Regulatory modules mediating the complex neural expression patterns of the homeobrain gene during Drosophila brain development

Abstract Background The homeobox gene homeobrain (hbn) is located in the 57B region together with two other homeobox genes, Drosophila Retinal homeobox (DRx) and orthopedia (otp). All three genes encode transcription factors with important functions in brain development. Hbn mutants are embryonic lethal and characterized by a reduction in the anterior protocerebrum, including the mushroom bodies, and a loss of the supraoesophageal brain commissure. Results In this study we conducted a detailed expression analysis of Hbn in later developmental stages. In the larval brain, Hbn is expressed in all type II lineages and the optic lobes, including the medulla and lobula plug. The gene is expressed in the cortex of the medulla and the lobula rim in the adult brain. We generated a new hbnKOGal4 enhancer trap strain by reintegrating Gal4 in the hbn locus through gene targeting, which reflects the complete hbn expression during development. Eight different enhancer-Gal4 strains covering 12 kb upstream of hbn, the two large introns and 5 kb downstream of the gene, were established and hbn expression was investigated. We characterized several enhancers that drive expression in specific areas of the brain throughout development, from embryo to the adulthood. Finally, we generated deletions of four of these enhancer regions through gene targeting and analysed their effects on the expression and function of hbn. Conclusion The complex expression of Hbn in the developing brain is regulated by several specific enhancers within the hbn locus. Each enhancer fragment drives hbn expression in several specific cell lineages, and with largely overlapping patterns, suggesting the presence of shadow enhancers and enhancer redundancy. Specific enhancer deletion strains generated by gene targeting display developmental defects in the brain. This analysis opens an avenue for a deeper analysis of hbn regulatory elements in the future.

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Functional analysis of enhancer elements regulating the expression of the Drosophila homeodomain transcription factor DRx by gene targeting

Hereditas ◽

10.1186/s41065-021-00210-z ◽

2021 ◽

Vol 158 (1) ◽

Author(s):

Christine Klöppel ◽

Kirsten Hildebrandt ◽

Dieter Kolb ◽

Nora Fürst ◽

Isabelle Bley ◽

...

Keyword(s):

Transcription Factor ◽

Brain Development ◽

Gene Targeting ◽

Developmental Stages ◽

Expression Patterns ◽

Adult Brain ◽

Central Complex ◽

Larval Brain ◽

Endogenous Expression ◽

The Brain

Abstract Background The Drosophila brain is an ideal model system to study stem cells, here called neuroblasts, and the generation of neural lineages. Many transcriptional activators are involved in formation of the brain during the development of Drosophila melanogaster. The transcription factor Drosophila Retinal homeobox (DRx), a member of the 57B homeobox gene cluster, is also one of these factors for brain development. Results In this study a detailed expression analysis of DRx in different developmental stages was conducted. We show that DRx is expressed in the embryonic brain in the protocerebrum, in the larval brain in the DM and DL lineages, the medulla and the lobula complex and in the central complex of the adult brain. We generated a DRx enhancer trap strain by gene targeting and reintegration of Gal4, which mimics the endogenous expression of DRx. With the help of eight existing enhancer-Gal4 strains and one made by our group, we mapped various enhancers necessary for the expression of DRx during all stages of brain development from the embryo to the adult. We made an analysis of some larger enhancer regions by gene targeting. Deletion of three of these enhancers showing the most prominent expression patterns in the brain resulted in specific temporal and spatial loss of DRx expression in defined brain structures. Conclusion Our data show that DRx is expressed in specific neuroblasts and defined neural lineages and suggest that DRx is another important factor for Drosophila brain development.

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Genome-Wide Analysis and Characterization of the Aux/IAA Family Genes Related to Floral Scent Formation in Hedychium coronarium

International Journal of Molecular Sciences ◽

10.3390/ijms20133235 ◽

2019 ◽

Vol 20 (13) ◽

pp. 3235 ◽

Cited By ~ 2

Author(s):

Yanguo Ke ◽

Farhat Abbas ◽

Yiwei Zhou ◽

Rangcai Yu ◽

Yuechong Yue ◽

...

