scholarly journals Genetic reagents for making split-GAL4 lines in Drosophila

2017 ◽  
Author(s):  
Heather Dionne ◽  
Karen L. Hibbard ◽  
Amanda Cavallaro ◽  
Jui-Chun Kao ◽  
Gerald M. Rubin
Keyword(s):  
Nervous System ◽  
Genomic Dna ◽  
Expression Patterns ◽  
Cell Types ◽  
Biological Processes ◽  
Single Segment ◽  
Distinct Cell ◽  
Experimental Tool ◽  
Transgenic Lines

AbstractThe ability to reproducibly target expression of transgenes to small, defined subsets of cells is a key experimental tool for understanding many biological processes. The Drosophila nervous system contains thousands of distinct cell types and it has generally not been possible to limit expression to one or a few cell types when using a single segment of genomic DNA as an enhancer to drive expression. Intersectional methods, in which expression of the transgene only occurs where two different enhancers overlap in their expression patterns, can be used to achieve the desired specificity. This report describes a set of over 2,800 transgenic lines for use with the split-GAL4 intersectional method.

scholarly journals Probing spermatogenesis in Drosophila with P-element enhancer detectors

Development ◽  
1992 ◽  
Vol 114 (1) ◽  
pp. 89-98 ◽  
Author(s):  
P. Gonczy ◽  
S. Viswanathan ◽  
S. DiNardo
Keyword(s):  
Germ Line ◽  
Expression Patterns ◽  
Cell Types ◽  
P Element ◽  
Structural Reorganization ◽  
Germ Cell Differentiation ◽  
Distinct Cell ◽  
Morphological Criteria ◽  
Cyst Cells ◽  
A Cell

Formation of motile sperm in Drosophila melanogaster requires the coordination of processes such as stem cell division, mitotic and meiotic control and structural reorganization of a cell. Proper execution of spermatogenesis entails the differentiation of cells derived from two distinct embryonic lineages, the germ line and the somatic mesoderm. Through an analysis of homozygous viable and fertile enhancer detector lines, we have identified molecular markers for the different cell types present in testes. Some lines label germ cells or somatic cyst cells in a stage-specific manner during their differentiation program. These expression patterns reveal transient identities for the cyst cells that had not been previously recognized by morphological criteria. A marker line labels early stages of male but not female germ cell differentiation and proves useful in the analysis of germ line sex-determination. Other lines label the hub of somatic cells around which germ line stem cells are anchored. By analyzing the fate of the somatic hub in an agametic background, we show that the germ line plays some role in directing its size and its position in the testis. We also describe how marker lines enable us to identify presumptive cells in the embryonic gonadal mesoderm before they give rise to morphologically distinct cell types. Finally, this collection of marker lines will allow the characterization of genes expressed either in the germ line or in the soma during spermatogenesis.

Developmentally distinct MYB genes encode functionally equivalent proteins in Arabidopsis

Development ◽  
2001 ◽  
Vol 128 (9) ◽  
pp. 1539-1546 ◽  
Author(s):  
M.M. Lee ◽  
J. Schiefelbein
Keyword(s):  
Functional Relationship ◽  
Expression Patterns ◽  
Cell Types ◽  
Myb Transcription Factor ◽  
Regulatory Sequences ◽  
Myb Gene ◽  
Distinct Cell ◽  
Transcription Factor Genes ◽  
Myb Genes ◽  
Multicellular Organisms

The duplication and divergence of developmental control genes is thought to have driven morphological diversification during the evolution of multicellular organisms. To examine the molecular basis of this process, we analyzed the functional relationship between two paralogous MYB transcription factor genes, WEREWOLF (WER) and GLABROUS1 (GL1), in Arabidopsis. The WER and GL1 genes specify distinct cell types and exhibit non-overlapping expression patterns during Arabidopsis development. Nevertheless, reciprocal complementation experiments with a series of gene fusions showed that WER and GL1 encode functionally equivalent proteins, and their unique roles in plant development are entirely due to differences in their cis-regulatory sequences. Similar experiments with a distantly related MYB gene (MYB2) showed that its product cannot functionally substitute for WER or GL1. Furthermore, an analysis of the WER and GL1 proteins shows that conserved sequences correspond to specific functional domains. These results provide new insights into the evolution of the MYB gene family in Arabidopsis, and, more generally, they demonstrate that novel developmental gene function may arise solely by the modification of cis-regulatory sequences.

