scholarly journals Phosphoglucomutase 1: a gene with two promoters and a duplicated first exon

1993 ◽  
Vol 296 (2) ◽  
pp. 417-422 ◽  
Author(s):  
W Putt ◽  
J H Ives ◽  
M Hollyoake ◽  
D A Hopkinson ◽  
D B Whitehouse ◽  
...  
Keyword(s):  
Cell Types ◽  
Fast Muscle ◽  
Intragenic Recombination ◽  
Amino Acid Residues ◽  
Sequence Comparisons ◽  
Exon 2 ◽  
Extra Amino Acid ◽  
Alternatively Spliced ◽  
The Common ◽  
A Site

In view of its central role in glycolysis and gluconeogenesis and its polymorphic genetic variability, the phosphoglucomutase 1 (PGM1) gene in man has been the target of protein structural studies and genetic analysis for more than 25 years. We have now isolated genomic clones containing the complete PGM1 gene and have shown that it spans over 65 kb and contains 11 exons. We have also shown that the sites of the two mutations which form the molecular basis for the common PGM1 protein polymorphism lie in exons 4 and 8 and are 18 kb apart. Within this region there is a site of intragenic recombination. We have discovered two alternatively spliced first exons, one of which, exon 1A, is transcribed in a wide variety of cell types; the other, exon 1B, is transcribed in fast muscle. Exon 1A is transcribed from a promoter which has the structural hallmarks of a housekeeping promoter but lies more than 35 kb upstream of exon 2. Exon 1B lies 6 kb upstream of exon 2 within the large first intron of the ubiquitously expressed PGM1 transcript. The fast-muscle form of PGM1 is characterized by 18 extra amino acid residues at its N-terminal end. Sequence comparisons show that exons 1A and 1B are structurally related and have arisen by duplication.

scholarly journals Exon-skipping in BCR/ABL is induced by ABL exon 2

2000 ◽  
Vol 348 (1) ◽  
pp. 63-69 ◽  
Author(s):  
Brian D. LICHTY ◽  
Suzanne KAMEL-REID
Keyword(s):  
Alternative Splicing ◽  
Genomic Dna ◽  
Chronic Myelogenous Leukaemia ◽  
Fusion Gene ◽  
Exon Skipping ◽  
Cell Types ◽  
Myelogenous Leukaemia ◽  
Sequence Element ◽  
Exon 2 ◽  
Alternatively Spliced

The BCR/ABL fusion gene is pathognomonic for chronic myelogenous leukaemia (CML). We have previously reported alternative splicing of BCR/ABL, as indicated by the detection of both p190- and p210-encoding transcripts, in about 60% of CML patient samples. These exon-skipping events involved the joining of ABL exon 2 to variable upstream BCR exons. Similarly, ABL exon 2 is alternatively spliced to either of two upstream ABL exons (1a or 1b) in c-ABL. We have constructed BCR and BCR/ABL minigenes to study this phenomenon in more detail. These constructs were transfected into various cell types and splicing was assessed by reverse transcriptase PCR. Whereas the basic BCR minigene expressed exon-inclusive transcripts only, insertion of genomic DNA spanning ABL exon 2 induced exon-skipping but only when expressed in the CML cell lines K562 and EM3. In this study we localized the required sequence element to ABL exon 2 itself. These results mimic the splicing phenotype displayed by most CML patients. We propose a model where a trans-factor present in some CML cells interacts with ABL exon 2 pre-mRNA to promote skipping of upstream BCR exons.

CELL AND TISSUE THERAPY OF HEART

2019 ◽  
pp. 77-84
Keyword(s):  
Extracellular Matrix ◽  
Cell Types ◽  
Heart Tissue ◽  
Decellularized Extracellular Matrix ◽  
Tissue Therapy ◽  
Essential Components ◽  
Long Time ◽  
Different Populations ◽  
A Site ◽  
Recipient Tissue

The strategy of heart tissue engineering is simple enough: first remove all the cells from a organ then take the protein scaffold left behind and repopulate it with stem cells immunologically matched to the patient in need. While various suc- cessful methods for decellularization have been developed, and the feasibility of using decellularized whole hearts and extracellular matrix to support cells has been demonstrated, the reality of creating whole hearts for transplantation and of clinical application of decellularized extracellular matrix-based scaffolds will require much more research. For example, further investigations into how lineage-restricted progenitors repopulate the decellularized heart and differentiate in a site-specific manner into different populations of the native heart would be essential. The scaffold heart does not have to be human. Pig hearts carries all the essential components of the extracellular matrix. Through trial and error, scaling up the concentration, timing and pressure of the detergents, researchers have refined the decellularization process on hundreds of hearts and other organs, but this is only the first step. Further, the framework must be populated with human cells. Most researchers in the field use a mixture of two or more cell types, such as endothelial precursor cells to line blood vessels and muscle progenitors to seed the walls of the chambers. The final challenge is one of the hardest: vasculariza- tion, placing a engineered heart into a living animal, integration with the recipient tissue, and keeping it beating for a long time. Much remains to be done before a bioartificial heart is available for transplantation in humans.