Keyword(s):

Gene Family ◽

Developmental Stages ◽

Floral Scent ◽

Expression Profiles ◽

Expression Patterns ◽

Regulatory Elements ◽

Virus Induced Gene Silencing ◽

Cut Flowers ◽

Flower Opening ◽

Hedychium Coronarium

Auxin plays a key role in different plant growth and development processes, including flower opening and development. The perception and signaling of auxin depend on the cooperative action of various components, among which auxin/indole-3-acetic acid (Aux/IAA) proteins play an imperative role. In a recent study, the entire Aux/IAA gene family was identified and comprehensively analyzed in Hedychium coronarium, a scented species used as an ornamental plant for cut flowers. Phylogenetic analysis showed that the Aux/IAA gene family in H. coronarium is slightly contracted compared to Arabidopsis, with low levels of non-canonical proteins. Sequence analysis of promoters showed numerous cis-regulatory elements related to various phytohormones. HcIAA genes showed distinct expression patterns in different tissues and flower developmental stages, and some HcIAA genes showed significant responses to auxin and ethylene, indicating that Aux/IAAs may play an important role in linking hormone signaling pathways. Based on the expression profiles, HcIAA2, HcIAA4, HcIAA6 and HcIAA12, were selected as candidate genes and HcIAA2 and HcIAA4 were screened for further characterization. Downregulation of HcIAA2 and HcIAA4 by virus-induced gene silencing in H. coronarium flowers modified the total volatile compound content, suggesting that HcIAA2 and HcIAA4 play important roles in H. coronarium floral scent formation. The results presented here will provide insights into the putative roles of HcIAA genes and will assist the elucidation of their precise roles during floral scent formation.

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Human library of cardiac promoters and enhancers

10.1101/2020.06.14.150904 ◽

2020 ◽

Author(s):

Ruslan M. Deviatiiarov ◽

Anna Gams ◽

Roman Syunyaev ◽

Tatiana V. Tatarinova ◽

Oleg Gusev ◽

...

Keyword(s):

Heart Diseases ◽

Association Studies ◽

Expression Patterns ◽

Critical Role ◽

Regulatory Elements ◽

Specific Cell ◽

Genome Wide Association Studies ◽

Specific Expression ◽

Enhancer Activity ◽

Human Donor

AbstractGenome regulatory elements play a critical role during cardiac development and maintenance of normal physiological homeostasis, and genome-wide association studies identified a large number of SNPs associated with cardiovascular diseases localized in intergenic zones. We used cap analysis of gene expression (CAGE) to identify transcription start sites (TSS) with one nucleotide resolution that effectively maps genome regulatory elements in a representative collection of human heart tissues. Here we present a comprehensive and fully annotated CAGE atlas of human promoters and enhancers from four chambers of the non-diseased human donor hearts, including both atria and ventricles. We have identified 10,528 novel regulatory elements, where 2,750 are classified as TSS and 4,258 novel enhancers, which were validated with ChIP-seq libraries and motif enrichment analysis. We found that heart-region specific expression patterns are primarily based on the alternative promoter and specific enhancer activity. Our study significantly increased evidence of the association of regulatory elements-located variants with heart morphology and pathologies. The precise location of cardiac disease-related SNPs within the regulatory regions and their correlation with a specific cell type offers a new understanding of genetic heart diseases.

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MRI of Capn15 knockout mice and analysis of Capn15 distribution reveal possible roles in brain development and plasticity

10.1101/763888 ◽

2019 ◽

Cited By ~ 1

Author(s):

Congyao Zha ◽

Carole A Farah ◽

Vladimir Fonov ◽

David A. Rudko ◽

Wayne S Sossin

Keyword(s):

