scholarly journals Temperature-Sensitive Transformation by an Abelson Virus Mutant Encoding an Altered SH2 Domain

2001 ◽  
Vol 75 (4) ◽  
pp. 1816-1823 ◽  
Author(s):  
Celine A. Mainville ◽  
Kalindi Parmar ◽  
Indira Unnikrishnan ◽  
Li Gong ◽  
Glen D. Raffel ◽  
...  
Keyword(s):  
B Cells ◽  
Leukemia Virus ◽  
Binding Pocket ◽  
Sh2 Domain ◽  
Wild Type Virus ◽  
Temperature Sensitive ◽  
Temperature Dependent ◽  
Abelson Virus ◽  
Nih 3T3 ◽  
The Impact

ABSTRACT Abelson murine leukemia virus (Ab-MLV) encodes the v-Abl protein tyrosine kinase and induces transformation of immortalized fibroblast lines and pre-B cells. Temperature-sensitive mutations affecting the kinase domain of the protein have demonstrated that the kinase activity is absolutely required for transformation. Despite this requirement, mutations affecting other regions of v-Abl modulate transformation activity. The SH2 domain and the highly conserved FLVRES motif within it form a phosphotyrosine-binding pocket that is required for interactions between the kinase and cellular substrates. To understand the impact of SH2 alterations on Ab-MLV-mediated transformation, we studied the Ab-MLV mutant P120/R273K. This mutant encodes a v-Abl protein in which the βB5 arginine at the base of the phosphotyrosine-binding pocket has been replaced by a lysine. Unexpectedly, infection of NIH 3T3 or pre-B cells with P120/R273K revealed a temperature-dependent transformation phenotype. At 34°C, P120/R273K transformed about 10-fold fewer cells than wild-type virus of equivalent titer; at 39.5°C, 300-fold fewer NIH 3T3 cells were transformed and pre-B cells were refractory to transformation. Temperature-dependent transformation was accompanied by decreased phosphorylation of Shc, a protein that interacts with the v-Abl SH2 and links the protein to Ras, and decreased induction of c-Myc expression. These data suggest that alteration of the FLVRES pocket affects the ability of v-Abl to interact with at least some of its substrates in a temperature-dependent fashion and identify a novel type of temperature-sensitive Abelson virus.

scholarly journals Absence of p53 Complements Defects in Abelson Murine Leukemia Virus Signaling

2003 ◽  
Vol 77 (11) ◽  
pp. 6208-6215 ◽  
Author(s):  
Indira Unnikrishnan ◽  
Naomi Rosenberg
Keyword(s):  
B Cells ◽  
Murine Leukemia Virus ◽  
Cellular Response ◽  
Leukemia Virus ◽  
Binding Pocket ◽  
Transformation Process ◽  
Sh2 Domain ◽  
Wild Type ◽  
Abelson Murine Leukemia Virus ◽  
Murine Leukemia

ABSTRACT The v-Abl protein encoded by Abelson murine leukemia virus (Ab-MLV) induces transformation of pre-B cells via a two-stage process. An initial proliferative phase during which cells with limited tumorigenic potential expand is followed by a crisis period marked by high levels of apoptosis and erratic growth. Transformants that survive this phase emerge as fully malignant cells and usually contain mutations that disable the p53 tumor suppressor pathway. Consistent with the importance of p53 in this process, pre-B cells from p53 null animals bypass crisis. Thus, the transformation process reflects a balance between signals from the v-Abl protein that drive transformation and those coming from the cellular response to inappropriate growth. One prediction of this hypothesis is that Ab-MLV mutants that are compromised in their ability to transform cells may be less equipped to overcome the effects of p53. To test this idea, we examined the ability of the P120/R273K mutant to transform pre-B cells from wild-type, p53 null, and Ink4a/Arf null mice. The SH2 domain of the v-Abl protein encoded by this mutant contains a substitution that affects the phosphotyrosine-binding pocket, and this mutant is compromised in its ability to transform NIH 3T3 and pre-B cells, especially at 39.5°C. Our data reveal that loss of p53 or Ink4a/Arf locus products complements the transforming defect of the P120/R273K mutant, but it does not completely restore wild-type function. These results indicate that one important transforming function of v-Abl proteins is overcoming the effects of a functional p53 pathway.

