scholarly journals Genetic analysis of transvection effects involving cis-regulatory elements of the Drosophila Ultrabithorax gene.

Genetics ◽  
1990 ◽  
Vol 126 (2) ◽  
pp. 365-373
Author(s):  
J L Micol ◽  
J E Castelli-Gair ◽  
A García-Bellido
Keyword(s):  
Regulatory Elements ◽  
Transcription Unit ◽  
Null Alleles ◽  
Loss Of Function ◽  
Partial Loss ◽  
Protein Coding ◽  
Critical Elements ◽  
Regulatory Mutations ◽  
In Trans

Abstract The Ultrabithorax (Ubx) gene of Drosophila melanogaster contains two functionally distinguishable regions: the protein-coding Ubx transcription unit and, upstream of it, the transcribed but nonprotein-coding bxd region. Numerous recessive, partial loss-of-function mutations which appear to be regulatory mutations map within the bxd region and within the introns of the Ubx transcription unit. In addition, mutations within the Ubx unit exons are known and most of these behave as null alleles. Ubx1 is one such allele. We have confirmed that, although the Ubx1 allele does not produce detectable Ubx proteins (UBX), it does retain other genetic functions detectable by their effects on the expression of a paired, homologous Ubx allele, i.e., by transvection. We have extended previous analyses made by E. B. Lewis by mapping the critical elements of the Ubx gene which participate in transvection effects. Our results show that the Ubx1 allele retains wild-type functions whose effectiveness can be reduced (1) by additional cis mutations in the bxd region or in introns of the Ubx transcription unit, as well as (2) by rearrangements disturbing pairing between homologous Ubx genes. Our results suggest that those remnant functions in Ubx1 are able to modulate the activity of the allele located in the homologous chromosome. We discuss the normal cis regulatory role of these functions involved in trans interactions between homologous Ubx genes, as well as the implications of our results for the current models on transvection.

scholarly journals Genetic structure of the abd-A gene of Drosophila

Development ◽  
1989 ◽  
Vol 107 (3) ◽  
pp. 575-583 ◽  
Author(s):  
A. Busturia ◽  
J. Casanova ◽  
E. Sanchez-Herrero ◽  
R. Gonzalez ◽  
G. Morata
Keyword(s):  
Genetic Structure ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Transcription Unit ◽  
Bithorax Complex ◽  
Coding Region ◽  
Regulatory Regions ◽  
Protein Coding ◽  
Spatial Regulation ◽  
Correct Level

We report the embryonic and adult phenotypes of a number of mutations of the abd-A gene of the bithorax complex. Some of them result in loss of abd-A function in the whole abd-A domain and are usually lethal. These probably eliminate or inactivate abd-A protein products. Other mutations affect only part of the abd-A domain. These are viable, appear to map outside the abd-A transcription unit, and presumably alter the normal spatial regulation of abd-A products. We propose a model of abd-A structure based on a protein-coding region and two cis-regulatory regions. Regulatory region 1, 3′ to the transcription unit, contains positive and negative regulatory elements. Regulatory region 2, 5′ to the transcription unit, establishes the correct level of abd-A activity in the abdominal metameres.

scholarly journals Mosaic CRISPR-stop enables rapid phenotyping of nonsense mutations in essential genes

Development ◽  
2021 ◽  
Vol 148 (5) ◽  
pp. dev196899
Author(s):  
Guangqin Wang ◽  
Chao Li ◽  
Shunji He ◽  
Zhiyong Liu
Keyword(s):  
Outer Hair Cells ◽  
Conditional Knockout ◽  
Rapid Screening ◽  
Lethal Gene ◽  
Loss Of Function ◽  
Cell Stage ◽  
Nonsense Mutations ◽  
Protein Coding ◽  
Lethal Genes

ABSTRACTCRISPR-stop converts protein-coding sequences into stop codons, which, in the appropriate location, results in a null allele. CRISPR-stop induction in one-cell-stage zygotes generates Founder 0 (F0) mice that are homozygous mutants; this avoids mouse breeding and serves as a rapid screening approach for nonlethal genes. However, loss of function of 25% of mammalian genes causes early lethality. Here, we induced CRISPR-stop in one of the two blastomeres of the zygote, a method we name mosaic CRISPR-stop, to produce mosaic Atoh1 and Sox10 F0 mice; these mice not only survived longer than regular Atoh1/Sox10 knockout mice but also displayed their recognized cochlear phenotypes. Moreover, by using mosaic CRISPR-stop, we uncovered a previously unknown role of another lethal gene, Rbm24, in the survival of cochlear outer hair cells (OHCs), and we further validated the importance of Rbm24 in OHCs by using our Rbm24 conditional knockout model. Together, our results demonstrated that mosaic CRISPR-stop is reliable and rapid, and we believe this method will facilitate rapid genetic screening of developmentally lethal genes in the mouse inner ear and also in other organs.