Brain Development ◽

Knockout Mice ◽

Brain Plasticity ◽

Small Animal ◽

Cell Types ◽

Cysteine Proteases ◽

Brain Regions ◽

Conditional Knockout ◽

Specific Cell ◽

The Brain

AbstractPurposeThe non-classical Small Optic Lobe (SOL) family of calpains are intracellular cysteine proteases that are expressed in the nervous system and appear to play an important role in neuronal development in both Drosophila, where loss of this calpain leads to the eponymous small optic lobes, and in mouse and human, where loss of this calpain (Capn15) leads to eye anomalies. However, the brain regions where this calpain is expressed and the areas most affected by the loss of this calpain have not been carefully examined.ProceduresWe utilize an insert strain where lacZ is expressed under the control of the Capn15 promoter, together with immunocytochemistry with markers of specific cell types to address where Capn 15 is expressed in the brain. We use small animal MRI comparing WT, Capn15 knockout and Capn15 conditional knockout mice to address the brain regions that are affected when Capn 15 is not present, either in early development of the adult.ResultsCapn15 is expressed in diverse brain regions, many of them involved in plasticity such as the hippocampus, lateral amygdala and Purkinje neurons. Capn15 knockout mice have smaller brains, and present specific deficits in the thalamus and hippocampal regions. There are no deficits revealed by MRI in brain regions when Capn15 is knocked out after development.ConclusionsAreas where Capn15 is expressed in the adult are not good markers for the specific regions where the loss of Capn15 specifically affects brain development. Thus, it is likely that this calpain plays distinct roles in brain development and brain plasticity.

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Deletion of lrrk2 causes early developmental abnormalities and age-dependent increase of monoamine catabolism in the zebrafish brain

PLoS Genetics ◽

10.1371/journal.pgen.1009794 ◽

2021 ◽

Vol 17 (9) ◽

pp. e1009794

Author(s):

Stefano Suzzi ◽

Reiner Ahrendt ◽

Stefan Hans ◽

Svetlana A. Semenova ◽

Avinash Chekuru ◽

...

Keyword(s):

Developmental Stages ◽

Biological Activities ◽

Specific Cell ◽

Loss Of Function ◽

Developmental Abnormalities ◽

Age Dependent ◽

Paralog Gene ◽

Early Developmental ◽

The Brain

LRRK2 gain-of-function is considered a major cause of Parkinson’s disease (PD) in humans. However, pathogenicity of LRRK2 loss-of-function in animal models is controversial. Here we show that deletion of the entire zebrafish lrrk2 locus elicits a pleomorphic transient brain phenotype in maternal-zygotic mutant embryos (mzLrrk2). In contrast to lrrk2, the paralog gene lrrk1 is virtually not expressed in the brain of both wild-type and mzLrrk2 fish at different developmental stages. Notably, we found reduced catecholaminergic neurons, the main target of PD, in specific cell populations in the brains of mzLrrk2 larvae, but not adult fish. Strikingly, age-dependent accumulation of monoamine oxidase (MAO)-dependent catabolic signatures within mzLrrk2 brains revealed a previously undescribed interaction between LRRK2 and MAO biological activities. Our results highlight mzLrrk2 zebrafish as a tractable tool to study LRRK2 loss-of-function in vivo, and suggest a link between LRRK2 and MAO, potentially of relevance in the prodromic stages of PD.

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A Novel Drosophila Alkaline Phosphatase Specific to the Ellipsoid Body of the Adult Brain and the Lower Malpighian (Renal) Tubule

Genetics ◽

10.1093/genetics/154.1.285 ◽

2000 ◽

Vol 154 (1) ◽

pp. 285-297 ◽

Cited By ~ 1

Author(s):

Ming Yao Yang ◽

Zongsheng Wang ◽

Matthew MacPherson ◽

Julian A T Dow ◽

Kim Kaiser

Keyword(s):

