scholarly journals Opposing Wnt signals regulate cervical squamocolumnar homeostasis and emergence of metaplasia

2021 ◽  
Vol 23 (2) ◽  
pp. 184-197 ◽  
Author(s):  
Cindrilla Chumduri ◽  
Rajendra Kumar Gurumurthy ◽  
Hilmar Berger ◽  
Oliver Dietrich ◽  
Naveen Kumar ◽  
...  
Keyword(s):  
Transition Zone ◽  
Single Molecule ◽  
Spatial Organization ◽  
Lineage Tracing ◽  
Stromal Compartment ◽  
Transition Zones ◽  
Wnt Inhibitor ◽  
Stem Cell Populations ◽  
In Situ Hybridizations

AbstractThe transition zones of the squamous and columnar epithelia constitute hotspots for the emergence of cancer, often preceded by metaplasia, in which one epithelial type is replaced by another. It remains unclear how the epithelial spatial organization is maintained and how the transition zone niche is remodelled during metaplasia. Here we used single-cell RNA sequencing to characterize epithelial subpopulations and the underlying stromal compartment of endo- and ectocervix, encompassing the transition zone. Mouse lineage tracing, organoid culture and single-molecule RNA in situ hybridizations revealed that the two epithelia derive from separate cervix-resident lineage-specific stem cell populations regulated by opposing Wnt signals from the stroma. Using a mouse model of cervical metaplasia, we further show that the endocervical stroma undergoes remodelling and increases expression of the Wnt inhibitor Dickkopf-2 (DKK2), promoting the outgrowth of ectocervical stem cells. Our data indicate that homeostasis at the transition zone results from divergent stromal signals, driving the differential proliferation of resident epithelial lineages.

scholarly journals Spatial organisation and homeostasis of epithelial lineages at the gastroesophageal junction is regulated by the divergent Wnt mucosal microenvironment

2021 ◽  
Author(s):  
Naveen Kumar ◽  
Rajendra Kumar Gurumurthy ◽  
Pon Ganish Prakash ◽  
Shilpa Mary Kurian ◽  
Christian Wentland ◽  
...  
Keyword(s):  
Stem Cells ◽  
Single Molecule ◽  
Gastroesophageal Junction ◽  
Lineage Tracing ◽  
Gastric Epithelium ◽  
Spatial Organisation ◽  
Stromal Compartment ◽  
Fundamental Process ◽  
Resident Stem Cells

The gastroesophageal junction (GEJ), where squamous and columnar epithelia meet, is a hotspot for Barretts metaplasia development, dysbiosis and carcinogenesis. However, the mechanisms regulating GEJ homeostasis remain unclear. Here, by employing organoids, bulk and single-cell transcriptomics, single-molecule RNA in situ hybridisations and lineage tracing, we identified the spatial organisation of the epithelial, stromal compartment and the regulators that maintain the normal GEJ homeostasis. During development, common KRT8 progenitors generate committed unilineage p63/KRT5-squamous and KRT8-columnar stem cells responsible for the regeneration of postnatal esophagus and gastric epithelium that meet at GEJ. A unique spatial distribution of Wnt regulators in the underlying stromal compartment of these stem cells creates diverging Wnt microenvironments at GEJ and supports their differential regeneration. Further, we show that these tissue-resident stem cells do not possess the plasticity to transdifferentiate to the other lineage with the altered Wnt signals. Our study provides invaluable insights into the fundamental process of GEJ homeostasis and is crucial for understanding disease development.

scholarly journals Spatial organization of the somatosensory cortex revealed by cyclic smFISH

2018 ◽  
Author(s):  
Simone Codeluppi ◽  
Lars E. Borm ◽  
Amit Zeisel ◽  
Gioele La Manno ◽  
Josina A. van Lunteren ◽  
...  
Keyword(s):  
Single Cell ◽  
Somatosensory Cortex ◽  
Single Molecule ◽  
Spatial Organization ◽  
Spatial Information ◽  
Cell Types ◽  
Cellular Organization ◽  
New Methods ◽  
The Brain