Mammary Epithelial Cell Transcriptome Reveals Potential Roles of lncRNAs in Regulating Milk Synthesis Pathways in Jersey and Kashmiri Cattle

2021 ◽  
Author(s):  
Peerzada Tajamul Mumtaz ◽  
Basharat Bhat ◽  
Eveline M. Ibeagha-Awemu ◽  
Qamar Taban ◽  
Mengqi Wang ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Mammary Gland Development ◽  
Expression Profiles ◽  
Regulatory Elements ◽  
Milk Quality ◽  
Mammary Epithelial ◽  
Protein Coding ◽  
Protein Coding Genes ◽  
Gland Development

Abstract Background Long noncoding RNAs (lncRNAs) are now proven as essential regulatory elements, playing diverse role in many biological processes including mammary gland development. However, little is known about their roles in bovine lactation process. There are very few reports available to date on the role of lncRNAs in lactation physiology and mammary glands development in cattle. Results To identify and characterize the roles of lncRNAs in bovine lactation, milk derived mammary epithelial cells (MEC) from Jersey (high milk producer) and Kashmiri cattle (low milk producer) at early, mid and late lactation stages were used. The lncRNA transcriptome of the samples (n=18) was studied using next generation RNA sequencing technology. 633 putative lncRNAs were identified, 76 of which were differentially expressed (DE) between comparison between the three stages of lactation. Additionally, 56 DE lncRNAs were identified from 9 Jersey and 9 Kashmir samples. Correlation of DE lncRNAs with protein-coding genes resulted in a comprehensive list of lncRNA-mRNA co-expressed pairs. Most of the DE lncRNAs showed positive correlations with protein coding genes in Jersey compared to Kashmiri cattle where they were mainly negatively correlated, which could be one of the underlying mechanisms responsible for the differential milking performance between the two breeds. In addition, a number of the DE lncRNAs were paired with the most DE milk quality genes like GPAM, LPL, ABCG2, etc. indicative of their potential regulatory effects on milk quality genes. KEGG pathways analysis of potential cis and trans target genes of DE lncRNAs indicated that 27 and 48 pathways were significantly enriched in Kashmiri and Jersey respectively, including mTOR signaling, PI3K-Akt signaling and RAP1 signaling pathways. These pathways have been proven to play key roles in lactation biology and mammary gland development. Conclusions Our study mapped the expression profiles of lncRNAs across lactation stages and their relationships with candidate genes related to milk quality and yield traits in Jersey and Kashmiri cattle. These findings provide a valuable resource for the study of the regulatory mechanisms involved in the lactation process as well as facilitate understanding of the role of lncRNAs in bovine lactation biology.

scholarly journals Domain-Specific Response of Imprinted Genes to Reduced DNMT1

2010 ◽  
Vol 30 (16) ◽  
pp. 3916-3928 ◽  
Author(s):  
Jamie R. Weaver ◽  
Garnik Sarkisian ◽  
Christopher Krapp ◽  
Jesse Mager ◽  
Mellissa R. W. Mann ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Dna Methyltransferase ◽  
Critical Role ◽  
Imprinted Genes ◽  
Specific Response ◽  
Loss Of Function ◽  
Partial Loss ◽  
Domain Specific ◽  
Parent Of Origin

