Determination of the migratory capacity of embryonic cortical cells lacking the transcription factor Pax-6

Development ◽  
1997 ◽  
Vol 124 (24) ◽  
pp. 5087-5096 ◽  
Author(s):  
D. Caric ◽  
D. Gooday ◽  
R.E. Hill ◽  
S.K. McConnell ◽  
D.J. Price
Keyword(s):  
Transcription Factor ◽  
Ventricular Zone ◽  
Wild Type Mouse ◽  
Wild Type ◽  
Cortical Cells ◽  
Developmental Potential ◽  
Neuronal Precursors ◽  
Cortical Precursors

The cerebral cortex forms by the orderly migration and subsequent differentiation of neuronal precursors generated in the proliferative ventricular zone. We studied the role of the transcription factor Pax-6, which is expressed in the ventricular zone, in cortical development. Embryos homozygous for a mutation of Pax-6 (Small eye; Sey) had abnormalities suggesting defective migration of late-born cortical precursors. When late-born Sey/Sey precursors were transplanted into wild-type embryonic rat cortex, they showed similar integrative, migrational and differentiative abilities to those of transplanted wild-type mouse precursors. These results suggest that postmitotic cortical cells do not need Pax-6 to acquire the capacity to migrate and differentiate, but that Pax-6 generates a cortical environment that permits later-born precursors to express their full developmental potential.

scholarly journals Advanced Spectroscopy and APBS Modeling for Determination of the Role of His190 and Trp103 in Mouse Thymidylate Synthase Interaction with Selected dUMP Analogues

2021 ◽  
Vol 22 (5) ◽  
pp. 2661
Author(s):  
Małgorzata Prokopowicz ◽  
Adam Jarmuła ◽  
Yannick Casamayou-Boucau ◽  
Fiona Gordon ◽  
Alan Ryder ◽  
...  
Keyword(s):  
Electrostatic Potential ◽  
Thymidylate Synthase ◽  
Wild Type Mouse ◽  
Inhibitory Mechanism ◽  
Wild Type ◽  
Theoretical Investigations ◽  
Ligand Interactions ◽  
Poisson Boltzmann

A homo-dimeric enzyme, thymidylate synthase (TS), has been a long-standing molecular target in chemotherapy. To further elucidate properties and interactions with ligands of wild-type mouse thymidylate synthase (mTS) and its two single mutants, H190A and W103G, spectroscopic and theoretical investigations have been employed. In these mutants, histidine at position 190 and tryptophan at position 103 are substituted with alanine and glycine, respectively. Several emission-based spectroscopy methods used in the paper demonstrate an especially important role for Trp 103 in TS ligands binding. In addition, the Advanced Poisson–Boltzmann Solver (APBS) results show considerable differences in the distribution of electrostatic potential around Trp 103, as compared to distributions observed for all remaining Trp residues in the mTS family of structures. Together, spectroscopic and APBS results reveal a possible interplay between Trp 103 and His190, which contributes to a reduction in enzymatic activity in the case of H190A mutation. Comparison of electrostatic potential for mTS complexes, and their mutants, with the substrate, dUMP, and inhibitors, FdUMP and N4-OH-dCMP, suggests its weaker influence on the enzyme–ligand interactions in N4OH-dCMP-mTS compared to dUMP-mTS and FdUMP-mTS complexes. This difference may be crucial for the explanation of the ”abortive reaction” inhibitory mechanism of N4OH-dCMP towards TS. In addition, based on structural analyses and the H190A mutant capacity to form a denaturation-resistant complex with N4-OH-dCMP in the mTHF-dependent reaction, His190 is apparently responsible for a strong preference of the enzyme active center for the anti rotamer of the imino inhibitor form.

scholarly journals The Na+/H+ exchanger isoform 3 is required for active paracellular and transcellular Ca2+ transport across murine cecum

2013 ◽  
Vol 305 (4) ◽  
pp. G303-G313 ◽  
Author(s):  
Juraj Rievaj ◽  
Wanling Pan ◽  
Emmanuelle Cordat ◽  
R. Todd Alexander
Keyword(s):  
Water Movement ◽  
Water Flux ◽  
Wild Type Mouse ◽  
Wild Type ◽  
Rt Pcr ◽  
Ussing Chambers ◽  
Voltage Clamps ◽  
Paracellular Absorption ◽  
Mouse Intestine