A mutation of the β-domain in POU1F1 causes pituitary deficiency due to dominant PIT-1β expression

2021 ◽  
Author(s):  
Shigeru Suzuki ◽  
Kumihiro Matsuo ◽  
Yoshiya Ito ◽  
Atsushi Kobayashi ◽  
Takahide Kokumai ◽  
...  
Keyword(s):  
Hormone Deficiency ◽  
Luciferase Reporter ◽  
Heterozygous Mutation ◽  
Pituitary Hormone ◽  
Pituitary Cells ◽  
Transactivation Domain ◽  
Exon 2 ◽  
Pituitary Development ◽  
Long Term Follow Up ◽  
Alternatively Spliced

Background: POU1F1 encodes both PIT-1α, which plays pivotal roles in pituitary development and GH, PRL and TSHB expression, and the alternatively spliced isoform PIT-1β, which contains an insertion of 26-amino acids (β-domain) in the transactivation domain of PIT-1α due to the use of an alternative splice acceptor at the end of the first intron. PIT-1β is expressed at much lower levels than PIT-1α and represses endogenous PIT-1α transcriptional activity. Although POU1F1 mutations lead to combined pituitary hormone deficiency (CPHD), no patients with β-domain mutations have been reported. Results: Here, we report that a three-generation family exhibited different degrees of CPHD, including growth hormone deficiency with intrafamilial variability of prolactin/TSH insufficiency and unexpected prolactinoma occurrence. The CPHD was due to a novel POU1F1 heterozygous variant (c.143-69T>G) in intron 1 of PIT-1α (RefSeq number NM_000306) or as c.152T>G (p.Ile51Ser) in exon 2 of PIT-1β (NM_001122757). Gene splicing experiments showed that this mutation yielded the PIT-1β transcript without other transcripts. Lymphocyte PIT-1β mRNA expression was significantly higher in the patients with the heterozygous mutation than a control. A luciferase reporter assay revealed that the PIT-1β-Ile51Ser mutant repressed PIT-1α and abolished transactivation capacity for the rat prolactin promoter in GH3 pituitary cells. Conclusions: We describe, for the first time, that PIT-1β mutation can cause CPHD through a novel genetic mechanism, such as PIT-1β overexpression, and that POU1F1 mutation might be associated with a prolactinoma. Analysis of new patients and long-term follow-up are needed to clarify the characteristics of PIT-1β mutations.

scholarly journals Cloning and functional characterization of the human fractalkine receptor promoter regions

2002 ◽  
Vol 368 (3) ◽  
pp. 753-760 ◽  
Author(s):  
Alexandre GARIN ◽  
Philippe PELLET ◽  
Philippe DETERRE ◽  
Patrice DEBRÉ ◽  
Christophe COMBADIÈRE
Keyword(s):  
Functional Characterization ◽  
Cell Types ◽  
Promoter Regions ◽  
Exon 2 ◽  
Reading Frame ◽  
Fractalkine Receptor ◽  
A Cell ◽  
Exon 4 ◽  
Coronary Events

We have previously shown that reduced expression of the fractalkine receptor, CX3CR1, is correlated with rapid HIV disease progression and with reduced susceptibility to acute coronary events. In order to elucidate the mechanisms underlying transcriptional regulation of CX3CR1 expression, we structurally and functionally characterized the CX3CR1 gene. It consists of four exons and three introns spanning over 18kb. Three transcripts are produced by splicing the three untranslated exons with exon 4, which contains the complete open reading frame. The transcript predominantly found in leucocytes corresponds to the splicing of exon 2 with exon 4. Transcripts corresponding to splicing of exons 1 and 4 are less abundant in leucocytes and splicing of exons 3 and 4 are rare longer transcripts. A constitutive promoter activity was found in the regions extending upstream from untranslated exons 1 and 2. Interestingly, exons 1 and 2 enhanced the activity of their respective promoters in a cell-specific manner. These data show that the CX3CR1 gene is controlled by three distinct promoter regions, which are regulated by their respective untranslated exons and that lead to the transcription of three mature messengers. This highly complex regulation may allow versatile and precise expression of CX3CR1 in various cell types.

