Biology subjects

Southall, T. D.

Publications and source records attributed to Southall, T. D..

8 recordsLinked to original sources

bioRxiv2025

Sex-lethal is recruited to chromatin to promote neuronal tRNA synthesis in males through RNA Polymerase III regulation

The RNA-binding protein Sex-lethal (Sxl) is classically known as a master regulator of sex determination and mRNA splicing in Drosophila melanogaster. However, this role is not conserved across species, and functions beyond this canonical pathway remain poorly understood. In this study, we uncover a splicing-independent role for Sxl at the chromatin level in the Drosophila brain. Using Targeted DamID (TaDa) profiling in neurons, we identify widespread recruitment of Sxl to promoter regions, independent of sex or RNA binding activity. Notably, Sxl chromatin occupancy exhibits near-complete overlap with Polr3E (RPC37), an RNA Polymerase III subunit, with Sxl binding abolished upon Polr3E knockdown. Depletion of Sxl in mature male neurons induces widespread transcriptional changes, particularly in metabolic genes, and improves negative geotaxis during ageing, phenotypes that closely mirror Polr3E knockdown. Conversely, overexpression of the brain-specific SxlRAC transcript leads to enhanced tRNA synthesis and upregulated metabolic gene expression. Together, these findings reveal a previously unrecognised role for Sxl in regulating Pol III activity via Polr3E, regulating tRNA synthesis and supporting neuronal metabolism. Given the emerging tie between Pol III regulation and neuronal ageing, our study highlights Sxl as a novel modulator of neuronal homeostasis.

molecular biology↗

bioRxiv2022

A serine-folate metabolic unit controls resistance and tolerance of infection

Immune activation drives metabolic change in most animals. Immune-induced metabolic change is most conspicuous as a driver of pathology in serious or prolonged infection, but it is normally expected to be important to support immune function and recovery. Many of the signalling mechanisms linking immune detection with metabolic regulation, and their specific consequences, are unknown. Here, we show that Drosophila melanogaster respond to many bacterial infections by altering expression of genes of the folate cycle and associated enzymes of amino acid metabolism. The net result of these changes is increased flow of carbon from glycolysis into serine and glycine synthesis and a shift of folate cycle activity from the cytosol into the mitochondrion. Immune-induced transcriptional induction of astray and Nmdmc, the two most-induced of these enzymes, depends on Dif and foxo. Loss of astray or Nmdmc results in infection-specific immune defects. Our work thus shows a key mechanism that connects immune-induced changes in metabolic signalling with the serine-folate metabolic unit to result in changed immune function.

immunology↗

bioRxiv2022

NuRD independent Mi-2 activity represses ectopic gene expression during neuronal maturation.

During neuronal development, extensive changes to chromatin states occur to regulate lineage-specific gene expression. The molecular factors underlying the repression of non-neuronal genes in differentiated neurons are poorly characterised. The Mi2/NuRD complex is a multiprotein complex with nucleosome remodelling and histone deacetylase activity. Whilst NuRD has previously been implicated in the development of nervous system tissues the precise nature of the gene expression programmes that it coordinates are ill-defined. Furthermore, evidence from several species suggests that Mi-2 may be incorporated into multiple complexes that may not possess histone deacetylase activity. We show that Mi-2 activity is required for suppressing ectopic expression of germline genes in neurons independently of HDAC1/NuRD, whilst components of NuRD, including Mi-2, regulate neural gene expression to ensure proper development of the larval nervous system. We find that Mi-2 binding in the genome is dynamic during neuronal maturation and Mi-2 mediated repression of ectopic gene expression is restricted to the early stages of neuronal development, indicating that Mi-2/NuRD is required for establishing stable neuronal transcriptomes during the early stages of neuronal differentiation.

developmental biology↗

bioRxiv2021

AGES: An auxin-inducible, GAL4-compatible, gene expression system for Drosophila

The ability to control transgene expression, both spatially and temporally, is essential for studying model organisms. In Drosophila, spatial control is primarily provided by the GAL4/UAS system, whilst temporal control relies on a temperature-sensitive GAL80 (which inhibits GAL4) and drug-inducible systems. However, these are not ideal. Shifting temperature can impact on many physiological and behavioural traits, and the current drug-inducible systems are either leaky, toxic, incompatible with existing GAL4-driver lines, or do not generate effective levels of expression. Here we describe the Auxin-inducible Gene Expression System (AGES). AGES relies on the auxin-dependent degradation of a ubiquitously expressed GAL80, and therefore, is compatible with existing GAL4-driver lines. Water-soluble auxin is added to fly food at a low, non-lethal, concentration, which induces expression comparable to uninhibited GAL4 expression. The system works in both larvae and adults, providing a stringent, non-lethal, cost-effective, and convenient method for temporally controlling GAL4 activity in Drosophila.