The global efforts towards the creation of a molecular census of the brain using single-cell transcriptomics is generating a large catalog of molecularly defined cell types lacking spatial information. Thus, new methods are needed to map a large number of cell-specific markers simultaneously on large tissue areas. Here, we developed a cyclic single molecule fluorescence in situ hybridization methodology and defined the cellular organization of the somatosensory cortex using markers identified by single-cell transcriptomics.

scholarly journals Single-molecule imaging suggests compact and spliceosome dependent organization of long introns

2021 ◽  
Author(s):  
Srivathsan Adivarahan ◽  
A.M.S.Kalhara Abeykoon ◽  
Daniel Zenklusen
Keyword(s):  
Rna Polymerase Ii ◽  
Single Molecule ◽  
Splice Site ◽  
Spatial Organization ◽  
Spliceosome Assembly ◽  
U2 Snrnp ◽  
Rna In Situ Hybridization ◽  
Intron Removal ◽  
In Cells

Intron removal from pre-mRNAs is a critical step in the processing of RNA polymerase II transcripts, required to create translation competent mRNAs. In humans, introns account for large portions of the pre-mRNA, with intronic sequences representing about 95% of most pre-mRNA. Intron length varies considerably; introns can be as short as a few to hundreds of thousands of nucleotides in length. How nascent long intronic RNA is organized during transcription to facilitate the communication between 5′ and 3′ splice-sites required for spliceosome assembly however is still poorly understood. Here, we use single-molecule fluorescent RNA in situ hybridization (smFISH) to investigate the spatial organization of co- and post-transcriptional long introns in cells. Using two long introns within the POLA1 pre-mRNA as a model, we show that introns are packaged into compact assemblies, and when fully transcribed, are organized in a looped conformation with their ends in proximity. This organization is observed for nascent and nucleoplasmic pre-mRNAs and requires spliceosome assembly, as disruption of U2 snRNP binding results in introns with separated 5′ and 3′ ends. Moreover, interrogating the spatial organization of partially transcribed co-transcriptional POLA1 intron 35 indicates that the 5′ splice site is maintained proximal to the 3′ splice site during transcription, supporting a model that 5′ splice site tethering to the elongating polymerase might contribute to spliceosome assembly at long introns. Together, our study reveals details of intron and pre-mRNA organization in cells and provides a tool to investigate mechanisms of splicing for long introns.

scholarly journals A stem cell population at the anorectal junction maintains homeostasis and participates in tissue regeneration

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Louciné Mitoyan ◽  
Véronique Chevrier ◽  
Hector Hernandez-Vargas ◽  
Alexane Ollivier ◽  
Zeinab Homayed ◽  
...  
Keyword(s):  
Transition Zone ◽  
Tissue Regeneration ◽  
Adult Stem Cells ◽  
Tissue Injury ◽  
Lineage Tracing ◽  
The Body ◽  
Stem Cell Population ◽  
Basal Cells ◽  
Transition Zones ◽  
Anorectal Junction

AbstractAt numerous locations of the body, transition zones are localized at the crossroad between two types of epithelium and are frequently associated with neoplasia involving both type of tissues. These transition zones contain cells expressing markers of adult stem cells that can be the target of early transformation. The mere fact that transition zone cells can merge different architecture with separate functions implies for a unique plasticity that these cells must display in steady state. However, their roles during tissue regeneration in normal and injured state remain unknown. Here, by using in vivo lineage tracing, single-cell transcriptomics, computational modeling and a three-dimensional organoid culture system of transition zone cells, we identify a population of Krt17+ basal cells with multipotent properties at the squamo-columnar anorectal junction that maintain a squamous epithelium during normal homeostasis and can participate in the repair of a glandular epithelium following tissue injury.

scholarly journals DnaA, the Initiator of Escherichia coliChromosomal Replication, Is Located at the Cell Membrane

2000 ◽  
Vol 182 (9) ◽  
pp. 2604-2610 ◽  
Author(s):  
Gillian Newman ◽  
Elliott Crooke
Keyword(s):  
Cell Membrane ◽  
Spatial Organization ◽  
Active Form ◽  
Chromosomal Replication ◽  
E Coli ◽  
Dnaa Protein ◽  
Section Electron ◽  
Acidic Phospholipids