ABSTRACT Imprinted genes are expressed in a monoallelic, parent-of-origin-specific manner. Clusters of imprinted genes are regulated by imprinting control regions (ICRs) characterized by DNA methylation of one allele. This methylation is critical for imprinting; a reduction in the DNA methyltransferase DNMT1 causes a widespread loss of imprinting. To better understand the role of DNA methylation in the regulation of imprinting, we characterized the effects of Dnmt1 mutations on the expression of a panel of imprinted genes in the embryo and placenta. We found striking differences among imprinted domains. The Igf2 and Peg3 domains showed imprinting perturbations with both null and partial loss-of-function mutations, and both domains had pairs of coordinately regulated genes with opposite responses to loss of DNMT1 function, suggesting these domains employ similar regulatory mechanisms. Genes in the Kcnq1 domain were less sensitive to the absence of DNMT1. Cdkn1c exhibited imprinting perturbations only in null mutants, while Kcnq1 and Ascl2 were largely unaffected by a loss of DNMT1 function. These results emphasize the critical role for DNA methylation in imprinting and reveal the different ways it controls gene expression.

scholarly journals Investigating rare pathogenic/likely pathogenic exonic variation in bipolar disorder

2021 ◽  
Author(s):  
Xiaoming Jia ◽  
Fernando S. Goes ◽  
Adam E. Locke ◽  
Duncan Palmer ◽  
Weiqing Wang ◽  
...  
Keyword(s):  
Bipolar Disorder ◽  
Meta Analysis ◽  
Common Variant ◽  
European Ancestry ◽  
Loss Of Function ◽  
Single Nucleotide Variants ◽  
Protein Coding ◽  
Significant Enrichment ◽  
Coding Variants

AbstractBipolar disorder (BD) is a serious mental illness with substantial common variant heritability. However, the role of rare coding variation in BD is not well established. We examined the protein-coding (exonic) sequences of 3,987 unrelated individuals with BD and 5,322 controls of predominantly European ancestry across four cohorts from the Bipolar Sequencing Consortium (BSC). We assessed the burden of rare, protein-altering, single nucleotide variants classified as pathogenic or likely pathogenic (P-LP) both exome-wide and within several groups of genes with phenotypic or biologic plausibility in BD. While we observed an increased burden of rare coding P-LP variants within 165 genes identified as BD GWAS regions in 3,987 BD cases (meta-analysis OR = 1.9, 95% CI = 1.3–2.8, one-sided p = 6.0 × 10−4), this enrichment did not replicate in an additional 9,929 BD cases and 14,018 controls (OR = 0.9, one-side p = 0.70). Although BD shares common variant heritability with schizophrenia, in the BSC sample we did not observe a significant enrichment of P-LP variants in SCZ GWAS genes, in two classes of neuronal synaptic genes (RBFOX2 and FMRP) associated with SCZ or in loss-of-function intolerant genes. In this study, the largest analysis of exonic variation in BD, individuals with BD do not carry a replicable enrichment of rare P-LP variants across the exome or in any of several groups of genes with biologic plausibility. Moreover, despite a strong shared susceptibility between BD and SCZ through common genetic variation, we do not observe an association between BD risk and rare P-LP coding variants in genes known to modulate risk for SCZ.

scholarly journals LINC00174 Facilitates Cell Proliferation, Cell Migration and Tumor Growth of Osteosarcoma via Regulating the TGF-β/SMAD Signaling Pathway and Upregulating SSH2 Expression

2021 ◽  
Vol 8 ◽  
Author(s):  
Changjun Zheng ◽  
Ronghang Li ◽  
Shuang Zheng ◽  
Hongjuan Fang ◽  
Meng Xu ◽  
...  
Keyword(s):  
Cell Growth ◽  
Tumor Growth ◽  
Inhibitory Effect ◽  
Loss Of Function ◽  
Protein Coding ◽  
Smad Pathway ◽  
Smad Signaling Pathway ◽  
Protein Phosphatase 2

Osteosarcoma (OS), a frequent malignant tumor which mainly occurs in the bone. The roles of long noncoding RNAs (lncRNAs) have been revealed in cancers, including OS. LncRNA long intergenic non-protein coding RNA (LINC00174) has been validated as an oncogene in several cancers. However, the role of LINC00174 in OS has not been explored. In our research, loss-of-function assays were conducted to explore the function of LINC00174 in OS cells. Then, we explored the downstream pathway of LINC00174 in OS cells. Bioinformatics, RNA pull-down and RIP experiments investigated the downstream mechanism of LINC00174 in OS cells. Finally, in vivo assays clarified the effect of LINC00174 on tumorigenesis. We found that LINC00174 was upregulated in OS tissues and cells. LINC00174 knockdown repressed OS cell growth. Mechanistically, LINC00174 knockdown suppressed the TGF-β/SMAD pathway. LINC00174 interacted with miR-378a-3p, and slingshot protein phosphatase 2 (SSH2) 3′UTR was targeted by miR-378a-3p in OS cells. Rescue assays showed that SSH2 upregulation or miR-378a-3p inhibition counteracted the inhibitory effect of LINC00174 depletion in OS cell growth. Additionally, LINC00174 depletion suppressed tumor growth in mice. In conclusion, LINC00174 promotes OS cellular malignancy and tumorigenesis via the miR-378a-3p/SSH2 axis and the TGF-β/SMAD pathway, which might provide a novel insight for OS treatment.