scholarly journals Active Akt and Functional p53 Modulate Apoptosis in Abelson Virus-Transformed Pre-B Cells

2004 ◽  
Vol 78 (4) ◽  
pp. 1636-1644 ◽  
Author(s):  
Li Gong ◽  
Indira Unnikrishnan ◽  
Anuradha Raghavan ◽  
Kalindi Parmar ◽  
Naomi Rosenberg
Keyword(s):  
B Cells ◽  
Protein Tyrosine Kinase ◽  
Leukemia Virus ◽  
Temperature Shift ◽  
Kinase Domain ◽  
Transformation Process ◽  
Sh2 Domain ◽  
Temperature Sensitive ◽  
Nonpermissive Temperature ◽  
Src Homology

ABSTRACT Suppression of apoptosis is an important feature of the Abelson murine leukemia virus (Ab-MLV) transformation process. During multistep transformation, Ab-MLV-infected pre-B cells undergo p53-dependent apoptosis during the crisis phase of transformation. Even once cells are fully transformed, an active v-Abl protein tyrosine kinase is required to suppress apoptosis because cells transformed by temperature-sensitive (ts) kinase mutants undergo rapid apoptosis after a shift to the nonpermissive temperature. However, inactivation of the v-Abl protein by a temperature shift interrupts signals transmitted via multiple pathways, making it difficult to identify those that are critically important for the suppression of apoptosis. To begin to dissect these pathways, we tested the ability of an SH2 domain Ab-MLV mutant, P120/R273K, to rescue aspects of the ts phenotype of pre-B cells transformed by the conditional kinase domain mutant. The P120/R273K mutant suppressed apoptosis at the nonpermissive temperature, a phenotype correlated with its ability to activate Akt. Apoptosis also was suppressed at the nonpermissive temperature by constitutively active Akt and in p53-null pre-B cells transformed with the ts kinase domain mutant. These data indicate that an intact Src homology 2 (SH2) domain is not critical for apoptosis suppression and suggest that signals transmitted through Akt and p53 play an important role in the response.

scholarly journals The Carboxyl Terminus of v-Abl Protein Can Augment SH2 Domain Function

2000 ◽  
Vol 74 (10) ◽  
pp. 4495-4504 ◽  
Author(s):  
David Warren ◽  
Andrew J. Heilpern ◽  
Kent Berg ◽  
Naomi Rosenberg
Keyword(s):  
Map Kinase ◽  
Protein Interactions ◽  
Leukemia Virus ◽  
Sh2 Domain ◽  
Carboxyl Terminus ◽  
Abl Tyrosine Kinase ◽  
Mitogen Activated Protein ◽  
Map Kinase Pathway ◽  
Nih 3T3 ◽  
Kinase Pathway