scholarly journals Transcriptional Profiling of Identified Neurons in Leech

2020 ◽  
Author(s):  
Elizabeth Heath-Heckman ◽  
Shinja Yoo ◽  
Christopher Winchell ◽  
Maurizio Pellegrino ◽  
James Angstadt ◽  
...  
Keyword(s):  
Nervous System ◽  
Transient Receptor Potential ◽  
Transcriptional Profiling ◽  
Expression Patterns ◽  
Gene Families ◽  
Cell Types ◽  
Sensory Transduction ◽  
System Structure ◽  
Identified Neurons ◽  
Hcn Channels

ABSTRACTWhile leeches in the genus Hirudo have long been models for neurobiology, the molecular underpinnings of nervous system structure and function in this group remain largely unknown. To begin to bridge this gap, we performed RNASeq on pools of identified neurons of the central nervous system (CNS): sensory T (touch), P (pressure) and N (nociception) neurons; neurosecretory Retzius cells; and ganglia from which these four cell types had been removed. Bioinformatic analyses identified 2,812 putative genes whose expression differed significantly among the samples. These genes clustered into 7 groups which could be associated with one or more of the identified cell types. We verified predicted expression patterns through in situ hybridization on whole CNS ganglia, and found that orthologous genes were for the most part similarly expressed in a divergent leech genus, suggesting evolutionarily conserved roles for these genes. Transcriptional profiling allowed us to identify candidate phenotype-defining genes from expanded gene families. Thus, we identified one of eight hyperpolarization-activated cyclic-nucleotide gated (HCN) channels as a candidate for mediating the prominent sag current in P neurons, and found that one of five inositol triphosphate receptors (IP3Rs), representing a sub-family of IP3Rs absent from vertebrate genomes, is expressed with high specificity in T cells. We also identified one of two piezo genes, two of ~65 deg/enac genes, and one of at least 16 transient receptor potential (trp) genes as prime candidates for involvement in sensory transduction in the three distinct classes of leech mechanosensory neurons.

scholarly journals Protein network analysis reveals selectively vulnerable regions and biological processes in FTD

2018 ◽  
Vol 4 (5) ◽  
pp. e266 ◽  
Author(s):  
Luke W. Bonham ◽  
Natasha Z.R. Steele ◽  
Celeste M. Karch ◽  
Claudia Manzoni ◽  
Ethan G. Geier ◽  
...  
Keyword(s):  
Association Studies ◽  
Expression Patterns ◽  
Interaction Network ◽  
Analytical Techniques ◽  
Cell Types ◽  
Brain Regions ◽  
Protein Network ◽  
Genome Wide Association Studies ◽  
Biological Processes ◽  
Human Brain Tissue

ObjectiveThe neuroanatomical profile of behavioral variant frontotemporal dementia (bvFTD) suggests a common biological etiology of disease despite disparate pathologic causes; we investigated the genetic underpinnings of this selective regional vulnerability to identify new risk factors for bvFTD.MethodsWe used recently developed analytical techniques designed to address the limitations of genome-wide association studies to generate a protein interaction network of 63 bvFTD risk genes. We characterized this network using gene expression data from healthy and diseased human brain tissue, evaluating regional network expression patterns across the lifespan as well as the cell types and biological processes most affected in bvFTD.ResultsWe found that bvFTD network genes show enriched expression across the human lifespan in vulnerable neuronal populations, are implicated in cell signaling, cell cycle, immune function, and development, and are differentially expressed in pathologically confirmed frontotemporal lobar degeneration cases. Five of the genes highlighted by our differential expression analyses, BAIAP2, ERBB3, POU2F2, SMARCA2, and CDC37, appear to be novel bvFTD risk loci.ConclusionsOur findings suggest that the cumulative burden of common genetic variation in an interacting protein network expressed in specific brain regions across the lifespan may influence susceptibility to bvFTD.

