Biology subjects

Sim, M.

Publications and source records attributed to Sim, M..

4 recordsLinked to original sources

bioRxiv2025

Social isolation upregulates takeout expression in female Drosophila melanogaster to promote sucrose feeding

Drosophila melanogaster provides a model system to examine how environmental stress interacts with sex to induce changes in brain function and behavior. Previous research suggests that social isolation induces changes in gene expression that encode a starvation-like brain state and reduce sleep. However, the extent to which social isolation alters behaviors via sex-specific brain changes is unclear. Here, we use Drosophila melanogaster to explore sex differences in chronic social isolation-induced behavioral and transcriptomic changes. We focused on takeout (to), a gene encoding a putative juvenile hormone-binding protein, as a target that is upregulated solely in females following social isolation. Male and female adult flies were exposed to chronic social isolation, and multiple behavioral sex differences were identified through tests of activity, motivation, aggression, and sugar consumption. RNA-seq analysis also identified several candidate genes that were associated with sex differences in isolation-induced behavioral changes. Our findings suggest that social isolation is sufficiently stressful to reveal latent sex differences in behavior, despite having no impact on survival. To expression and sucrose consumption were upregulated exclusively in females following social isolation. Following to knockdown in to-expressing cells, sucrose consumption decreased in socially isolated females but increased in males. However, to knock down, to overexpression, and transformer knock down in neurons did not change sucrose-feeding behavior between control and isolated females. Overall, our results suggest that manipulating to expression influences sucrose-feeding in opposite directions between females and males following social isolation, and that isolation-induced to overexpression in non-neuronal cells in the brain or head may play a role in communicating information about females nutritional status to the brain. Additional roles for to in stress-related behaviors and behavioral sex differences should be explored, as well as whether to participates in signaling pathways that may be functionally conserved in human disorders. Author SummaryChronic stress contributes to detrimental health effects, but our understanding of how stress induces sex differences in brain gene expression and behavior is incomplete. Here, we use a combination of behavioral testing, RNA sequencing, and genetic manipulations in Drosophila melanogaster to explore how social isolation reveals latent sex differences in gene expression and stress-relevant behaviors. We found the most pronounced sex differences in behaviors related to feeding and motivation. RNA profiling revealed isolated female-specific upregulation of over 100 genes, with many of them relating to reproduction and energy metabolism. We manipulated expression of the candidate gene takeout (to) and found that downregulating to in all to-expressing cells decreases sucrose-feeding in isolated females but increases it in isolated males. Our results suggest that within a chronic stress context, sex-specific effectors in the head may regulate gene expression related to feeding and macronutrient choice to ensure that females prioritize survival over reproduction. Learning more about this system in flies could provide insight into functionally analogous pathways in humans that may be dysregulated in female-biased stress-related disorders.

neuroscience↗

bioRxiv2024

A highly conserved sRNA downregulates multiple genes, including a sigma54 transcriptional activator, in the virulence mode of Bordetella pertussis