ABSTRACT Given the lack of a nucleus in prokaryotic cells, the significance of spatial organization in bacterial chromosome replication is only beginning to be fully appreciated. DnaA protein, the initiator of chromosomal replication in Escherichia coli, is purified as a soluble protein, and in vitro it efficiently initiates replication of minichromosomes in membrane-free DNA synthesis reactions. However, its conversion from a replicatively inactive to an active form in vitro occurs through its association with acidic phospholipids in a lipid bilayer. To determine whether the in situ residence of DnaA protein is cytoplasmic, membrane associated, or both, we examined the cellular location of DnaA using immunogold cryothin-section electron microscopy and immunofluorescence. Both of these methods revealed that DnaA is localized at the cell membrane, further suggesting that initiation of chromosomal replication in E. coli is a membrane-affiliated event.

scholarly journals Germline Transformation of Drosophila virilis With the Transposable Element mariner

Genetics ◽  
1996 ◽  
Vol 143 (1) ◽  
pp. 365-374 ◽  
Author(s):  
Allan R Lohe ◽  
Daniel L Hartl
Keyword(s):  
Transposable Element ◽  
Common Ancestor ◽  
Pcr Amplification ◽  
Drosophila Virilis ◽  
Cell Region ◽  
Diverse Species ◽  
Drosophila Mauritiana ◽  
Characteristic 2 ◽  
In Situ Hybridizations

Abstract An important goal in molecular genetics has been to identify a transposable element that might serve as an efficient transformation vector in diverse species of insects. The transposable element mariner occurs naturally in a wide variety of insects. Although virtually all mariner elements are nonfunctional, the Mosl element isolated from Drosophila mauritiana is functional. Mosl was injected into the pole-cell region of embryos of D. virilis, which last shared a common ancestor with D. mauritiana 40 million years ago. Mosl PCR fragments were detected in several pools of DNA from progeny of injected animals, and backcross lines were established. Because Go lines were pooled, possibly only one transformation event was actually obtained, yielding a minimum frequency of 4%. Mosl segregated in a Mendelian fashion, demonstrating chromosomal integration. The copy number increased by spontaneous mobilization. In situ hybridization confirmed multiple polymorphic locations of Mosl. Integration results in a characteristic 2-bp TA duplication. One Mosl element integrated into a tandem array of 370-bp repeats. Some copies may have integrated into heterochromatin, as evidenced by their ability to support PCR amplification despite absence of a signal in Southern and in situ hybridizations.

scholarly journals Petiole-Lamina Transition Zone: A Functionally Crucial but Often Overlooked Leaf Trait

Plants ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 774
Author(s):  
Max Langer ◽  
Thomas Speck ◽  
Olga Speck
Keyword(s):  
Transition Zone ◽  
Three Dimensional ◽  
Spatial Arrangement ◽  
Body Plan ◽  
The Body ◽  
Vascular Bundles ◽  
Cross Sectional ◽  
Thin Sections ◽  
Transition Zones ◽  
The Cross

Although both the petiole and lamina of foliage leaves have been thoroughly studied, the transition zone between them has often been overlooked. We aimed to identify objectively measurable morphological and anatomical criteria for a generally valid definition of the petiole–lamina transition zone by comparing foliage leaves with various body plans (monocotyledons vs. dicotyledons) and spatial arrangements of petiole and lamina (two-dimensional vs. three-dimensional configurations). Cross-sectional geometry and tissue arrangement of petioles and transition zones were investigated via serial thin-sections and µCT. The changes in the cross-sectional geometries from the petiole to the transition zone and the course of the vascular bundles in the transition zone apparently depend on the spatial arrangement, while the arrangement of the vascular bundles in the petioles depends on the body plan. We found an exponential acropetal increase in the cross-sectional area and axial and polar second moments of area to be the defining characteristic of all transition zones studied, regardless of body plan or spatial arrangement. In conclusion, a variety of terms is used in the literature for describing the region between petiole and lamina. We prefer the term “petiole–lamina transition zone” to underline its three-dimensional nature and the integration of multiple gradients of geometry, shape, and size.

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