ABSTRACT Abelson murine leukemia virus (Ab-MLV) transforms NIH 3T3 and pre-B cells via expression of the v-Abl tyrosine kinase. Although the enzymatic activity of this molecule is absolutely required for transformation, other regions of the protein are also important for this response. Among these are the SH2 domain, involved in phosphotyrosine-dependent protein-protein interactions, and the long carboxyl terminus, which plays an important role in transformation of hematopoietic cells. Important signals are sent from each of these regions, and transformation is most likely orchestrated by the concerted action of these different parts of the protein. To explore this idea, we compared the ability of the v-Src SH2 domain to substitute for that of v-Abl in the full-length P120 v-Abl protein and in P70 v-Abl, a protein that lacks the carboxyl terminus characteristic of Abl family members. Ab-MLV strains expressing P70/S2 failed to transform NIH 3T3 cells and demonstrated a greatly reduced capacity to mediate signaling events associated with the Ras-dependent mitogen-activated protein (MAP) kinase pathway. In contrast, Ab-MLV strains expressing P120/S2 were indistinguishable from P120 with respect to these features. Analyses of additional mutants demonstrated that the last 162 amino acids of the carboxyl terminus were sufficient to restore transformation. These data demonstrate that an SH2 domain with v-Abl substrate specificity is required for NIH 3T3 transformation in the absence of the carboxyl terminus and suggest that cooperativity between the extreme carboxyl terminus and the SH2 domain facilitates the transmission of transforming signals via the MAP kinase pathway.

Type-C oncornavirus assembly in NIH/3T3 cells infected with Temperature-sensitive mutants of rauscher murine leukemia virus

1978 ◽  
Vol 36 (2) ◽  
pp. 356-357
Author(s):  
H. Yeger ◽  
V. I. Kalnins ◽  
J. R. Stephenson
Keyword(s):  
Murine Leukemia Virus ◽  
Leukemia Virus ◽  
Class Ii ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
3T3 Cells ◽  
Type C ◽  
Nih 3T3 ◽  
Murine Leukemia ◽  
Nih 3T3 Cells

Selected conditional-lethal mutants of mammalian leukemia virus have been used in an ultrastructural study to investigate type-C virus assembly at the cell membrane. Several temperature-sensitive (ts) mutants of Rauscher murine leukemia virus (R-MuLV) have been previously isolated and characterized by biochemical methods and electron microscopy. Biochemical analysis of one particular class of these mutants, the Class II ts mutants, which are defective in late replication functions, indicates that they are defective in cleavage of the gag-gene coded polypeptide precursor, Pr65, of the viral structural proteins at the non-permissive temperature. Because of this defect, NIH/3T3 cells infected with the Class II mutants, ts24, ts25, and ts29 produce very low levels of virus at the non-permissive temperature as compared to wt infected cells.

scholarly journals The Extreme Carboxyl Terminus of v-Abl Is Required for Lymphoid Cell Transformation by Abelson Virus

2003 ◽  
Vol 77 (8) ◽  
pp. 4617-4625 ◽  
Author(s):  
David Warren ◽  
Deborah S. Griffin ◽  
Celine Mainville ◽  
Naomi Rosenberg
Keyword(s):  
B Cells ◽  
Protein Tyrosine Kinase ◽  
Cell Transformation ◽  
Leukemia Virus ◽  
Fibroblast Cell ◽  
Lymphoid Cells ◽  
Carboxyl Terminus ◽  
Nih 3T3

ABSTRACT The v-Abl protein tyrosine kinase encoded by Abelson murine leukemia virus (Ab-MLV) induces transformation of pre-B cells in vivo and in vitro and can transform immortalized fibroblast cell lines in vitro. Although the kinase activity of the protein is required for these events, most previously studied mutants encoding truncated v-Abl proteins that lack the extreme carboxyl terminus retain high transforming capacity in NIH 3T3 cells but transform lymphocytes poorly. To understand the mechanisms responsible for poor lymphoid transformation, mutants expressing a v-Abl protein lacking portions of the COOH terminus were compared for their ability to transform pre-B cells. Although all mutants lacking sequences within the COOH terminus were compromised for lymphoid transformation, loss of amino acids in the central region of the COOH terminus, including those implicated in JAK interaction and DNA binding, decreased transformation twofold or less. In contrast, loss of the extreme COOH terminus rendered the protein unstable and led to rapid proteosome-mediated degradation, a feature that was more prominent when the protein was expressed in Ab-MLV-transformed lymphoid cells. These data indicate that the central portion of the COOH terminus is not essential for lymphoid transformation and reveal that one important function of the COOH terminus is to stabilize the v-Abl protein in lymphoid cells.