Intestinal calcium (Ca2+) absorption occurs via paracellular and transcellular pathways. Although the transcellular route has been extensively studied, mechanisms mediating paracellular absorption are largely unexplored. Unlike passive diffusion, secondarily active paracellular Ca2+ uptake occurs against an electrochemical gradient with water flux providing the driving force. Water movement is dictated by concentration differences that are largely determined by Na+ fluxes. Consequently, we hypothesized that Na+ absorption mediates Ca2+ flux. NHE3 is central to intestinal Na+ absorption. NHE3 knockout mice (NHE3−/−) display impaired intestinal Na+, water, and Ca2+ absorption. However, the mechanism mediating this latter abnormality is not clear. To investigate this, we used Ussing chambers to measure net Ca2+ absorption across different segments of wild-type mouse intestine. The cecum was the only segment with net Ca2+ absorption. Quantitative RT-PCR measurements revealed cecal expression of all genes implicated in intestinal Ca2+ absorption, including NHE3. We therefore employed this segment for further studies. Inhibition of NHE3 with 100 μM 5-( N-ethyl- N-isopropyl) amiloride decreased luminal-to-serosal and increased serosal-to-luminal Ca2+ flux. NHE3−/− mice had a >60% decrease in luminal-to-serosal Ca2+ flux. Ussing chambers experiments under altered voltage clamps (−25, 0, +25 mV) showed decreased transcellular and secondarily active paracellular Ca2+ absorption in NHE3−/− mice relative to wild-type animals. Consistent with this, cecal Trpv6 expression was diminished in NHE3−/− mice. Together these results implicate NHE3 in intestinal Ca2+ absorption and support the theory that this is, at least partially, due to the role of NHE3 in Na+ and water absorption.

scholarly journals Novel Role of MCP-Induced Protein 1 in Ischemic Brain Damage in Animals of Transient Focal Ischemia

2018 ◽  
Author(s):  
Zhuqing Jin ◽  
Jian Liang ◽  
Jiaqi Li ◽  
Pappachan E. Kolattukudy
Keyword(s):  
Cerebral Ischemia ◽  
Matrix Metalloproteinases ◽  
Tight Junction ◽  
Western Blot ◽  
Blood Brain Barrier ◽  
Wild Type Mouse ◽  
Brain Barrier ◽  
Wild Type ◽  
Blood Brain

Focal cerebral ischemia can lead to blood-brain barrier (BBB) breakdown, which is implicated in neuroinflammation and elevation of matrix metalloproteinases (MMPs). The role of the anti-inflammatory protein, monocyte chemotactic protein–induced protein 1 (MCPIP1) plays in the injury of BBB in stroke has not yet been reported. This study was conducted to identify and characterize the role MCPIP1 plays in BBB breakdown. Transient middle cerebral artery occlusion (MCAO) is induced in both wild-type and Mcpip1-/- mice for 2 hours of occlusion periods followed by reperfusion for 24 or 48 hours. BBB permeability was measured by FITC-dextran extravasation, MMP-9/3 expression was analyzed by western blot, and claudin-5 and zonula occludens-1 (ZO-1) were analyzed by immunohistochemistry and western blot. After MCAO in wild type mouse is induced, there is significantly increase in MCPIP1 mRNA and protein levels. Absence of MCPIP1 leaded to significant increase in FITC-dextran leakage in peri-infarct brain, significant upregulation of MMP-9, MMP-3 and reduced levels of tight junction components, claudin-5 and ZO-1 in the brain after MCAO. Our data demonstrate that absence of MCPIP1 exacerbates ischemia-induced blood-brain barrier disruption by enhancing the expression of matrix metalloproteinases and degradation of tight junction proteins. Overall data indicate that MCPIP1 is important protective role against BBB disruption in cerebral ischemia.

scholarly journals Diagnostic and Treatment Approaches Involving Transthyretin in Amyloidogenic Diseases

2019 ◽  
Vol 20 (12) ◽  
pp. 2982 ◽  
Author(s):  
Gil Yong Park ◽  
Angelo Jamerlan ◽  
Kyu Hwan Shim ◽  
Seong Soo A. An
Keyword(s):  
Cerebrospinal Fluid ◽  
Genetic Mutations ◽  
Hormone Binding ◽  
Wild Type ◽  
Autonomic Motor ◽  
Hormone Binding Protein ◽  
The Brain ◽  
Tetrameric Form