Hox-7 expression during murine craniofacial development

Development ◽  
1991 ◽  
Vol 113 (2) ◽  
pp. 601-611 ◽  
Author(s):  
A. MacKenzie ◽  
M.W. Ferguson ◽  
P.T. Sharpe
Keyword(s):  
Nervous System ◽  
Choroid Plexus ◽  
Expression Pattern ◽  
Neural Tube ◽  
Anterior Pituitary ◽  
Expression Patterns ◽  
In Situ Hybridisation ◽  
Cell Types ◽  
Subsequent Formation ◽  
Dorsal Midline

We have used in situ hybridisation to establish the temporal and spatial expression patterns of the mouse homeobox-containing gene; Hox-7, in the developing embryonic cranium and nervous system of the mouse between embryonic days 9.5 (E9.5) and E15.5. Hox-7 has previously been associated with areas of mesenchymal-epithelial interaction and cell migration especially in neural crest ectomesenchymal cells. Aside from the expression patterns seen in the facial anlage at E9.5, Hox-7 transcripts were also detected in the neuroepithelium including cells of the dorsal midline of the neural tube. This expression pattern persisted throughout the embryonic time span studied. At E11.5, expression of Hox-7 became obvious in the neuroepithelium of the forming tela choroida and the telencephelii in areas destined to form the choroid plexus before any atrophy of the neuroepithelium took place. High expression of Hox-7 was also present in the mesenchyme cells invading the pouch formed by the involuting choroid plexus neuroepithelium. A second major site where Hox-7 was expressed was the anlage of the anterior pituitary; the Rathke's pouch. Expression became obvious at E10.5 throughout the pouch but by E12.5 became more regionalised in areas of the pouch destined to form the pars distalis. Hox-7 was also expressed in the forming meninges and skull bone precursors from E10.5 onwards. Expression of the Hox-7 gene is also seen in the external ear, the forming eye, the nasal pits and forming Jacobson's organs. When these expression patterns are considered together with characterised human and mouse retinoic acid embryopathies and the congenital malformations seen in human children associated with deletion of chromosome 4p16.1 (Wolf-Hirschhorn syndrome), Hox-7 may be a good candidate as one of the genes involved in the initiation of the choroid plexus phenotype and its subsequent formation, the formation of the outer ear, formation of the dentition and the differentiation of the cell types of the anterior pituitary. The expression pattern of Hox-7 in the dorsal midline of the neural tube further suggests that it may also be involved in the specification of the dorsal-ventral axis of the developing nervous system.

scholarly journals Selective interaction of peripheral and central nervous system cells with two distinct cell-binding domains of fibronectin.

1987 ◽  
Vol 105 (3) ◽  
pp. 1435-1442 ◽  
Author(s):  
S L Rogers ◽  
P C Letourneau ◽  
B A Peterson ◽  
L T Furcht ◽  
J B McCarthy
Keyword(s):  
Nervous System ◽  
Cell Attachment ◽  
Cell Types ◽  
Cell Interaction ◽  
Cell Binding ◽  
Domain Specific ◽  
Binding Domains ◽  
Distinct Cell ◽  
Initial Adhesion ◽  
Fibronectin Domains

