signals have been described to require enzymatic activities of histone acetyltransferases and deacetylases (HDACs), the mechanisms by which HDACs confine the utilization of the zygotic genome remain to be elucidated. Here, we show that histone deacetylase 1 (Hdac1) progressively binds to the zygotic genome from mid blastula and onward. The recruitment of Hdac1 to the genome at blastula is instructed maternally. Cis-regulatory modules (CRMs) bound by Hdac1 possess epigenetic signatures underlying distinct functions. We highlight a dual function model of Hdac1 where Hdac1 not only represses gene expression by sustaining a histone hypoacetylation state on inactive chromatin, but also maintains gene expression through participating in dynamic histone acetylation-deacetylation cycles on active chromatin. As a result, Hdac1 maintains differential histone acetylation states of bound CRMs between different germ layers and reinforces the transcriptional program underlying cell lineage identities, both in time and space. Taken together, our study reveals a comprehensive role for Hdac1 during early vertebrate embryogenesis.

PMID:36971347 | DOI:10.7554/eLife.79380

Cell Genom. 2023 Feb 2;3(3):100262. doi: 10.1016/j.xgen.2023.100262. eCollection 2023 Mar 8.

ABSTRACT

Precision oncology promises accurate prediction of disease trajectories by utilizing molecular features of tumors. We present a systematic analysis of the prognostic potential of diverse molecular features across large cancer cohorts. We find that the mRNA expression of biologically coherent sets of genes (modules) is substantially more predictive of patient survival than single-locus genomic and transcriptomic aberrations. Extending our analysis beyond existing curated gene modules, we find a large novel class of highly prognostic DNA/RNA cis-regulatory modules associated with dynamic gene expression within cancers. Remarkably, in more than 82% of cancers, modules substantially improve survival stratification compared with conventional clinical factors and prominent genomic aberrations. The prognostic potential of cancer modules generalizes to external cohorts better than conventionally used single-gene features. Finally, a machine-learning framework demonstrates the combined predictive power of multiple modules, yielding prognostic models that perform substantially better than existing histopathological and clinical factors in common use.

PMID:36950380 | PMC:PMC10025453 | DOI:10.1016/j.xgen.2023.100262

Dev Biol. 2023 Mar 10:S0012-1606(23)00045-3. doi: 10.1016/j.ydbio.2023.03.002. Online ahead of print.

ABSTRACT

The establishment of body pattern is a fundamental process in developmental biology. In Drosophila, the wing disc is subdivided into dorsal (D) and ventral (V) compartments by the D/V boundary. The dorsal fate is adopted by expressing the selector gene apterous (ap). ap expression is regulated by three combinational cis-regulatory modules which are activated by EGFR pathway, Ap-Vg auto-regulatory and epigenetic mechanisms. Here, we found that the Tbx family transcription factor Optomotor-blind (Omb) restricted ap expression in the ventral compartment. Loss of omb induced autonomous initiation of ap expression in the middle third-instar larvae in the ventral compartment. Oppositely, over-activation of omb inhibited ap in the medial pouch. All three enhancers apE, apDV and apP were upregulated in omb null mutants, indicating a combinational regulation of ap modulators. However, Omb affected ap expression neither by directly regulating EGFR signaling, nor via Vg regulation. Therefore, a genetic screen of epigenetic regulators, including the Trithorax group (TrxG) and Polycomb group (PcG) genes was performed. We found that knocking down the TrxG gene kohtalo (kto), domino (dom) or expressing the PcG gene grainy head (grh), the ectopic ap in omb mutants was repressed. The inhibition of apDV by kto knockdown and grh activation could contribute to ap repression. Moreover, Omb and the EGFR pathway are genetically parallel in ap regulation in the ventral compartment. Collectively, Omb is a repressive signal for ap expression in the ventral compartment, which requires TrxG and PcG genes.

PMID:36907311 | DOI:10.1016/j.ydbio.2023.03.002

Genes (Basel). 2023 Jan 24;14(2):304. doi: 10.3390/genes14020304.