scholarly journals A Nonerythroid Isoform of Protein 4.1R Interacts with Components of the Contractile Apparatus in Skeletal Myofibers

2000 ◽  
Vol 11 (11) ◽  
pp. 3805-3817 ◽  
Author(s):  
Aikaterini Kontrogianni-Konstantopoulos ◽  
Shu-Ching Huang ◽  
Edward J. Benz
Keyword(s):  
Skeletal Muscle ◽  
Protein S ◽  
Binding Assays ◽  
Adult Skeletal Muscle ◽  
Exon 2 ◽  
Alternatively Spliced ◽  
Translation Initiation Codon ◽  
Protein 4.1R

The ∼80-kDa erythroid 4.1R protein is a major component of the erythrocyte cytoskeleton, where it links transmembrane proteins to the underlying spectrin/actin complexes. A diverse collection of 4.1R isoforms has been described in nonerythroid cells, ranging from ∼30 to ∼210 kDa. In the current study, we identified the number and primary structure of 4.1R isoforms expressed in adult skeletal muscle and characterized the localization patterns of 4.1R message and protein. Skeletal muscle 4.1R appears to originate solely from the upstream translation initiation codon (AUG-1) residing in exon 2′. Combinations of alternatively spliced downstream exons generate an array of distinct 4.1R spliceoforms. Two major isoform classes of ∼105/110 and ∼135 kDa are present in muscle homogenates. 4.1R transcripts are distributed in highly ordered signal stripes, whereas 4.1R protein(s) decorate the sarcoplasm in transverse striations that are in register with A-bands. An ∼105/110-kDa 4.1R isoform appears to occur in vivo in a supramolecular complex with major sarcomeric proteins, including myosin, α-actin, and α-tropomyosin. In vitro binding assays showed that 4.1R may interact directly with the aforementioned contractile proteins through its 10-kDa domain. All of these observations suggest a topological model whereby 4.1R may play a scaffolding role by anchoring the actomyosin myofilaments and possibly modulating their displacements during contraction/relaxation.

scholarly journals Understanding axial progenitor biology in vivo and in vitro

Development ◽  
2021 ◽  
Vol 148 (4) ◽  
pp. dev180612
Author(s):  
Filip J. Wymeersch ◽  
Valerie Wilson ◽  
Anestis Tsakiridis
Keyword(s):  
Spinal Cord ◽  
Vertebral Column ◽  
Cell Types ◽  
Disease Modelling ◽  
Recent Developments ◽  
The Common ◽  
Key Features ◽  
Trunk Musculature

ABSTRACTThe generation of the components that make up the embryonic body axis, such as the spinal cord and vertebral column, takes place in an anterior-to-posterior (head-to-tail) direction. This process is driven by the coordinated production of various cell types from a pool of posteriorly-located axial progenitors. Here, we review the key features of this process and the biology of axial progenitors, including neuromesodermal progenitors, the common precursors of the spinal cord and trunk musculature. We discuss recent developments in the in vitro production of axial progenitors and their potential implications in disease modelling and regenerative medicine.

scholarly journals Identification of Amino Acid Residues in the α, β, and γ Subunits of the Epithelial Sodium Channel (ENaC) Involved in Amiloride Block and Ion Permeation

1997 ◽  
Vol 109 (1) ◽  
pp. 15-26 ◽  
Author(s):  
Laurent Schild ◽  
Estelle Schneeberger ◽  
Ivan Gautschi ◽  
Dmitri Firsov
Keyword(s):  
Amino Acid ◽  
Ion Selectivity ◽  
Site Directed Mutagenesis ◽  
Ion Permeation ◽  
Amino Acid Residues ◽  
Α Subunit ◽  
Na Ions ◽  
Membrane Spanning ◽  
A Site ◽  
Epithelial Sodium Channel Enac

The amiloride-sensitive epithelial Nachannel (ENaC) is a heteromultimeric channel made of three αβγ subunits. The structures involved in the ion permeation pathway have only been partially identified, and the respective contributions of each subunit in the formation of the conduction pore has not yet been established. Using a site-directed mutagenesis approach, we have identified in a short segment preceding the second membrane-spanning domain (the pre-M2 segment) amino acid residues involved in ion permeation and critical for channel block by amiloride. Cys substitutions of Gly residues in β and γ subunits at position βG525 and γG537 increased the apparent inhibitory constant (Ki) for amiloride by >1,000-fold and decreased channel unitary current without affecting ion selectivity. The corresponding mutation S583 to C in the α subunit increased amiloride Ki by 20-fold, without changing channel conducting properties. Coexpression of these mutated αβγ subunits resulted in a nonconducting channel expressed at the cell surface. Finally, these Cys substitutions increased channel affinity for block by externalZn2+ ions, in particular the αS583C mutant showing a Ki for Zn2+of 29 μM. Mutations of residues αW582L or βG522D also increased amiloride Ki, the later mutation generating a Ca2+blocking site located 15% within the membrane electric field. These experiments provide strong evidence that αβγ ENaCs are pore-forming subunits involved in ion permeation through the channel. The pre-M2 segment of αβγ subunits may form a pore loop structure at the extracellular face of the channel, where amiloride binds within the channel lumen. We propose that amiloride interacts with Na+ions at an external Na+binding site preventing ion permeation through the channel pore.