Bacterial sRNAs together with the RNA chaperone Hfq post-transcriptionally regulate gene expression by affecting ribosome binding or mRNA stability. In the human pathogen Bordetella pertussis, the causative agent of whooping cough, hundreds of sRNAs have been identified, but their roles in B. pertussis biology are mostly unknown. Here we characterize a Hfq-dependent sRNA (S17), whose level is dramatically higher in the virulence (Bvg+) mode. We show that transcription from a {sigma}A-dependent promoter yields a long form of 190 nucleotides (nts) that is processed by RNase E to generate a shorter, more stable form (S17S) of 67 nts. Using RNA-seq and RT-qPCR, we identify 92 genes whose expression significantly increases in the absence of S17. Of these genes, 70 contain sequences at/near their ribosome binding sites (RBSs) that are complementary to single-stranded (ss) regions (Sites 1 or 2) of S17S. The identified genes include those encoding multiple transporters and 3 transcriptional regulators. Using a lacZ translational reporter system, we demonstrate that S17S directly represses one of these genes, BP2158, a {sigma}54- dependent transcriptional regulator, suggesting the repression of a {sigma}54 regulon in the Bvg+ mode. We find that the S17S region containing Sites 1 and 2 is 100% conserved throughout various Betaproteobacteria species, and the S17S target sites are often conserved in the homologs of the B. pertussis target genes. We speculate that S17S regulation represents a highly conserved process that fine-tunes gene expression in the Bvg+ mode of B. pertussis and perhaps under other conditions in related bacteria. IMPORTANCERegulation of gene expression involves controlling transcription, translation, and transcript degradation. sRNAs with short sequences complementary to an mRNA sequence are involved in post-transcriptional regulation by aiding or interfering with either ribosome binding or nuclease attack. In the human pathogen Bordetella pertussis, the causative agent of whooping cough, hundreds of sRNA have been identified, but their functions remain largely unknown. We have characterized a sRNA that is abundant in the virulence mode of B. pertussis and serves to down-regulate multiple genes including transcriptional regulators and various transporters. We demonstrate that this sRNA directly represses a transcriptional factor, suggesting that it influences the regulation of specific B. pertussis regulons. The high conservation of this sRNA and its targets within Betaproteobacteria suggests a conserved pathway for gene regulation.

microbiology↗

bioRxiv2020

Codon optimisation for maximising gene expression in multiple species and microbial consortia

MotivationCodon optimisation, the process of adapting the codon composition of a coding sequence, is often used in synthetic biology to increase expression of a heterologous protein. Recently, a number of synthetic biology approaches that allow synthetic constructs to be deployed in multiple organisms have been published. However, so far, design tools for codon optimisation have not been updated to reflect these new approaches. ApproachWe designed an evolutionary algorithm (EA) to design coding sequences (CDSs) that encode a target protein for one or more target organisms, based on the Chimera average repetitive substring (ARS) metric -- a correlate of gene expression. A parameter scan was then used to find optimal parameter sets. Using the optimal parameter sets, three heterologous proteins were repeatedly optimised Bacillus subtilis 168 and Escherichia coli MG1655. The ARS scores of the resulting sequences were compared to the ARS scores of coding sequences that had been optimised for each organism individually (using Chimera Map). ResultsWe demonstrate that an EA is a valid approach to optimising a coding sequence for multiple organisms at once; both crossover and mutation operators were shown to be necessary for the best performance. In some scenarios, the EA generated CDSs that had higher ARS scores than CDSs optimised for the individual organisms, suggesting that the EA exploits the CDS design space in a way that Chimera Map does not. Availability and implementationThe implementation of the EA, with instructions, is available on GitHub: https://github.com/intbio-ncl/chimera_evolve.

synthetic biology↗

bioRxiv2018

Driving the expression of the Salmonella enterica sv Typhimurium flagellum using flhDC from Escherichia coli results in key regulatory and cellular differences.

The flagellar systems of Escherichia coli and Salmonella enterica exhibit a significant level of genetic and functional synteny. Both systems are controlled by the flagellar specific master regulator FlhD4C2. Since the early days of genetic analyses of flagellar systems it has been known that E. coli flhDC can complement a {Delta}flhDC mutant in S. enterica. The genomic revolution has identified how genetic changes to transcription factors and / or DNA binding sites can impact the phenotypic outcome across related species. We were therefore interested in asking: using modern tools to interrogate flagellar gene expression and assembly, what would the impact be of replacing the flhDC coding sequences in S. enterica for the E. coli genes at the flhDC S. entercia chromosomal locus? We show that even though all strains created are motile, flagellar gene expression is measurably lower when flhDCEC are present. These changes can be attributed to the impact of FlhD4C2 DNA recognition and the protein-protein interactions required to generate a stable FlhD4C2 complex. Furthermore, our data suggests that in E. coli the internal flagellar FliT regulatory feedback loop has a marked difference with respect to output of the flagellar systems. We argue due diligence is required in making assumptions based on heterologous expression of regulators and that even systems showing significant synteny may not behave in exactly the same manner.

microbiology↗