scholarly journals Disruption of the Shc/Grb2 Complex during Abelson Virus Transformation Affects Proliferation, but Not Apoptosis

2005 ◽  
Vol 79 (4) ◽  
pp. 2325-2334 ◽  
Author(s):  
Linda B. Baughn ◽  
Naomi Rosenberg
Keyword(s):  
B Cell ◽  
Protein Tyrosine Kinase ◽  
Cell Transformation ◽  
Leukemia Virus ◽  
Sh2 Domain ◽  
Dominant Negative ◽  
Growth And Survival ◽  
Erk Phosphorylation ◽  
Ras Activation ◽  
Abelson Virus

ABSTRACT The v-Abl protein tyrosine kinase encoded by Abelson murine leukemia virus (Ab-MLV) induces pre-B-cell transformation. Signals emanating from the SH2 domain of the protein are required for transformation, and several proteins bind this region of v-Abl. One such protein is the adaptor molecule Shc, a protein that complexes with Grb2/Sos and facilitates Ras activation, an event associated with Ab-MLV transformation. To test the role this interaction plays in growth and survival of infected pre-B cells, dominant-negative (DN) Shc proteins were coexpressed with v-Abl and transformation was examined. Expression of DN Shc reduced Ab-MLV pre-B-cell transformation and decreased the ability of v-Abl to stimulate Ras activation and Erk phosphorylation in a Raf-dependent but Rac-independent fashion. Further analysis revealed that Shc is required for v-Abl-mediated Raf tyrosine 340 and 341 phosphorylation, an event associated with Erk phosphorylation. In contrast to effects on proliferation, survival of the cells and activation of Akt were not affected by expression of DN Shc. Together, these data reveal that v-Abl-Shc interactions are a critical part of the growth stimulatory signals delivered during transformation but that they do not affect antiapoptotic pathways. Furthermore, these data highlight a novel role for Shc in signaling from v-Abl to Raf.

B-cell and T-cell values in peripheral blood in Polish mixed-breed rabbits with addition of blood of meet breeds

2014 ◽  
Vol 17 (3) ◽  
pp. 421-426 ◽  
Author(s):  
B. Tokarz-Deptuła ◽  
P. Niedźwiedzka-Rystwej ◽  
B. Hukowska-Szematowicz ◽  
M. Adamiak ◽  
A. Trzeciak-Ryczek ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Laboratory Animals ◽  
Immunological Parameters ◽  
Four Seasons ◽  
Mixed Breed ◽  
The Impact

Abstract In Poland, rabbit is a highly valued animal, due to dietetic and flavour values of its meat, but above all, rabbits tend to be commonly used laboratory animals. The aim of the study was developing standards for counts of B-cells with CD19+ receptor, T-cells with CD5+ receptor, and their subpopulations, namely T-cells with CD4+, CD8+ and CD25+ receptor in the peripheral blood of mixed-breed Polish rabbits with addition of blood of meet breeds, including the assessment of the impact of four seasons of the year and animal sex on the values of the immunological parameters determined. The results showed that the counts of B- and T-cells and their subpopulations in peripheral blood remain within the following ranges: for CD19+ B-cells: 1.05 - 3.05%, for CD5+ T-cells: 34.00 - 43.07%, CD4+ T-cells: 23.52 - 33.23%, CD8+ T-cells: 12.55 - 17.30%, whereas for CD25+ T-cells: 0.72 - 2.81%. As it comes to the season of the year, it was observed that it principally affects the values of CD25+ T-cells, while in the case of rabbit sex, more changes were found in females.

scholarly journals Natural Infection of Dairy Cows with Bovine Leukemia Virus Affects Immunoglobulin Levels in Saliva and Serum but Not Milk