Transthyretin (TTR) is a thyroid hormone-binding protein which transports thyroxine from the bloodstream to the brain. The structural stability of TTR in tetrameric form is crucial for maintaining its original functions in blood or cerebrospinal fluid (CSF). The altered structure of TTR due to genetic mutations or its deposits due to aggregation could cause several deadly diseases such as cardiomyopathy and neuropathy in autonomic, motor, and sensory systems. The early diagnoses for hereditary amyloid TTR with cardiomyopathy (ATTR-CM) and wild-type amyloid TTR (ATTRwt) amyloidosis, which result from amyloid TTR (ATTR) deposition, are difficult to distinguish due to the close similarities of symptoms. Thus, many researchers investigated the role of ATTR as a biomarker, especially its potential for differential diagnosis due to its varying pathogenic involvement in hereditary ATTR-CM and ATTRwt amyloidosis. As a result, the detection of ATTR became valuable in the diagnosis and determination of the best course of treatment for ATTR amyloidoses. Assessing the extent of ATTR deposition and genetic analysis could help in determining disease progression, and thus survival rate could be improved following the determination of the appropriate course of treatment for the patient. Here, the perspectives of ATTR in various diseases were presented.

scholarly journals Complement Receptor 1 Expression on Mouse Erythrocytes Mediates Clearance of Streptococcus pneumoniae by Immune Adherence

2010 ◽  
Vol 78 (7) ◽  
pp. 3129-3135 ◽  
Author(s):  
Jie Li ◽  
Jennifer P. Wang ◽  
Ionita Ghiran ◽  
Anna Cerny ◽  
Alexander J. Szalai ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Direct Evidence ◽  
Wild Type Mouse ◽  
Complement Receptor ◽  
Blood Clearance ◽  
Wild Type ◽  
Immune Adherence ◽  
Complement Receptor 1

ABSTRACT Complement-containing immune complexes can be presented to phagocytes by human erythrocytes bearing complement receptor 1 (CR1). Although this has long been assumed to be a mechanism by which humans are able to protect themselves from “extracellular” bacteria such as pneumococci, there is little direct evidence. In these studies we have investigated this question by comparing results for erythrocytes from transgenic mice expressing human CR1 on their erythrocytes to the results for wild-type mouse erythrocytes that do not express CR1. We demonstrate that human CR1 expression on murine erythrocytes allows immune adherence to beads opsonized with either mouse or human serum as a source of complement. The role of CR1 in immune adherence was supported by studies showing that it was blocked by the addition of antibody to human CR1. Furthermore, human CR1 expression enhances the immune adherence of opsonized pneumococci to erythrocytes in vitro, and the pneumococci attached to erythrocytes via CR1 can be transferred in vitro to live macrophages. Even more importantly, we observed that if complement-opsonized pneumococci are injected intravenously with CR1+ mouse erythrocytes into wild-type mice (after a short in vitro incubation), they are cleared faster than opsonized pneumococci similarly injected with wild-type mouse erythrocytes. Finally, we have shown that the intravenous (i.v.) injection of pneumococci into CR1+ mice also results in more rapid blood clearance than in wild-type mice. These data support that immune adherence via CR1 on erythrocytes likely plays an important role in the clearance of opsonized bacteria from human blood.

scholarly journals Regulation of the Later Stages of Nodulation Stimulated by IPD3/CYCLOPS Transcription Factor and Cytokinin in Pea Pisum sativum L.

Plants ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 56
Author(s):  
Elizaveta S. Rudaya ◽  
Polina Yu. Kozyulina ◽  
Olga A. Pavlova ◽  
Alexandra V. Dolgikh ◽  
Alexandra N. Ivanova ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Comparative Analysis ◽  
Pisum Sativum ◽  
Nodule Development ◽  
Rna Seq ◽  
Wild Type ◽  
Development Stages ◽  
Pisum Sativum L ◽  
Pea Mutant

