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The EMBO Journal Jul 2020Semaphorin ligands interact with plexin receptors to contribute to functions in the development of myriad tissues including neurite guidance and synaptic organisation...
Semaphorin ligands interact with plexin receptors to contribute to functions in the development of myriad tissues including neurite guidance and synaptic organisation within the nervous system. Cell-attached semaphorins interact in trans with plexins on opposing cells, but also in cis on the same cell. The interplay between trans and cis interactions is crucial for the regulated development of complex neural circuitry, but the underlying molecular mechanisms are uncharacterised. We have discovered a distinct mode of interaction through which the Drosophila semaphorin Sema1b and mouse Sema6A mediate binding in cis to their cognate plexin receptors. Our high-resolution structural, biophysical and in vitro analyses demonstrate that monomeric semaphorins can mediate a distinctive plexin binding mode. These findings suggest the interplay between monomeric vs dimeric states has a hereto unappreciated role in semaphorin biology, providing a mechanism by which Sema6s may balance cis and trans functionalities.
Topics: Animals; COS Cells; Cell Adhesion Molecules; Chlorocebus aethiops; Drosophila Proteins; Drosophila melanogaster; Mice; Nerve Tissue Proteins; Protein Structure, Quaternary; Semaphorins; Structure-Activity Relationship
PubMed: 32500924
DOI: 10.15252/embj.2019102926 -
Current Topics in Developmental Biology 2020Terminal regions of the early Drosophila embryo are patterned by the highly conserved ERK cascade, giving rise to the nonsegmented terminal structures of the future... (Review)
Review
Terminal regions of the early Drosophila embryo are patterned by the highly conserved ERK cascade, giving rise to the nonsegmented terminal structures of the future larva. In less than an hour, this signaling event establishes several gene expression boundaries and sets in motion a sequence of elaborate morphogenetic events. Genetic studies of terminal patterning discovered signaling components and transcription factors that are involved in numerous developmental contexts and deregulated in human diseases. This review summarizes current understanding of signaling and morphogenesis during terminal patterning and discusses several open questions that can now be rigorously investigated using live imaging, omics, and optogenetic approaches. The anatomical simplicity of the terminal patterning system and its amenability to a broad range of increasingly sophisticated genetic perturbations will continue to make it a premier quantitative model for studying multiple aspects of tissue patterning by dynamically controlled cell signaling pathways.
Topics: Animals; Body Patterning; Drosophila Proteins; Drosophila melanogaster; Embryo, Nonmammalian; Gene Expression Regulation, Developmental; MAP Kinase Signaling System; Transcription Factors
PubMed: 32143743
DOI: 10.1016/bs.ctdb.2019.11.008 -
EMBO Reports Jul 2023We report that preexisting (old) and newly synthesized (new) histones H3 and H4 are asymmetrically partitioned during the division of Drosophila intestinal stem cells...
We report that preexisting (old) and newly synthesized (new) histones H3 and H4 are asymmetrically partitioned during the division of Drosophila intestinal stem cells (ISCs). Furthermore, the inheritance patterns of old and new H3 and H4 in postmitotic cell pairs correlate with distinct expression patterns of Delta, an important cell fate gene. To understand the biological significance of this phenomenon, we expressed a mutant H3T3A to compromise asymmetric histone inheritance. Under this condition, we observe an increase in Delta-symmetric cell pairs and overpopulated ISC-like, Delta-positive cells. Single-cell RNA-seq assays further indicate that H3T3A expression compromises ISC differentiation. Together, our results indicate that asymmetric histone inheritance potentially contributes to establishing distinct cell identities in a somatic stem cell lineage, consistent with previous findings in Drosophila male germline stem cells.
Topics: Animals; Drosophila; Histones; Intestines; Cell Differentiation; Drosophila Proteins; Cell Division
PubMed: 37255015
DOI: 10.15252/embr.202256404 -
Autophagy May 2024The selective clearance of unwanted, damaged or dangerous components by macroautophagy/autophagy is critical for maintaining cellular homeostasis in organisms from yeast...
The selective clearance of unwanted, damaged or dangerous components by macroautophagy/autophagy is critical for maintaining cellular homeostasis in organisms from yeast to humans. In recent years, significant progress has been made in understanding how phagophores selectively sequester specific cargo. Nevertheless, a fundamental question remains: Can distinct selective autophagy programs simultaneously operate within the same cell? A recent study from the Baehrecke lab has unveiled a developmentally programmed Pink1-dependent reticulophagy process in the Drosophila intestine. Furthermore, this study demonstrated that autophagy differentially clears mitochondria and ER in the same cell under the regulation of Pink1 through different E3 ubiquitin ligases, highlighting the need for further exploration in understanding the complexity of autophagic substrate selection and crosstalk between diverse autophagy programs.