genetics↗

bioRxiv2020

FlyORF-TaDa allows rapid generation of new lines for in vivo cell-type specific profiling of protein-DNA interactions in Drosophila melanogaster

Targeted DamID (TaDa) is an increasingly popular method of generating cell-type specific DNA binding profiles in vivo. Although sensitive and versatile, TaDa requires the generation of new transgenic fly lines for every protein that is profiled, which is both time-consuming and costly. Here, we describe the FlyORF-TaDa system for converting an existing FlyORF library of inducible open reading frames (ORFs) to TaDa lines via a genetic cross, with recombinant progeny easily identifiable by eye colour. Profiling the binding of the H3K36me3-associated chromatin protein MRG15 in larval neural stem cells using both FlyORF-TaDa and conventional TaDa demonstrates that new lines generated using this system provide accurate and highly-reproducible DamID binding profiles. Our data further show that MRG15 binds to a subset of active chromatin domains in vivo. Courtesy of the large coverage of the FlyORF library, the FlyORF-TaDa system enables the easy creation of TaDa lines for 74% of all transcription factors and chromatin modifying proteins within the Drosophila genome.

molecular biology↗

bioRxiv2020

Condensin I subunit Cap-G is essential for proper gene expression during the maturation of post-mitotic neurons

The condensin complex is essential for mitotic chromosome assembly and segregation during cell divisions, however, little is known about its function in post-mitotic, differentiated cells. Here we report a novel role for the condensin I subunit Cap-G in Drosophila neurons. We show that, despite not requiring condensin for mitotic chromosome compaction, post-mitotic neurons express Cap-G and that knockdown of Cap-G specifically in neurons (from their birth onwards) results in developmental arrest, behavioural defects, and dramatic gene expression changes. These include reduced expression of a subset of neuronal genes and aberrant expression of genes that are not normally expressed in the developing brain. Knockdown of Cap-G in more mature neurons also results in similar phenotypes but to a lesser degree. Furthermore, we see dynamic binding of Cap-G to chromatin at distinct loci in neural stem cells and differentiated neurons. Therefore, Cap-G is essential for proper gene expression in neurons and plays an important role during the early stages of neuronal development. O_FIG O_LINKSMALLFIG WIDTH=197 HEIGHT=200 SRC="FIGDIR/small/904409v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@11a48aborg.highwire.dtl.DTLVardef@9912b7org.highwire.dtl.DTLVardef@19bd19dorg.highwire.dtl.DTLVardef@deb7f7_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2019

Dynamic neurotransmitter specific transcription factor expression profiles during Drosophila development

The remarkable diversity of neurons in the nervous system is generated during development, when properties such as cell morphology, receptor profiles and neurotransmitter identities are specified. In order to gain a greater understanding of neurotransmitter specification we profiled the transcription state of cholinergic, GABAergic and glutamatergic neurons in vivo at three developmental time points. We identified 86 differentially expressed transcription factors that are uniquely enriched, or uniquely depleted, in a specific neurotransmitter type. Some transcription factors show a similar profile across development, others only show enrichment or depletion at specific developmental stages. Profiling of Acj6 (cholinergic enriched) and Ets65A (cholinergic depleted) binding sites in vivo reveals that they both directly bind the ChAT locus, in addition to a wide spectrum of other key neuronal differentiation genes. We also show that cholinergic enriched transcription factors are expressed in mostly non-overlapping populations in the adult brain, implying the absence of combinatorial regulation of neurotransmitter fate in this context. Furthermore, our data underlines that, similar to C. elegans, there are no simple transcription factor codes for neurotransmitter type specification.

developmental biology↗

bioRxiv2017

CATaDa reveals global remodelling of chromatin accessibility during stem cell differentiationin vivo

Regulation of eukaryotic gene expression is coordinated by dynamic changes to chromatin states throughout development. Measurements of accessible chromatin are used extensively to identify genomic regulatory elements. Whilst the chromatin landscapes of pluripotent stem cells are well characterised, chromatin accessibility changes in the development of somatic stem cell lineages are not well defined. Here we show that tissue specific chromatin accessibility data can be produced via ectopic expression of E. coli Dam methylase in vivo, without the requirement for cell-sorting. We have profiled chromatin accessibility in individual cell types of the Drosophila neural and midgut stem cell lineages. Functional cell-type specific enhancers were identified, as well as novel motifs enriched at diferent stages of development. Finally, we show global changes in the accessibility of chromatin between stem-cells and their diferentiated progeny. Our results demonstrate the dynamic nature of chromatin accessibility in somatic tissues during stem cell diferentiation and provide a novel approach to understanding the gene regulatory mechanisms underlying development.

developmental biology↗