ABSTRACT

How complex morphological patterns form is an intriguing question in developmental biology. However, the mechanisms that generate complex patterns remain largely unknown. Here, we sought to identify the genetic mechanisms that regulate the tan (t) gene in a multi-spotted pigmentation pattern on the abdomen and wings of Drosophila guttifera. Previously, we showed that yellow (y) gene expression completely prefigures the abdominal and wing pigment patterns of this species. In the current study, we demonstrate that the t gene is co-expressed with the y gene in nearly identical patterns, both transcripts foreshadowing the adult abdominal and wing melanin spot patterns. We identified cis-regulatory modules (CRMs) of t, one of which drives reporter expression in six longitudinal rows of spots on the developing pupal abdomen, while the second CRM activates the reporter gene in a spotted wing pattern. Comparing the abdominal spot CRMs of y and t, we found a similar composition of putative transcription factor binding sites that are thought to regulate the complex expression patterns of both terminal pigmentation genes y and t. In contrast, the y and t wing spots appear to be regulated by distinct upstream factors. Our results suggest that the D. guttifera abdominal and wing melanin spot patterns have been established through the co-regulation of y and t, shedding light on how complex morphological traits may be regulated through the parallel coordination of downstream target genes.

PMID:36833231 | DOI:10.3390/genes14020304

Front Genet. 2023 Feb 7;14:1135320. doi: 10.3389/fgene.2023.1135320. eCollection 2023.

NO ABSTRACT

PMID:36824436 | PMC:PMC9941176 | DOI:10.3389/fgene.2023.1135320

Cell Genom. 2022 Nov 9;2(11):100191. doi: 10.1016/j.xgen.2022.100191. Epub 2022 Oct 5.

ABSTRACT

Gene expression is controlled by transcription factors (TFs) that bind cognate DNA motif sequences in cis-regulatory elements (CREs). The combinations of DNA motifs acting within homeostasis and disease, however, are unclear. Gene expression, chromatin accessibility, TF footprinting, and H3K27ac-dependent DNA looping data were generated and a random-forest-based model was applied to identify 7,531 cell-type-specific cis-regulatory modules (CRMs) across 15 diploid human cell types. A co-enrichment framework within CRMs nominated 838 cell-type-specific, recurrent heterotypic DNA motif combinations (DMCs), which were functionally validated using massively parallel reporter assays. Cancer cells engaged DMCs linked to neoplasia-enabling processes operative in normal cells while also activating new DMCs only seen in the neoplastic state. This integrative approach identifies cell-type-specific cis-regulatory combinatorial DNA motifs in diverse normal and diseased human cells and represents a general framework for deciphering cis-regulatory sequence logic in gene regulation.

PMID:36742369 | PMC:PMC9894309 | DOI:10.1016/j.xgen.2022.100191

Chromosome Res. 2023 Jan 31;31(1):7. doi: 10.1007/s10577-023-09716-w.

ABSTRACT

The homeotic genes or Hox define the anterior-posterior (AP) body axis formation in bilaterians and are often present on the chromosome in an order collinear to their function across the AP axis. However, there are many cases wherein the Hox are not collinear, but their expression pattern is conserved across the AP axis. The expression pattern of Hox is attributed to the cis-regulatory modules (CRMs) consisting of enhancers, initiators, or repressor elements that regulate the genes in a segment-specific manner. In the Drosophila melanogaster Hox complex, the bithorax complex (BX-C) and even the CRMs are organized in an order that is collinear to their function in the thoracic and abdominal segments. In the present study, the regulatorily inert regions were targeted using CRISPR/Cas9 to generate a series of transgenic lines with the insertion of FRT sequences. These FRT lines are repurposed to shuffle the CRMs associated with Abd-B to generate modular deletion, duplication, or inversion of multiple CRMs. The rearrangements yielded entirely novel phenotypes in the fly suggesting the requirement of such complex manipulations to address the significance of higher order arrangement of the CRMs. The functional map and the transgenic flies generated in this study are important resources to decipher the collective ability of multiple regulatory elements in the eukaryotic genome to function as complex modules.

PMID:36719476 | DOI:10.1007/s10577-023-09716-w

PLoS One. 2022 Dec 19;17(12):e0279061. doi: 10.1371/journal.pone.0279061. eCollection 2022.

NO ABSTRACT

PMID:36534652 | DOI:10.1371/journal.pone.0279061

Comput Biol Med. 2022 Nov 29;152:106375. doi: 10.1016/j.compbiomed.2022.106375. Online ahead of print.

NO ABSTRACT

PMID:36502693 | DOI:10.1016/j.compbiomed.2022.106375