scholarly journals SIRT2-knockdown Rescues GARS-induced Charcot-Marie-Tooth Neuropathy

2019 ◽  
Author(s):  
Chao Shen ◽  
Qian Qi ◽  
Yicai Qin ◽  
Dejian Zhou ◽  
Xinyuan Chen ◽  
...  
Keyword(s):  
Trna Synthetase ◽  
Nerve Degeneration ◽  
Loss Of Function ◽  
Wild Type ◽  
Partial Loss ◽  
Charcot Marie Tooth ◽  
Charcot Marie Tooth Disease ◽  
Underlying Mechanisms ◽  
Tooth Disease

AbstractCharcot-Marie-Tooth disease is the most common inherited peripheral neuropathy. Dominant mutations in glycyl-tRNA synthetase (GARS) gene cause peripheral nerve degeneration and lead to CMT disease type 2D. Mutations in GARS (GARSCMT2D) show partial loss-of-function features, suggesting that tRNA-charging deficits play a role in disease pathogenesis, but the underlying mechanisms are not fully understood. In this study we report that wild-type GARS tightly binds the NAD+-dependent deacetylase SIRT2 and inhibits its deacetylation activity, resulting in the hyperacetylated α-tubulin, the major substrate of SIRT2. Previous studies showed that acetylation of α-tubulin protects microtubules from mechanical breakage and keep axonal transportation. However, CMT2D mutations in GARS can not inhibit SIRT2 deacetylation, which leads to decrease acetylated α-tubulin and severe axonal transport deficits. Genetic reduction of SIRT2 in the Drosophila model rescues the GARS–induced axonal CMT neuropathy and extends the life span. Our findings demonstrate the pathogenic role of SIRT2-dependent α-tubulin deacetylation in mutant GARS-induced neuropathies and provide new perspectives for targeting SIRT2 as a potential therapy against hereditary axonopathies.

scholarly journals Gene-Based Analysis in HRC Imputed Genome Wide Association Data Identifies Three Novel Genes For Alzheimer’s Disease

2018 ◽  
Author(s):  
Emily Baker ◽  
Rebecca Sims ◽  
Ganna Leonenko ◽  
Aura Frizzati ◽  
Janet Harwood ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Regulatory Elements ◽  
Specific Protein ◽  
Polygenic Risk Score ◽  
Loss Of Function ◽  
Protein Coding ◽  
Transcriptional Regulatory Elements ◽  
Risk Alleles ◽  
Association Data

AbstractA novel POLARIS gene-based analysis approach was employed to compute gene-based polygenic risk score (PRS) for all individuals in the latest HRC imputed GERAD (N cases=3,332 and N controls=9,832) data using the International Genomics of Alzheimer’s Project summary statistics (N cases=13,676 and N controls=27,322, excluding GERAD subjects) to identify the SNPs and weight their risk alleles for the PRS score. SNPs were assigned to known, protein coding genes using GENCODE (v19). SNPs are assigned using both 1) no window around the gene and 2) a window of 35kb upstream and 10kb downstream to include transcriptional regulatory elements. The overall association of a gene is determined using a logistic regression model, adjusting for population covariates.Three novel gene-wide significant genes were determined from the POLARIS gene-based analysis using a gene window; PPARGC1A, RORA and ZNF423. The ZNF423 gene resides in an Alzheimer’s disease (AD)-specific protein network which also includes other AD-related genes. The PPARGC1A gene has been linked to energy metabolism and the generation of amyloid beta plaques and the RORA has strong links with genes which are differentially expressed in the hippocampus. We also demonstrate no enrichment for genes in either loss of function intolerant or conserved noncoding sequence regions.