Alkaline Phosphatase ◽

Expression Patterns ◽

Fluid Secretion ◽

Regulatory Elements ◽

Enhancer Trap ◽

Adult Brain ◽

P Element ◽

Renal Tubules ◽

Genomic Locus ◽

Ellipsoid Body

Abstract Two independent Drosophila melanogaster P{GAL4} enhancer-trap lines revealed identical GAL4-directed expression patterns in the ellipsoid body of the brain and in the Malpighian (renal) tubules in the abdomen. Both P-element insertions mapped to the same chromosomal site (100B2). The genomic locus, as characterized by plasmid rescue of flanking DNA, restriction mapping, and DNA sequencing, revealed the two P{GAL4} elements to be inserted in opposite orientations, only 46 bp apart. Three genes flanking the insertions have been identified. Calcineurin A1 (previously mapped to 21E-F) lies to one side, and two very closely linked genes lie to the other. The nearer encodes Aph-4, the first Drosophila alkaline phosphatase gene to be identified; the more distant gene [l(3)96601] is novel, with a head-elevated expression, and with distant similarity to transcription regulatory elements. Both in situ hybridization with Aph-4 probes and direct histochemical determination of alkaline phosphatase activity precisely matches the enhancer-trap pattern reported by the original lines. Although the P-element insertions are not recessive lethals, they display tubule phenotypes in both heterozygotes and homozygotes. Rates of fluid secretion in tubules from c507 homozygotes are reduced, both basally, and after stimulation by CAP2b, cAMP, or Drosophila leucokinin. The P-element insertions also disrupt the expression of Aph-4, causing misexpression in the tubule main segment. This disruption extends to tubule pigmentation, with c507 homozygotes displaying white-like transparent main segments. These results suggest that Aph-4, while possessing a very narrow range of expression, nonetheless plays an important role in epithelial function.

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Activation of endogenous retroviruses during brain development causes neuroinflammation

10.1101/2020.07.07.191668 ◽

2020 ◽

Cited By ~ 2

Author(s):

Marie E Jönsson ◽

Raquel Garza ◽

Yogita Sharma ◽

Rebecca Petri ◽

Erik Södersten ◽

...

Keyword(s):

Brain Development ◽

Transcriptional Activation ◽

Neural Progenitor Cells ◽

Large Fraction ◽

Endogenous Retroviruses ◽

Adult Brain ◽

Potential Trigger ◽

Mammalian Genomes ◽

The Brain

AbstractEndogenous retroviruses (ERVs) make up a large fraction of mammalian genomes and are thought to contribute to human disease, including brain disorders. In the brain, aberrant activation of ERVs is a potential trigger for neuroinflammation, but mechanistic insight into this phenomenon remains lacking. Using CRISPR/Cas9-based gene disruption of the epigenetic co-repressor protein Trim28, we found a dynamic H3K9me3-dependent regulation of ERVs in proliferating neural progenitor cells (NPCs), but not in adult neurons. In vivo deletion of Trim28 in cortical NPCs during mouse brain development resulted in viable offspring expressing high levels of ERV expression in excitatory neurons in the adult brain. Neuronal ERV expression was linked to inflammation, including activated microglia, and aggregates of ERV-derived proteins. This study demonstrates that brain development is a critical period for the silencing of ERVs and provides causal in vivo evidence demonstrating that transcriptional activation of ERV in neurons results in neuroinflammation.

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Whole-brain optical access in small adult vertebrates with two- and three-photon microscopy

10.1101/2021.12.09.471956 ◽

2021 ◽

Author(s):

Najva Akbari ◽

Rose L Tatarsky ◽

Andrew H Bass ◽

Chris Xu

Keyword(s):

Developmental Stages ◽

Multiphoton Microscopy ◽

Adult Brain ◽

High Signal ◽

Larval Brain ◽

Optical Access ◽

Low Contrast ◽

Two Photon Microscopy ◽

Vertebrate Brain ◽

Entire Brain

Although optical microscopy has allowed us to study the entire brain in early developmental stages, access to the brains of live, adult vertebrates has been limited. Danionella, a genus of miniature, transparent fish closely related to zebrafish has been introduced as a neuroscience model to study the entire adult vertebrate brain. However, the extent of optically accessible depth in these animals has not been quantitatively characterized. Here, we show that two- and three-photon microscopy can be used to access the entire depth of the adult wild type Danionella dracula brain without any modifications to the animal other than mechanical stabilization. Three-photon microscopy provides high signal to background ratio and optical sectioning through the deepest part of the brain. While vasculature can be observed with two-photon microscopy, the deeper regions have low contrast. We show that multiphoton microscopy is ideal for readily penetrating the entire adult brain within the geometry of these animals' head structures and without the need for pigment removal. With multiphoton microscopy enabling optical access to the entire adult brain and a repertoire of methods that allow observation of the larval brain, Danionella provides a model system for readily studying the entire brain over the lifetime of a vertebrate.