Mechanisms of cell interaction with fibronectin have been studied with proteolytic fibronectin fragments that have well-defined ligand binding properties. Results of a previous study (Rogers, S. L., J. B. McCarthy, S. L. Palm, L. T. Furcht, and P. C. Letourneau, 1985, J. Neurosci., 5:369-378) demonstrated that (a) central (CNS) and peripheral (PNS) nervous system neurons adhere to, and extend neurites on a 33-kD carboxyl terminal fibronectin fragment that also binds heparin, and (b) neurons from the PNS, but not the CNS, have stable interactions with a 75-kD cell-binding fragment and with intact fibronectin. In the present study domain-specific reagents were used in inhibition assays to further differentiate cell surface interactions with the two fibronectin domains, and to define the significance of these domains to cell interactions with the intact fibronectin molecule. These reagents are (a) a soluble synthetic tetrapeptide Arg-Gly-Asp-Ser (RGDS; Pierschbacher, M. D., and E. Ruoslahti, 1984, Nature (Lond.), 309:30-33) representing a cell-binding determinant in the 75-kD fragment, and (b) an antibody raised against the 33-kD fragment that binds specifically to that fragment. Initial cell attachment to, and neurite extension upon, fibronectin and the two different fragments was evaluated in the presence and absence of the two reagents. Attachment of both PNS and CNS cells to intact fibronectin was reduced in the presence of RGDS, the former more so than the latter. In contrast, the antibody to the 33-kD fragment did not affect attachment of PNS cells to fibronectin, but significantly decreased attachment of CNS cells to the molecule. RGDS inhibited attachment of CNS cells to the molecule. RGDS inhibited attachment of both cell types to the 75-kD fragment to a greater degree than it did attachment to the intact molecule. Cell interaction with the 33-kD fragment was not affected by RGDS. Reduction of neurite lengths (determined after 24 h of culture) by the domain-specific reagents paralleled the reduction in initial adhesion to each substratum. Therefore, it appears that (a) both PNS and CNS cells have receptors for each cell-binding domain of fibronectin, (b) the receptor(s) for the two domains are distinct, with attachment to the 33-kD fragment being independent of RGDS, and (c) the relative importance of each domain to cell interaction with intact fibronectin is different for CNS and PNS cells.

scholarly journals Transcriptional profiling of identified neurons in leech

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Elizabeth Heath-Heckman ◽  
Shinja Yoo ◽  
Christopher Winchell ◽  
Maurizio Pellegrino ◽  
James Angstadt ◽  
...  
Keyword(s):  
Nervous System ◽  
Transient Receptor Potential ◽  
Transcriptional Profiling ◽  
Expression Patterns ◽  
Gene Families ◽  
Cell Types ◽  
Sensory Transduction ◽  
System Structure ◽  
Identified Neurons ◽  
Hcn Channels

Abstract Background While leeches in the genus Hirudo have long been models for neurobiology, the molecular underpinnings of nervous system structure and function in this group remain largely unknown. To begin to bridge this gap, we performed RNASeq on pools of identified neurons of the central nervous system (CNS): sensory T (touch), P (pressure) and N (nociception) neurons; neurosecretory Retzius cells; and ganglia from which these four cell types had been removed. Results Bioinformatic analyses identified 3565 putative genes whose expression differed significantly among the samples. These genes clustered into 9 groups which could be associated with one or more of the identified cell types. We verified predicted expression patterns through in situ hybridization on whole CNS ganglia, and found that orthologous genes were for the most part similarly expressed in a divergent leech genus, suggesting evolutionarily conserved roles for these genes. Transcriptional profiling allowed us to identify candidate phenotype-defining genes from expanded gene families. Thus, we identified one of eight hyperpolarization-activated cyclic-nucleotide gated (HCN) channels as a candidate for mediating the prominent sag current in P neurons, and found that one of five inositol triphosphate receptors (IP3Rs), representing a sub-family of IP3Rs absent from vertebrate genomes, is expressed with high specificity in T cells. We also identified one of two piezo genes, two of ~ 65 deg/enac genes, and one of at least 16 transient receptor potential (trp) genes as prime candidates for involvement in sensory transduction in the three distinct classes of leech mechanosensory neurons. Conclusions Our study defines distinct transcriptional profiles for four different neuronal types within the leech CNS, in addition to providing a second ganglionic transcriptome for the species. From these data we identified five gene families that may facilitate the sensory capabilities of these neurons, thus laying the basis for future work leveraging the strengths of the leech system to investigate the molecular processes underlying and linking mechanosensation, cell type specification, and behavior.

scholarly journals Cell type-specific modulation of healthspan by Forkhead family transcription factors in the nervous system

2021 ◽  
Vol 118 (8) ◽  
pp. e2011491118 ◽  
Author(s):  
Ekin Bolukbasi ◽  
Nathaniel S. Woodling ◽  
Dobril K. Ivanov ◽  
Jennifer Adcott ◽  
Andrea Foley ◽  
...  
Keyword(s):  
Nervous System ◽  
Transcription Factors ◽  
Cell Types ◽  
Growth Factor Signaling ◽  
Cell Type ◽  
Model Of Alzheimer’S Disease ◽  
Distinct Cell ◽  
Evolutionarily Conserved ◽  
Cell Type Specific ◽  
Specific Mapping

