scholarly journals The Drosophila gene stand still encodes a germline chromatin-associated protein that controls the transcription of the ovarian tumor gene

Development ◽  
1999 ◽  
Vol 126 (9) ◽  
pp. 1917-1926 ◽  
Author(s):  
I. Sahut-Barnola ◽  
D. Pauli
Keyword(s):  
Rna Polymerase Ii ◽  
Germ Cells ◽  
Ovarian Tumor ◽  
Polytene Chromosomes ◽  
Ectopic Expression ◽  
Point Mutations ◽  
Subnuclear Localization ◽  
Drosophila Gene ◽  
Germline Differentiation ◽  
Tumor Gene

The Drosophila gene stand still (stil) encodes a novel protein required for survival, sexual identity and differentiation of female germ cells. Using specific antibodies, we show that the Stil protein accumulates in the nucleus of all female germ cells throughout development, and is transiently expressed during early stages of male germline differentiation. Changes of Stil subnuclear localization during oogenesis suggest an association with chromatin. Several mutant alleles, which are point mutations in the Stil N-terminal domain, encode proteins that no longer co-localized with chromatin. We find that Stil binds to many sites on polytene chromosomes with strong preference for decondensed chromatin. This localization is very similar to that of RNA polymerase II. We show that Stil is required for high levels of transcription of the ovarian tumor gene in germ cells. Expression of ovarian tumor in somatic cells can be induced by ectopic expression of Stil. Finally, we find that transient ubiquitous somatic expression of Stil results in lethality of the fly at all stages of development.

Regulatory and functional interactions between the somatic sex regulatory gene transformer and the germline genes ovo and ovarian tumor

Development ◽  
1999 ◽  
Vol 126 (5) ◽  
pp. 861-871 ◽  
Author(s):  
S. Hinson ◽  
R.N. Nagoshi
Keyword(s):  
Germ Cells ◽  
Ovarian Tumor ◽  
Sex Chromosome ◽  
Sex Differentiation ◽  
Regulatory Gene ◽  
Tumor Promoter ◽  
Germline Genes ◽  
Germline Differentiation ◽  
Tumor Expression ◽  
Female Germline

In Drosophila, compatibility between the sexually differentiated state of the soma and the sex chromosome constitution of the germline is required for normal gametogenesis. In this study, we defined important aspects of the soma-germline interactions controlling early oogenesis. In particular, the sex-specific germline activity of the ovarian tumor promoter was found to be dependent upon somatic factors controlled by the somatic sex differentiation gene transformer. This regulation defines whether there is sufficient ovarian tumor expression in adult XX germ cells to support oogenesis. In addition, the ovarian tumor function required for female germline differentiation is dependent on the activity of another germline gene, ovo, whose regulation is transformer-independent. These and other data indicate that ovarian tumor plays a central role in coordinating regulatory inputs from the soma (as regulated by transformer) with those from the germline (involving ovo). We also demonstrate that transformer-dependent interactions influence whether XX germ cells require ovarian tumor or ovo functions to undergo early gametogenic differentiation. These results are incorporated into a model hypothesizing that the functions of ovarian tumor and ovo are dependent on an early sex determination decision in the XX germline that is at least partially controlled by somatic transformer activity.

scholarly journals The Phosphatidylserine Receptor TIM-1 Enhances Authentic Chikungunya Virus Cell Entry

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1828
Author(s):  
Jared Kirui ◽  
Yara Abidine ◽  
Annasara Lenman ◽  
Koushikul Islam ◽  
Yong-Dae Gwon ◽  
...  
Keyword(s):  
Chikungunya Virus ◽  
Molecular Mechanisms ◽  
Rna Virus ◽  
Ectopic Expression ◽  
Point Mutations ◽  
Cell Entry ◽  
Target Cells ◽  
Human Hepatoma Cells ◽  
Chikv Infection ◽  
Phosphatidylserine Receptor