Pathogens ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 907
Author(s):  
Monika Dziuba ◽  
Vickie J. Ruggiero ◽  
Catherine Wilson ◽  
Paul C. Bartlett ◽  
Paul M. Coussens
Keyword(s):  
Pilot Study ◽  
Dairy Cows ◽  
Bovine Leukemia Virus ◽  
Leukemia Virus ◽  
Natural Infection ◽  
Total Serum ◽  
Serum Samples ◽  
Retroviral Infection ◽  
Serum Iga ◽  
The Impact

Bovine leukemia virus (BLV) is a retroviral infection that disrupts the immune function of infected animals. It is widespread among U.S. dairy cattle. In this pilot study, the average total IgA and IgM concentrations in milk, saliva, and serum samples from BLV ELISA-positive (ELISA+) dairy cows were compared against samples from BLV ELISA-negative (ELISA−) cows using the Kruskal–Wallis test (with ties). The results from ELISA+ cows were also stratified by lymphocyte count (LC) and proviral load (PVL). In milk and saliva from ELISA+ cows, the average total IgA and IgM concentrations were decreased compared to ELISA− cows, although this was only statistically significant for saliva IgM in cows with low PVL (p = 0.0424). Numerically, the average total IgA concentrations were 33.6% lower in milk and 23.7% lower in saliva, and the average total IgM concentrations were 42.4% lower in milk and 15.5% lower in saliva. No significant differences were observed in the total serum IgA concentrations, regardless of PVL and LC. The total serum IgM from ELISA+ cows was significantly decreased (p = 0.0223), with the largest decreases occurring in the highest PVL and LC subgroups. This pilot study is a first step in investigating the impact of BLV on mucosal immunity and will require further exploration in each of the various stages of disease progression.

scholarly journals Temperature Characterization of Liquid Crystal Dielectric Image Line Phase Shifter for Millimeter-Wave Applications

Crystals ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 63
Author(s):  
Henning Tesmer ◽  
Rani Razzouk ◽  
Ersin Polat ◽  
Dongwei Wang ◽  
Rolf Jakoby ◽  
...  
Keyword(s):  
Liquid Crystal ◽  
Millimeter Wave ◽  
Insertion Loss ◽  
Phase Shifter ◽  
Ground Plane ◽  
Temperature Dependent ◽  
Differential Phase ◽  
Dependent Behavior ◽  
The Impact ◽  
Temperature Dependent Behavior

In this paper we investigate the temperature dependent behavior of a liquid crystal (LC) loaded tunable dielectric image guide (DIG) phase shifter at millimeter-wave frequencies from 80 GHz to 110 GHz for future high data rate communications. The adhesive, necessary for precise fabrication, is analyzed before temperature dependent behavior of the component is shown, using the nematic LC-mixture GT7-29001. The temperature characterization is conducted by changing the temperature of the LC DIG’s ground plane between −10∘C and 80 ∘C. The orientation of the LC molecules, and therefore the effective macroscopic relative permittivity of the DIG, is changed by inserting the temperature setup in a fixture with rotatable magnets. Temperature independent matching can be observed, while the insertion loss gradually increases with temperature for both highest and lowest permittivity of the LC. At 80 ∘C the insertion loss is up to 1.3dB higher and at −10∘C it is 0.6dB lower than the insertion loss present at 20 ∘C. In addition, the achievable differential phase is reduced with increasing temperature. The impact of molecule alignment to this reduction is shown for the phase shifter and an estimated 85% of the anisotropy is still usable with an LC DIG phase shifter when increasing the temperature from 20 ∘C to 80 ∘C. Higher reduction of differential phase is present at higher frequencies as the electrical length of the phase shifter increases. A maximum difference in differential phase of 72∘ is present at 110 GHz, when increasing the temperature from 20 ∘C to 80 ∘C. Nevertheless, a well predictable, quasi-linear behavior can be observed at the covered temperature range, highlighting the potential of LC-based dielectric components at millimeter wave frequencies.

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