Topics: Ubiquitin-Protein Ligases; Mitophagy; Animals; Humans; Drosophila Proteins; Autophagy; Protein Serine-Threonine Kinases; Mitochondria; Drosophila melanogaster
PubMed: 38456640
DOI: 10.1080/15548627.2024.2323294 -
Proceedings of the National Academy of... Jun 2023The split-Gal4 system allows for intersectional genetic labeling of highly specific cell types and tissues in . However, the existing split-Gal4 system, unlike the...
The split-Gal4 system allows for intersectional genetic labeling of highly specific cell types and tissues in . However, the existing split-Gal4 system, unlike the standard Gal4 system, cannot be repressed by Gal80, and therefore cannot be controlled temporally. This lack of temporal control precludes split-Gal4 experiments in which a genetic manipulation must be restricted to specific timepoints. Here, we describe a split-Gal4 system based on a self-excising split-intein, which drives transgene expression as strongly as the current split-Gal4 system and Gal4 reagents, yet which is repressible by Gal80. We demonstrate the potent inducibility of "split-intein Gal4" in vivo using both fluorescent reporters and via reversible tumor induction in the gut. Further, we show that our split-intein Gal4 can be extended to the drug-inducible GeneSwitch system, providing an independent method for intersectional labeling with inducible control. We also show that the split-intein Gal4 system can be used to generate highly cell type-specific genetic drivers based on in silico predictions generated by single-cell RNAseq (scRNAseq) datasets, and we describe an algorithm ("Two Against Background" or TAB) to predict cluster-specific gene pairs across multiple tissue-specific scRNA datasets. We provide a plasmid toolkit to efficiently create split-intein Gal4 drivers based on either CRISPR knock-ins to target genes or using enhancer fragments. Altogether, the split-intein Gal4 system allows for the creation of highly specific intersectional genetic drivers that are inducible/repressible.
Topics: Animals; Transcription Factors; Inteins; Drosophila; Protein Splicing; Transgenes; Drosophila Proteins
PubMed: 37276389
DOI: 10.1073/pnas.2304730120 -
Genes & Development Jul 2021The prototypic and ubiquitous microtubule motor, kinesin-1, uses a variety of adaptor proteins to facilitate the selective transport of diverse cargo within the cell.... (Review)
Review
The prototypic and ubiquitous microtubule motor, kinesin-1, uses a variety of adaptor proteins to facilitate the selective transport of diverse cargo within the cell. These cargo adaptors bind to the motor complex through interactions with the kinesin light or heavy chains (KLCs or KHCs). In this issue of , Dimitrova-Paternoga et al. (pp. 976-991) present the first structural characterization of a KHC-cargo adaptor interface. They describe an antiparallel heterotrimeric coiled-coil complex between the carboxy tail of KHC and Tm1-I/C (Tm1), the atypical tropomyosin that is important for mRNA transport in oocytes. This interaction enhances direct binding between KHC and RNA. Their findings demonstrate the structural plasticity of the KHC tail as a platform for protein-protein interactions and reveal how a cargo adaptor protein can modify a motor-RNA interface to promote transport.
Topics: Animals; Drosophila; Drosophila Proteins; Kinesins; Microtubules; RNA
PubMed: 34210804
DOI: 10.1101/gad.348691.121 -
Developmental Biology Apr 2022The ability of stem cells to divide asymmetrically is crucial for cell-type diversity and tissue homeostasis. Drosophila neural stem cells, also knowns as neuroblasts,... (Review)
Review
The ability of stem cells to divide asymmetrically is crucial for cell-type diversity and tissue homeostasis. Drosophila neural stem cells, also knowns as neuroblasts, utilize asymmetric cell division to self-renew and give rise to differentiated daughter cells. Drosophila neuroblasts relies on the polarized protein complexes on the apical and basal cortex to govern cell polarity and asymmetry. Here, we review recent advances in our understanding of the neuroblast polarity focusing on how actin cytoskeleton, phosphoinositide lipids and liquid-liquid phase separation regulate the asymmetric cell division of Drosophila neuroblasts.
Topics: Animals; Cell Division; Drosophila; Drosophila Proteins; Neural Stem Cells; Neurons
PubMed: 35181298
DOI: 10.1016/j.ydbio.2022.02.006 -
PloS One 2021During formation of the Hedgehog (Hh) signaling proteins, cooperative activities of the Hedgehog INTein (Hint) fold and Sterol Recognition Region (SRR) couple...