Chikungunya virus (CHIKV) is a re-emerging, mosquito-transmitted, enveloped positive stranded RNA virus. Chikungunya fever is characterized by acute and chronic debilitating arthritis. Although multiple host factors have been shown to enhance CHIKV infection, the molecular mechanisms of cell entry and entry factors remain poorly understood. The phosphatidylserine-dependent receptors, T-cell immunoglobulin and mucin domain 1 (TIM-1) and Axl receptor tyrosine kinase (Axl), are transmembrane proteins that can serve as entry factors for enveloped viruses. Previous studies used pseudoviruses to delineate the role of TIM-1 and Axl in CHIKV entry. Conversely, here, we use the authentic CHIKV and cells ectopically expressing TIM-1 or Axl and demonstrate a role for TIM-1 in CHIKV infection. To further characterize TIM-1-dependent CHIKV infection, we generated cells expressing domain mutants of TIM-1. We show that point mutations in the phosphatidylserine binding site of TIM-1 lead to reduced binding, entry, and infection of CHIKV. Ectopic expression of TIM-1 renders immortalized keratinocytes permissive to CHIKV, whereas silencing of endogenously expressed TIM-1 in human hepatoma cells reduces CHIKV infection. Altogether, our findings indicate that, unlike Axl, TIM-1 readily promotes the productive entry of authentic CHIKV into target cells.

scholarly journals Drosophila Mi-2 Negatively Regulates dDREF by Inhibiting Its DNA-Binding Activity

2002 ◽  
Vol 22 (14) ◽  
pp. 5182-5193 ◽  
Author(s):  
Fumiko Hirose ◽  
Nobuko Ohshima ◽  
Eun-Jeong Kwon ◽  
Hideki Yoshida ◽  
Masamitsu Yamaguchi
Keyword(s):  
Dna Binding ◽  
Polytene Chromosomes ◽  
Ectopic Expression ◽  
Binding Activity ◽  
Mobility Shift ◽  
Interacting Protein ◽  
Reciprocal Regulation ◽  
Dna Binding Activity ◽  
Expression Of Genes ◽  
Biochemical Analyses

ABSTRACT Drosophila melanogaster DNA replication-related element (DRE) factor (dDREF) is a transcriptional regulatory factor required for the expression of genes carrying the 5′-TATCGATA DRE. dDREF has been reported to bind to a sequence in the chromatin boundary element, and thus, dDREF may play a part in regulating insulator activity. To generate further insights into dDREF function, we carried out a Saccharomyces cerevisiae two-hybrid screening with DREF polypeptide as bait and identified Mi-2 as a DREF-interacting protein. Biochemical analyses revealed that the C-terminal region of Drosophila Mi-2 (dMi-2) specifically binds to the DNA-binding domain of dDREF. Electrophoretic mobility shift assays showed that dMi-2 thereby inhibits the DNA-binding activity of dDREF. Ectopic expression of dDREF and dMi-2 in eye imaginal discs resulted in severe and mild rough-eye phenotypes, respectively, whereas flies simultaneously expressing both proteins exhibited almost-normal eye phenotypes. Half-dose reduction of the dMi-2 gene enhanced the DREF-induced rough-eye phenotype. Immunostaining of polytene chromosomes of salivary glands showed that dDREF and dMi-2 bind in mutually exclusive ways. These lines of evidence define a novel function of dMi-2 in the negative regulation of dDREF by its DNA-binding activity. Finally, we postulated that dDREF and dMi-2 may demonstrate reciprocal regulation of their functions.

zfh-1 is required for germ cell migration and gonadal mesoderm development in Drosophila

Development ◽  
1998 ◽  
Vol 125 (4) ◽  
pp. 655-666 ◽  
Author(s):  
H.T. Broihier ◽  
L.A. Moore ◽  
M. Van Doren ◽  
S. Newman ◽  
R. Lehmann
Keyword(s):  
Cell Migration ◽  
Germ Cell ◽  
Germ Cells ◽  
Ectopic Expression ◽  
Homeodomain Protein ◽  
Mesoderm Development ◽  
Visceral Mesoderm ◽  
Temporal Course ◽  
Germ Cell Migration