The early development of mystacial vibrissae in the mouse

Development ◽  
1984 ◽  
Vol 83 (1) ◽  
pp. 137-156
Author(s):  
Joan T. Wrenn ◽  
Norman K. Wessells
Keyword(s):  
Flat Surface ◽  
Temporal Correlation ◽  
Cell Types ◽  
Nerve Branch ◽  
Complex Process ◽  
Initial Generation ◽  
Mystacial Vibrissae ◽  
Vertical Row ◽  
Early Mouse ◽  
A Site

The initial generation of the pattern of mystacial vibrissae (whiskers) in the mouse is described. The maxillary process is present in 10-day embryos but has a relatively flat surface. Beginning at approximately 11·5 days, the first sign of vibrissal development is the formation of ridges and grooves on the maxillary and lateral nasal processes. The first vibrissal rudiment to form subsequently appears posterior to the most ventral groove on the maxillary process. It is the most ventral whisker of the posterior, vertical row. The next few rudiments appear: (1) dorsal to the first, also in the vertical row; and (2) anterior to the first, on the ventral-most ridge and in the groove beneath it. Formation of vibrissal rudiments continues in a dorsal and anterior progression usually by an apparent partitioning of the ridges into vibrissal units. The hypothesis that this patterning of mystacial vibrissae might be determined by the pattern of innervation in the early mouse snout was investigated. Nerve trunks and branches are present in the maxillary process well before any sign of vibrissal formation. Because innervation is so widespread there appears to be no immediate temporal correlation between the outgrowth of a nerve branch to a site and the generation of a vibrissa there. Furthermore, at the time just prior to the formation of the first follicle rudiment, there is little or no nerve branching to the presumptive site of that first follicle while branches are found more dorsally where vibrissae will not form until later. Thus, a one-to-one spatial correlation between nerve and follicle sites also appears to be lacking. The developmental changes in ultrastructure within the neurites of the trunks and branches as well as the apparent rearrangements of the nerve trunks suggest that early innervation of the snout is a labile phenomenon and that the vibrissal pattern begins to be established before the neural pattern is completely developed. The results indicate that vibrissal pattern formation is likely to be a complex process relying on the interplay of the cells and tissues involved, rather than on unidirectional instructions from neurons to other cell types.

Bioprinting

2021 ◽  
Author(s):  
Kenneth Douglas
Keyword(s):  
Stem Cells ◽  
Neuromuscular Junctions ◽  
Cell Types ◽  
Passive Movement ◽  
Plain Language ◽  
High Concentration ◽  
Proliferation And Differentiation ◽  
The Common ◽  
Biological Environment ◽  
Lung And Kidney

Abstract: This book describes how bioprinting emerged from 3D printing and details the accomplishments and challenges in bioprinting tissues of cartilage, skin, bone, muscle, neuromuscular junctions, liver, heart, lung, and kidney. It explains how scientists are attempting to provide these bioprinted tissues with a blood supply and the ability to carry nerve signals so that the tissues might be used for transplantation into persons with diseased or damaged organs. The book presents all the common terms in the bioprinting field and clarifies their meaning using plain language. Readers will learn about bioink—a bioprinting material containing living cells and supportive biomaterials. In addition, readers will become at ease with concepts such as fugitive inks (sacrificial inks used to make channels for blood flow), extracellular matrices (the biological environment surrounding cells), decellularization (the process of isolating cells from their native environment), hydrogels (water-based substances that can substitute for the extracellular matrix), rheology (the flow properties of a bioink), and bioreactors (containers to provide the environment cells need to thrive and multiply). Further vocabulary that will become familiar includes diffusion (passive movement of oxygen and nutrients from regions of high concentration to regions of low concentration), stem cells (cells with the potential to develop into different bodily cell types), progenitor cells (early descendants of stem cells), gene expression (the process by which proteins develop from instructions in our DNA), and growth factors (substances—often proteins—that stimulate cell growth, proliferation, and differentiation). The book contains an extensive glossary for quick reference.

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