During formation of the Hedgehog (Hh) signaling proteins, cooperative activities of the Hedgehog INTein (Hint) fold and Sterol Recognition Region (SRR) couple autoproteolysis to cholesterol ligation. The cholesteroylated Hh morphogens play essential roles in embryogenesis, tissue regeneration, and tumorigenesis. Despite the centrality of cholesterol in Hh function, the full structure of the Hint-SRR ("Hog") domain that attaches cholesterol to the last residue of the active Hh morphogen remains enigmatic. In this work, we combine molecular dynamics simulations, photoaffinity crosslinking, and mutagenesis assays to model cholesterolysis intermediates in the human Sonic Hedgehog (hSHH) protein. Our results provide evidence for a hydrophobic Hint-SRR interface that forms a dynamic, non-covalent cholesterol-Hog complex. Using these models, we suggest a unified mechanism by which Hh proteins can recruit, sequester, and orient cholesterol, and offer a molecular basis for the effects of disease-causing hSHH mutations.
Topics: Animals; Cholesterol; Drosophila Proteins; Drosophila melanogaster; HEK293 Cells; Hedgehog Proteins; Humans; Molecular Dynamics Simulation; Protein Domains
PubMed: 33630857
DOI: 10.1371/journal.pone.0246814 -
Proteins Jun 2024Understanding the emergence and structural characteristics of de novo and random proteins is crucial for unraveling protein evolution and designing novel enzymes....
Understanding the emergence and structural characteristics of de novo and random proteins is crucial for unraveling protein evolution and designing novel enzymes. However, experimental determination of their structures remains challenging. Recent advancements in protein structure prediction, particularly with AlphaFold2 (AF2), have expanded our knowledge of protein structures, but their applicability to de novo and random proteins is unclear. In this study, we investigate the structural predictions and confidence scores of AF2 and protein language model-based predictor ESMFold for de novo and conserved proteins from Drosophila and a dataset of comparable random proteins. We find that the structural predictions for de novo and random proteins differ significantly from conserved proteins. Interestingly, a positive correlation between disorder and confidence scores (pLDDT) is observed for de novo and random proteins, in contrast to the negative correlation observed for conserved proteins. Furthermore, the performance of structure predictors for de novo and random proteins is hampered by the lack of sequence identity. We also observe fluctuating median predicted disorder among different sequence length quartiles for random proteins, suggesting an influence of sequence length on disorder predictions. In conclusion, while structure predictors provide initial insights into the structural composition of de novo and random proteins, their accuracy and applicability to such proteins remain limited. Experimental determination of their structures is necessary for a comprehensive understanding. The positive correlation between disorder and pLDDT could imply a potential for conditional folding and transient binding interactions of de novo and random proteins.
Topics: Animals; Protein Folding; Conserved Sequence; Drosophila Proteins; Databases, Protein; Models, Molecular; Computational Biology; Proteins; Intrinsically Disordered Proteins; Protein Conformation; Amino Acid Sequence; Algorithms; Drosophila
PubMed: 38226524
DOI: 10.1002/prot.26652 -
Biochimica Et Biophysica Acta. Gene... Jul 2020The Cyclin B1 gene encodes a G2/M cyclin that is deregulated in various human cancers, however, the transcriptional regulation of this gene is incompletely understood....
The Cyclin B1 gene encodes a G2/M cyclin that is deregulated in various human cancers, however, the transcriptional regulation of this gene is incompletely understood. The E2F and retinoblastoma family of proteins are involved in this gene's regulation, but there is disagreement on which of the E2F and retinoblastoma proteins interact with the promoter to regulate this gene. Here, we dissect the promoter region of the Drosophila CycB gene, and study the role of Rbf and E2F factors in its regulation. This gene exhibits remarkable features that distinguish it from G1/S regulated promoters, such as PCNA. The promoter is comprised of modular elements with dedicated repressor and activator functions, including a segment spanning the first intron that interferes with a 5' activator element. A highly active minimal promoter (-464, +100) is repressed by the Rbf1 retinoblastoma protein, but much more potently repressed by the Rbf2 protein, which has been linked in other studies to control of cell growth genes. Unlike many other cell-cycle genes, which are activated by E2F1 and repressed by E2F2, CycB is potently activated by E2F2, and repressed by E2F1. Although the bulk of Rbf binding is associated with a region 5' of the core promoter, E2F and retinoblastoma proteins functionally interact with the basal promoter region, in part through a conserved E2F site at -80 bp. The specific regulatory requirements of this late cell cycle promoter appear to be linked to the unique activities of E2F and retinoblastoma family members acting on a complex cis-regulatory circuit.
Topics: Animals; Cell Line; Conserved Sequence; Cyclin B; Drosophila Proteins; Drosophila melanogaster; Introns; Promoter Regions, Genetic; Protein Binding; Retinoblastoma Protein; Transcription Factors; Transcriptional Activation
PubMed: 32275964
DOI: 10.1016/j.bbagrm.2020.194549