In Drosophila as well as many vertebrate systems, germ cells form extraembryonically and migrate into the embryo before navigating toward gonadal mesodermal cells. How the gonadal mesoderm attracts migratory germ cells is not understood in any system. We have taken a genetic approach to identify genes required for germ cell migration in Drosophila. Here we describe the role of zfh-1 in germ cell migration to the gonadal mesoderm. In zfh-1 mutant embryos, the initial association of germ cells and gonadal mesoderm is blocked. Loss of zfh-1 activity disrupts the development of two distinct mesodermal populations: the caudal visceral mesoderm and the gonadal mesoderm. We demonstrate that the caudal visceral mesoderm facilitates the migration of germ cells from the endoderm to the mesoderm. Zfh-1 is also expressed in the gonadal mesoderm throughout the development of this tissue. Ectopic expression of Zfh-1 is sufficient to induce additional gonadal mesodermal cells and to alter the temporal course of gene expression within these cells. Finally, through analysis of a tinman zfh-1 double mutant, we show that zfh-1 acts in conjunction with tinman, another homeodomain protein, in the specification of lateral mesodermal derivatives, including the gonadal mesoderm.

Functional analysis of a stable transcription arrest site in the first intron of the murine adenosine deaminase gene

1993 ◽  
Vol 13 (5) ◽  
pp. 2718-2729
Author(s):  
S F Kash ◽  
J W Innis ◽  
A U Jackson ◽  
R E Kellems
Keyword(s):  
Rna Polymerase Ii ◽  
Adenosine Deaminase ◽  
Functional Characterization ◽  
Gel Filtration ◽  
Point Mutations ◽  
Elongation Factor ◽  
Dependent Manner ◽  
First Intron ◽  
Intron 1

Transcription arrest plays a role in regulating the expression of a number of genes, including the murine adenosine deaminase (ADA) gene. We have previously identified two prominent arrest sites at the 5' end of the ADA gene: one in the first exon and one in the first intron (J. W. Innis and R. E. Kellems, Mol. Cell. Biol. 11:5398-5409, 1991). Here we report the functional characterization of the intron 1 arrest site, located 137 to 145 nucleotides downstream of the cap site. We have determined, using gel filtration, that the intron 1 arrest site is a stable RNA polymerase II pause site and that the transcription elongation factor SII promotes read-through at this site. Additionally, the sequence determinants for the pause are located within a 37-bp fragment encompassing this site (+123 to +158) and can direct transcription arrest in an orientation-dependent manner in the context of the ADA and adenovirus major late promoters. Specific point mutations in this region increase or decrease the relative pausing efficiency. We also show that the sequence determinants for transcription arrest can function when placed an additional 104 bp downstream of their natural position.

scholarly journals Abnormal Meiosis Initiation in Germ Cell Caused by Aberrant Differentiation of Gonad Somatic Cell

2019 ◽  
Vol 2019 ◽  
pp. 1-8
Author(s):  
Min Chen ◽  
Min Chen ◽  
Suren Chen ◽  
Jingjing Zhou ◽  
Fangfang Dong ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Germ Cell ◽  
Somatic Cell ◽  
Germ Cells ◽  
Somatic Cells ◽  
Ectopic Expression ◽  
Genital Ridge ◽  
Male And Female ◽  
Germ Cell Differentiation ◽  
Exact Function