The IPD3/CYCLOPS transcription factor was shown to be involved in the regulation of nodule primordia development and subsequent stages of nodule differentiation. In contrast to early stages, the stages related to nodule differentiation remain less studied. Recently, we have shown that the accumulation of cytokinin at later stages may significantly impact nodule development. This conclusion was based on a comparative analysis of cytokinin localization between pea wild type and ipd3/cyclops mutants. However, the role of cytokinin at these later stages of nodulation is still far from understood. To determine a set of genes involved in the regulation of later stages of nodule development connected with infection progress, intracellular accommodation, as well as plant tissue and bacteroid differentiation, the RNA-seq analysis of pea mutant SGEFix--2 (sym33) nodules impaired in these processes compared to wild type SGE nodules was performed. To verify cytokinin’s influence on late nodule development stages, the comparative RNA-seq analysis of SGEFix--2 (sym33) mutant plants treated with cytokinin was also conducted. Findings suggest a significant role of cytokinin in the regulation of later stages of nodule development.

scholarly journals Transcriptional network orchestrating regional patterning of cortical progenitors

2021 ◽  
Vol 118 (51) ◽  
pp. e2024795118
Author(s):  
Athéna R. Ypsilanti ◽  
Kartik Pattabiraman ◽  
Rinaldo Catta-Preta ◽  
Olga Golonzhka ◽  
Susan Lindtner ◽  
...  
Keyword(s):  
Stem Cells ◽  
Transcription Factor ◽  
Ventricular Zone ◽  
Integrated Approach ◽  
Mutant Mice ◽  
Gene Interactions ◽  
Wild Type ◽  
Dna Accessibility ◽  
Mouse Cortex ◽  
Radial Glial

We uncovered a transcription factor (TF) network that regulates cortical regional patterning in radial glial stem cells. Screening the expression of hundreds of TFs in the developing mouse cortex identified 38 TFs that are expressed in gradients in the ventricular zone (VZ). We tested whether their cortical expression was altered in mutant mice with known patterning defects (Emx2, Nr2f1, and Pax6), which enabled us to define a cortical regionalization TF network (CRTFN). To identify genomic programming underlying this network, we performed TF ChIP-seq and chromatin-looping conformation to identify enhancer–gene interactions. To map enhancers involved in regional patterning of cortical progenitors, we performed assays for epigenomic marks and DNA accessibility in VZ cells purified from wild-type and patterning mutant mice. This integrated approach has identified a CRTFN and VZ enhancers involved in cortical regional patterning in the mouse.

PAX5/TEL Causes down Regulation of CD19 and TGFbeta Resistance in PreBI Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1416-1416
Author(s):  
Grazia Fazio ◽  
Chiara Palmi ◽  
Greta Giordano Attianese ◽  
Andrea Biondi ◽  
Antonius Rolink ◽  
...  
Keyword(s):  
Growth Rate ◽  
Cell Proliferation ◽  
Transcription Factor ◽  
Cell Differentiation ◽  
B Cell ◽  
Chimeric Protein ◽  
B Cell Differentiation ◽  
Wild Type ◽  
Down Regulation