Reduced activity of insulin/insulin-like growth factor signaling (IIS) increases healthy lifespan among diverse animal species. Downstream of IIS, multiple evolutionarily conserved transcription factors (TFs) are required; however, distinct TFs are likely responsible for these effects in different tissues. Here we have asked which TFs can extend healthy lifespan within distinct cell types of the adult nervous system in Drosophila. Starting from published single-cell transcriptomic data, we report that forkhead (FKH) is endogenously expressed in neurons, whereas forkhead-box-O (FOXO) is expressed in glial cells. Accordingly, we find that neuronal FKH and glial FOXO exert independent prolongevity effects. We have further explored the role of neuronal FKH in a model of Alzheimer’s disease-associated neuronal dysfunction, where we find that increased neuronal FKH preserves behavioral function and reduces ubiquitinated protein aggregation. Finally, using transcriptomic profiling, we identify Atg17, a member of the Atg1 autophagy initiation family, as one FKH-dependent target whose neuronal overexpression is sufficient to extend healthy lifespan. Taken together, our results underscore the importance of cell type-specific mapping of TF activity to preserve healthy function with age.

scholarly journals Mapping Transgene Insertion Sites Reveals Complex Interactions Between Mouse Transgenes and Neighboring Endogenous Genes

2018 ◽  
Author(s):  
Mallory A. Laboulaye ◽  
Xin Duan ◽  
Mu Qiao ◽  
Irene E. Whitney ◽  
Joshua R. Sanes
Keyword(s):  
Integration Site ◽  
Insertion Site ◽  
Cell Types ◽  
Regulatory Elements ◽  
Endogenous Gene ◽  
Complex Interactions ◽  
Distinct Cell ◽  
Transgenic Lines ◽  
Endogenous Genes ◽  
Insertion Sites

ABSTRACTTransgenic mouse lines are routinely employed to label and manipulate distinct cell types. The transgene generally comprises cell-type specific regulatory elements linked to a cDNA encoding a reporter or other proteins. However, off-target expression seemingly unrelated to the regulatory elements in the transgene is often observed, and sometimes suspected to reflect influences related to the site of transgene integration in the genome. To test this hypothesis, we used a proximity ligation-based method, Targeted Locus Amplification (TLA), to map the insertion sites of three well-characterized transgenes that appeared to exhibit insertion site-dependent expression in retina. The nearest endogenous genes to transgenes HB9-GFP, Mito-P, and TYW3 are Cdh6, Fat4 and Khdrbs2, respectively. For two lines, we demonstrate that expression reflects that of the closest endogenous gene (Fat4 and Cdh6), even though the distance between transgene and endogenous gene is 550 and 680 kb, respectively. In all three lines, the transgenes decrease expression of the neighboring endogenous genes. In each case, the affected endogenous gene was expressed in at least some of the cell types that the transgenic line has been used to mark and study. These results provide insights into the effects of transgenes and endogenous genes on each other’s expression, demonstrate that mapping insertion site is valuable for interpreting results obtained with transgenic lines, and indicate that TLA is a reliable method for integration site discovery.

scholarly journals Combined aptamer and transcriptome sequencing of single cells

2017 ◽  
Author(s):  
Cyrille L. Delley ◽  
Leqian Liu ◽  
Maen F. Sarhan ◽  
Adam R. Abate
Keyword(s):  
Single Cell ◽  
Single Cells ◽  
Expression Patterns ◽  
Cell Types ◽  
Single Cell Protein ◽  
Protein Characterization ◽  
Cell Protein ◽  
Surface Binding ◽  
Distinct Cell

AbstractThe transcriptome and proteome encode distinct information that is important for characterizing heterogeneous biological systems. We demonstrate a method to simultaneously characterize the transcriptomes and proteomes of single cells at high throughput using aptamer probes and droplet-based single cell sequencing. With our method, we differentiate distinct cell types based on aptamer surface binding and gene expression patterns. Aptamers provide advantages over antibodies for single cell protein characterization, including rapid, in vitro, and high-purity generation via SELEX, and the ability to amplify and detect them with PCR and sequencing.

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