The interaction between germ cell and somatic cell plays important roles in germ cell development. However, the exact function of gonad somatic cell in germ cell differentiation is unclear. In the present study, the function of gonad somatic cell in germ cell meiosis was examined by using mouse models with aberrant somatic cell differentiation. In Wt1R394W/R394W mice, the genital ridge is absent due to the apoptosis of coelomic epithelial cells. Interestingly, in both male and female Wt1R394W/R394W germ cells, STRA8 was detected at E12.5 and the scattered SYCP3 foci were observed at E13.5 which was consistent with control females. In Wt1-/flox; Cre-ERTM mice, Wt1 was inactivated by the injection of tamoxifen at E9.5 and the differentiation of Sertoli and granulosa cells was completely blocked. We found that most germ cells were located outside of genital ridge after Wt1 inactivation. STRA8, SYCP3, and γH2AX proteins were detected in germ cells of both male and female Wt1-/flox; Cre-ERTM gonads, whereas no thread-like SYCP3 signal was observed. Our study demonstrates that aberrant development of gonad somatic cells leads to ectopic expression of meiosis-associated genes in germ cells, but meiosis was arrested before prophase I. These results suggest that the proper differentiation of gonad somatic cells is essential for germ cell meiosis.

scholarly journals A three-dimensional structural dissection of Drosophila polytene chromosomes.

1995 ◽  
Vol 131 (2) ◽  
pp. 279-295 ◽  
Author(s):  
Y Urata ◽  
S J Parmelee ◽  
D A Agard ◽  
J W Sedat
Keyword(s):  
Rna Polymerase Ii ◽  
Polytene Chromosome ◽  
Polytene Chromosomes ◽  
Three Dimensional ◽  
Spatial Relationship ◽  
Band Shape ◽  
Structural Details ◽  
Novel Method ◽  
Relationship Of

We have analyzed the three-dimensional structural details of Drosophila melanogaster polytene chromosome bands and interbands using three-dimensional light microscopy and a novel method of sample preparation that does not involve flattening or stretching the chromosomes. Bands have been visualized in unfixed chromosomes stained with the DNA specific dye 4,6-Diamidino-2-phenylindole (DAPI). Interbands have been visualized using fixed chromosomes that have been immunostained with an antibody to RNA polymerase II. Additionally, these structures have been analyzed using in situ hybridization with probes from specific genetic loci (Notch and white). Bands are seen to be composed of approximately 36 substructural features that measure 0.2-0.4 micron in diameter. We suggest that these substructural features are in fact longitudinal fibers made up of bundles of chromatids. Band shape can be a reproducible characteristic of a particular band and is dependent on the spatial relationship of these bundles, varying from bands with a uniform distribution of bundles to bands with a peripheral concentration of chromatin. Interbands are composed of bundles of chromatids of a similar size and number as those seen in the bands. The distribution of bundles is similar between a band and the neighboring interband, implying that there is a long range organization to the DNA that includes both the coding and the noncoding portions of genes. Finally, we note that the polytene chromosome has a circular shape when viewed in cross section, whether there are one or two homologs present.

scholarly journals Proteomic and metabolomic analyses uncover sex-specific regulatory pathways in mouse fetal germline differentiation†

2020 ◽  
Vol 103 (4) ◽  
pp. 717-735
Author(s):  
Yohei Hayashi ◽  
Masaru Mori ◽  
Kaori Igarashi ◽  
Keiko Tanaka ◽  
Asuka Takehara ◽  
...  
Keyword(s):  
Oxidative Phosphorylation ◽  
Germ Cell ◽  
Germ Cells ◽  
Cell Development ◽  
Nucleotide Synthesis ◽  
Male Germ Cells ◽  
Regulatory Pathways ◽  
Germ Cell Development ◽  
Germ Cell Differentiation ◽  
Germline Differentiation