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Human atlas of cardiac promoters and enhancers reveals important role of regulatory elements in heritable diseases

10.21203/rs.3.rs-37530/v1 ◽

2020 ◽

Author(s):

Ruslan Deviatiiarov ◽

Anna Gams ◽

Roman Syunyaev ◽

Tatiana Tatarinova ◽

Oleg Gusev ◽

...

Keyword(s):

Heart Diseases ◽

Association Studies ◽

Expression Patterns ◽

Critical Role ◽

Regulatory Elements ◽

Specific Cell ◽

Genome Wide Association Studies ◽

Specific Expression ◽

Enhancer Activity ◽

Human Donor

Abstract Genome regulatory elements play a critical role during cardiac development and maintenance of normal physiological homeostasis, and genome-wide association studies identified a large number of SNPs associated with cardiovascular diseases localized in intergenic zones. We used cap analysis of gene expression (CAGE) to identify transcription start sites (TSS) with one nucleotide resolution that effectively maps genome regulatory elements in a representative collection of human heart tissues. Here we present a comprehensive and fully annotated CAGE atlas of human promoters and enhancers from four chambers of the non-diseased human donor hearts, including both atria and ventricles. We have identified 10,528 novel regulatory elements, where 2,750 are classified as TSS and 4,258 novel enhancers, which were validated with ChIP-seq libraries and motif enrichment analysis. We found that heart-region specific expression patterns are primarily based on the alternative promoter and specific enhancer activity. Our study significantly increased evidence of the association of regulatory elements-located variants with heart morphology and pathologies. The precise location of cardiac disease-related SNPs within the regulatory regions and their correlation with a specific cell type offers a new understanding of genetic heart diseases.

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Molecular regulation of fetal brain development in pigs

10.32469/10355/88112 ◽

2021 ◽

Author(s):

◽

Monica P. Strawn

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Transcription Factor ◽

Brain Development ◽

Early Development ◽

Expression Patterns ◽

Fetal Brain ◽

Molecular Regulation ◽

Fetal Brain Development ◽

The Brain

Two experiments were conducted to investigate molecular regulation that impacts fetal brain development in pigs. In the first experiment (Chapter 2), gene expression was profiled by RNA sequencing (RNA-seq) to examine the whole transcriptome of the male (M) and female (F) fetal brain at gestation day (d) 45, 60 and 90. The analysis showed fewer differentially expressed genes (DEGs) in the brain of male and female fetuses in earlier gestation (d45-d60) when compared to late gestation (d60-d90). The homeobox (HOX) A5 gene that regulates pattern formation in early development was in the top upregulated DEGs between d45 to d60 in fetuses of both sexes. This study also found HOX B5 and D3 genes were in the top upregulated genes between d45 and d60 of the fetal brain of females, but not males. The second experiment (Chapter 3) investigated DNA methylation in pigs. DNA methylation in the fetal brain of both sexes at the same three gestation days was performed by enzymatic methyl sequencing (EM-seq). Hotspots of methylation in specific chromosomal regions were observed in the analysis. The analysis identified 1,475 sites in the pig genome that were methylated in the fetal brain, irrespective of sex, during development. The same sites were methylated in a canonically correlated manner in the blood of the adult stage, both in sows and boars. This is consistent with the Dilman theory of developmental aging (DevAge), which suggests that aging and early development of the brain are regulated by common molecular processes. A comparative analysis (Chapter 4) compared the gene expression patterns in the fetal brain and placenta between pigs and mice. The analysis identified 112 genes that were expressed (mean FPKM > 10) in the fetal brain of both species but not expressed (mean FPKM < 1) in the placenta of either species, and 10 genes that were expressed in the placenta of both species but not expressed in the fetal brain. In-silico analysis of the transcription factor binding sites in the 500 bp of the upstream DNA of these common genes revealed that they were commonly regulated by the RE1 silencing transcription factor (REST), which is a multifaceted transcription factor that acts as a master regulator of neurogenesis as well as controls neural excitation and the aging processes.

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