Abstract 442: Generation of an Abcg1 Knock Out Mouse on the Reversa Background

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Gillian Douglas ◽  
Lucy Trelfa ◽  
Keith Channon ◽  
Ben Davies ◽  
Shoumo Bhattacharya
Keyword(s):  
Amino Acid ◽  
Intima Media Thickness ◽  
Null Alleles ◽  
Cholesterol Transport ◽  
Loss Of Function ◽  
Phosphate Binding ◽  
Knock Out ◽  
Plaque Regression ◽  
Genomic Editing

Background: Clinically HDL mediated reverse cholesterol transport (RCT) from macrophages has been shown to be inversely associated with carotid intima media thickness. Cholesterol efflux to mature HDL is mediated by ATP binding cassette transporter G1 (Abcg1). Abcg1 pays a key role in cholesterol transport with loss of function in macrophages and endothelial cells associated with significant cholesterol accumulation. However, mechanistic studies into the role of Abgc1 in plaque regression have been restricted due to the limitations of current regression models. Aims: To use TALENS mediated genomic editing to generate an Abcg1 knockout mouse on the REVERSA background to enable the investigation of its role in plaque regression. Methods and results: TALENs constructs were targeted to exon 3 upstream of the phosphate binding Walker A domain. TALEN mRNA was injected into REVERSA oocytes which were then implanted into foster mice. Founders were screened by Cel1 nuclease assay and sequencing. Three independent alleles were identified two of which create frameshift mutations (predicted to be null alleles) and one which resulted in a 3 amino acid deletion and a one amino acid substitution near the Walker A domain (potential hypomorphic allele). The two founder lines with frame shift mutations (KO 145 and 171) were taken forward for additional analysis. RNA extracted from primary macrophages from WT (REVERSA) and homozygous Abcg1 knockout mice was used to confirm the mutation was transcribed to RNA. Intron-spanning primers were designed and a product of the expected size was obtained and sequence analysis confirmed the insertion (KO-145) and deletion (KO-171) within the WALKER A domain of Abcg1. To ensure the mutations resulted in loss of function, a radioactive RCT assay was carried out in bone marrow derived primary macrophages. A significant decrease in RCT to HDL was observed in macrophages from both the KO-145 and KO-171 lines as expected there was no difference in RCT to ApoA1-I. Conclusions: Using genomic editing we have generated a gene specific knockout on the REVERSA background which will enable, for the first time, the investigation of the role of Abcg1 in plaque regression.

Genetic studies of mei-1 gene activity during the transition from meiosis to mitosis in Caenorhabditis elegans.

Genetics ◽  
1993 ◽  
Vol 134 (1) ◽  
pp. 199-210 ◽  
Author(s):  
T R Clandinin ◽  
P E Mains
Keyword(s):  
Caenorhabditis Elegans ◽  
Null Alleles ◽  
Mitotic Spindles ◽  
Dominant Effect ◽  
Genetic Studies ◽  
In Trans ◽  
In Cis ◽  
Normal Meiosis

Abstract Genetic evidence suggests that the mei-1 locus of Caenorhabditis elegans encodes a maternal product required for female meiosis. However, a dominant gain-of-function allele, mei-1(ct46), can support normal meiosis but causes defects in subsequent mitotic spindles. Previously identified intragenic suppressors of ct46 lack functional mei-1 activity; null alleles suppress only in cis but other alleles arise frequently and suppress both in cis and in trans. Using a different screen for suppressors of the dominant ct46 defect, the present study describes another type of intragenic mutation that also arises at high frequency. These latter alleles appear to have reduced meiotic activity and retain a weakened dominant effect. Characterization of these alleles in trans-heterozygous combinations with previously identified mei-1 alleles has enabled us to define more clearly the role of the mei-1 gene product during normal embryogenesis. We propose that a certain level of mei-1 activity is required for meiosis but must be eliminated prior to mitosis. The dominant mutation causes mei-1 activity to function at mitosis; intragenic trans-suppressors act in an antimorphic manner to inactivate multimeric mei-1 complexes. We propose that inactivation of meiosis-specific functions may be an essential precondition of mitosis; failure to eliminate such functions may allow ectopic meiotic activity during mitosis and cause embryonic lethality.

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