Abstract The PAX5/TEL chimeric gene was cloned from the translocation t(9;12)(q11;p13) in an ALL patient. Recent data indicate that the PAX5/TEL fusion defines the cytogenetic entity dic(9;12)(p13;p13), which accounts for about 1% of childhood ALL, almost exclusively B-progenitor ALL. PAX5/TEL is likely to be an aberrant transcription factor, resulting from joining the 5′ region of PAX5 (a transcription factor essential for B cell development) to the 3′ region of TEL/ETV6, containing the Ets-family DNA binding domain. We have cloned the FLAG-full length chimeric PAX5/TEL cDNA in the retroviral vector pMSCV-IRES-GFP (MigR1) to transduce target cells. We have demonstrated a specific nuclear localization of the chimeric protein in NIH3T3 by immunofluorescence analysis. Moreover, we observed a PAX5/TEL dependent decrease of the cellular growth rate in IL-3 dependent murine proB Ba/F3 cells. We further investigated the function of the PAX5/TEL chimeric protein as a potential oncoprotein in murine preBI cells, as a more physiological model. Murine PAX5 −/− preBI cells and wild type preBI cells were purified as B220+/c-KIT+ cells from mouse fetal liver and they were cultured on OP9 and DL1-OP9 stroma cells in presence of IL-7. The OP9 stroma supports B cell proliferation and survival; the DL1-OP9 stroma expresses Delta-like1, one of the Notch ligands, and it’s important to support T cell development. Both PAX5 −/− preBI cells and wild type preBI cells were transduced with the retroviral construct pMSCV-PAX5/TEL-IRES-GFP to analyze cell proliferation, differentiation and growth-dependence on IL-7. Wild type preBI cells expressing PAX5/TEL showed down modulation of CD19 when cultured on OP9 stroma in presence of IL-7; an inverse correlation was observed between the levels of expression of GFP and of CD19. The down modulation of CD19 can be involved in driving the preBI cell into differentiation block. A possible explanation of CD19 repression can rely on a potential competition between PAX5/TEL and endogenous PAX5 to bind PAX5 consensus region on DNA. On OP9 stroma, PAX5/TEL preBI cells are resistant to TGFbeta anti-proliferative and apoptotic effects, with a three-fold increased growth rate than control cells. Although the specific mechanism of PAX5/TEL disruption of TGFbeta signalling pathway remains to be investigated, we propose the TGFbeta resistance by PAX5/TEL as a way to evade the immunosurveillance. PAX5/TEL-preBI cells cultured on DL1-OP9 showed a different phenotype, with up-regulation of c-KIT and down-regulation of CD44. PAX5−/− preBI cells infected with PAX5TEL and grown on OP9 were CD19 negative even in the presence of PAX5TEL. On DL1-OP9 stroma, PAX5TEL cells were able to differentiate maintaining the developmental plasticity of PAX5 −/− preBI cells. These preliminary results indicate a role of PAX5/TEL as a transcription factor, potentially with a suppressor function, down regulating CD19 expression, thus suggesting a function on B cell differentiation. The chimera is able to interfere with TGFbeta pathway, inducing resistance and conferring an advantage in cell survival, evading the immunosurveillance. PAX5TEL do not replace PAX5 functions in PAX5−/− cells, it cannot activate PAX5 target genes as CD19, important for restoring B cell differentiation. Further analyeis are needed to better evaluate the role of PAX5/TEL protein, both in vivo and in vitro models.

scholarly journals Dissecting the Role of the N-Terminal Region of the Escherichia coli Global Transcription Factor FNR

2008 ◽  
Vol 190 (24) ◽  
pp. 8230-8233 ◽  
Author(s):  
Aixin Yan ◽  
Patricia J. Kiley
Keyword(s):  
Escherichia Coli ◽  
Transcription Factor ◽  
Terminal Region ◽  
Wild Type ◽  
Fnr Protein

ABSTRACT The role of the N-terminal region of the transcription factor FNR, which immediately precedes the first ligand (Cys20) of the [4Fe-4S] cluster, was investigated. We found that truncation mutants that removed residues 2 to 16 and 2 to 17 had wild-type levels of FNR protein but surprisingly altered O2 regulation.

scholarly journals Leukocyte integrin Mac-1 regulates thrombosis via interaction with platelet GPIbα

2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Yunmei Wang ◽  
Huiyun Gao ◽  
Can Shi ◽  
Paul W. Erhardt ◽  
Alexander Pavlovsky ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Carotid Artery ◽  
Binding Site ◽  
Small Molecule ◽  
Therapeutic Potential ◽  
Bleeding Risk ◽  
Disease Models ◽  
Wild Type ◽  
Microvascular Injury

Abstract Inflammation and thrombosis occur together in many diseases. The leukocyte integrin Mac-1 (also known as integrin αMβ2, or CD11b/CD18) is crucial for leukocyte recruitment to the endothelium, and Mac-1 engagement of platelet GPIbα is required for injury responses in diverse disease models. However, the role of Mac-1 in thrombosis is undefined. Here we report that mice with Mac-1 deficiency (Mac-1 −/− ) or mutation of the Mac-1-binding site for GPIbα have delayed thrombosis after carotid artery and cremaster microvascular injury without affecting parameters of haemostasis. Adoptive wild-type leukocyte transfer rescues the thrombosis defect in Mac-1 −/− mice, and Mac-1-dependent regulation of the transcription factor Foxp1 contributes to thrombosis as evidenced by delayed thrombosis in mice with monocyte-/macrophage-specific overexpression of Foxp1. Antibody and small-molecule targeting of Mac-1:GPIbα inhibits thrombosis. Our data identify a new pathway of thrombosis involving leukocyte Mac-1 and platelet GPIbα, and suggest that targeting this interaction has anti-thrombotic therapeutic potential with reduced bleeding risk.

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