Abstract Regulatory mechanisms of germline differentiation have generally been explained via the function of signaling pathways, transcription factors, and epigenetic regulation; however, little is known regarding proteomic and metabolomic regulation and their contribution to germ cell development. Here, we conducted integrated proteomic and metabolomic analyses of fetal germ cells in mice on embryonic day (E)13.5 and E18.5 and demonstrate sex- and developmental stage-dependent changes in these processes. In male germ cells, RNA processing, translation, oxidative phosphorylation, and nucleotide synthesis are dominant in E13.5 and then decline until E18.5, which corresponds to the prolonged cell division and more enhanced hyper-transcription/translation in male primordial germ cells and their subsequent repression. Tricarboxylic acid cycle and one-carbon pathway are consistently upregulated in fetal male germ cells, suggesting their involvement in epigenetic changes preceding in males. Increased protein stability and oxidative phosphorylation during female germ cell differentiation suggests an upregulation of aerobic energy metabolism, which likely contributes to the proteostasis required for oocyte maturation in subsequent stages. The features elucidated in this study shed light on the unrevealed mechanisms of germ cell development.

scholarly journals Genetic interaction mapping informs integrative structure determination of protein complexes

Science ◽  
2020 ◽  
Vol 370 (6522) ◽  
pp. eaaz4910 ◽  
Author(s):  
Hannes Braberg ◽  
Ignacia Echeverria ◽  
Stefan Bohn ◽  
Peter Cimermancic ◽  
Anthony Shiver ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Structure Determination ◽  
Genetic Interaction ◽  
Protein Complexes ◽  
Point Mutations ◽  
Genetic Interactions ◽  
Cellular Functions ◽  
Cross Links

Determining structures of protein complexes is crucial for understanding cellular functions. Here, we describe an integrative structure determination approach that relies on in vivo measurements of genetic interactions. We construct phenotypic profiles for point mutations crossed against gene deletions or exposed to environmental perturbations, followed by converting similarities between two profiles into an upper bound on the distance between the mutated residues. We determine the structure of the yeast histone H3-H4 complex based on ~500,000 genetic interactions of 350 mutants. We then apply the method to subunits Rpb1-Rpb2 of yeast RNA polymerase II and subunits RpoB-RpoC of bacterial RNA polymerase. The accuracy is comparable to that based on chemical cross-links; using restraints from both genetic interactions and cross-links further improves model accuracy and precision. The approach provides an efficient means to augment integrative structure determination with in vivo observations.

scholarly journals A cytological approach to the ordering of events in gene activation using the Sgs-4 locus of Drosophila melanogaster.

1984 ◽  
Vol 99 (1) ◽  
pp. 233-238 ◽  
Author(s):  
E K Steiner ◽  
J C Eissenberg ◽  
S C Elgin
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Topoisomerase I ◽  
Gene Activation ◽  
Polytene Chromosomes ◽  
Null Alleles ◽  
Major Salivary Gland ◽  
Wild Type ◽  
Fluorescent Band ◽  
Chromosomal Proteins

The polytene chromosomes of Drosophila strains that differ in the synthesis of the major salivary gland glue protein sgs-4 were examined by indirect immunofluorescence using antisera to several nonhistone chromosomal proteins. The Oregon-R X chromosome, which produces sgs-4 messenger RNA, showed a strong fluorescent band at locus 3C11-12 when stained with anti-RNA polymerase II, whereas the null mutant Berkeley 1 failed to exhibit fluorescence at that locus. The presence of another antigen (Band 2), normally associated with developmentally active loci, was clearly evident at locus 3C11-12 of both transcriptionally competent and null strains, indicating that the association of Band 2 antigen with the chromatin is an event independent of RNA polymerase II binding. Antibodies directed against Drosophila topoisomerase I stained 3C11-12 in the Sgs-4+ (wild-type) strain brightly, but gave significantly less staining in the null strain. This indicates that the high concentrations of topoisomerase I seen at active loci are closely associated with the transcriptional event. In some of these analyses, we have made use of flies heterozygous for the wild-type and null alleles in order to make internally controlled comparisons. The results suggest that this type of analysis will enable conclusions to be drawn concerning the interdependence and order of action of chromosomal proteins involved in developmental gene activation.

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