All three splice variants of the human sarco/endoplasmic reticulum Ca2+-ATPase 3 gene are translated to proteins: a study of their co-expression in platelets and lymphoid cells

2001 ◽  
Vol 358 (3) ◽  
pp. 559-568 ◽  
Author(s):  
Tünde KOVÁCS ◽  
Ferenc FELFÖLDI ◽  
Béla PAPP ◽  
Katalin PÁSZTY ◽  
Raymonde BREDOUX ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Splice Variants ◽  
Cell Types ◽  
Apparent Size ◽  
Lymphoid Cells ◽  
Jurkat Cells ◽  
Protein Isoforms ◽  
Published Data ◽  
Hek 293 ◽  
Tryptic Fragment

The molecular cloning of two previously unknown human sarco/endoplasmic reticulum Ca2+-ATPase 3 (SERCA3) 3′-end transcripts, 3b and 3c, has been recently published. Data were lacking, however, for the presence of these SERCA3 variants in different tissue or cell types at the protein level. Here we report the co-expression of three human SERCA3 protein isoforms in platelets and T lymphoid Jurkat cells. Isoform-specific polyclonal anti-peptide antibodies have been generated that recognize specifically the SERCA3a, 3b or 3c splice variants at their C-termini, and this has been confirmed by peptide-competition experiments as well. None of these antibodies cross-reacted with the housekeeping SERCA2b isoform co-expressed endogenously with SERCA3 proteins in non-muscle cells. Although all three SERCA3 isoforms could be detected in platelets, the 3a form was the most abundantly expressed species. Its size matched the apparent size of SERCA3a over-expressed in HEK-293 cells. Immunoprecipitation of the SERCA3 variants from platelet membranes using a PL/IM 430-affinity matrix provided evidence that the putative pan-anti-SERCA3 antibody, PL/IM 430, recognizes all SERCA3 protein isoforms. The epitope for the PL/IM 430 antibody could be localized in a 40kDa N-terminal tryptic fragment common to all three SERCA3 variants. Comparative Western-blot analysis showed that the expression level of the SERCA3a, 3b and 3c isoforms was more than 10 times lower in Jurkat cells than in platelets, whereas expression of the ubiquitous SERCA2b was nearly identical. This work highlights new Ca2+-transporting proteins of haematopoietic cells and provides specific antibodies for their detection.

scholarly journals All three splice variants of the human sarco/endoplasmic reticulum Ca2+-ATPase 3 gene are translated to proteins: a study of their co-expression in platelets and lymphoid cells

2001 ◽  
Vol 358 (3) ◽  
pp. 559 ◽  
Author(s):  
Tünde KOVÁCS ◽  
Ferenc FELFÖLDI ◽  
Béla PAPP ◽  
Katalin PÁSZTY ◽  
Raymonde BREDOUX ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Splice Variants ◽  
Lymphoid Cells

scholarly journals Divergent Mitochondrial and Endoplasmic Reticulum Association of DMPK Splice Isoforms Depends on Unique Sequence Arrangements in Tail Anchors

2005 ◽  
Vol 25 (4) ◽  
pp. 1402-1414 ◽  
Author(s):  
René E. M. A. van Herpen ◽  
Ralph J. A. Oude Ophuis ◽  
Mietske Wijers ◽  
Miranda B. Bennink ◽  
Fons A. J. van de Loo ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Kinase ◽  
Myotonic Dystrophy ◽  
Mammalian Cells ◽  
Cell Types ◽  
Unique Sequence ◽  
Protein Isoforms ◽  
Mitochondrial Outer Membrane ◽  
Splice Isoforms ◽  
Triplet Repeat Expansion

ABSTRACT Myotonic dystrophy protein kinase (DMPK) is a Ser/Thr-type protein kinase with unknown function, originally identified as the product of the gene that is mutated by triplet repeat expansion in patients with myotonic dystrophy type 1 (DM1). Alternative splicing of DMPK transcripts results in multiple protein isoforms carrying distinct C termini. Here, we demonstrate by expressing individual DMPKs in various cell types, including C2C12 and DMPK −/− myoblast cells, that unique sequence arrangements in these tails control the specificity of anchoring into intracellular membranes. Mouse DMPK A and C were found to associate specifically with either the endoplasmic reticulum (ER) or the mitochondrial outer membrane, whereas the corresponding human DMPK A and C proteins both localized to mitochondria. Expression of mouse and human DMPK A—but not C—isoforms in mammalian cells caused clustering of ER or mitochondria. Membrane association of DMPK isoforms was resistant to alkaline conditions, and mutagenesis analysis showed that proper anchoring was differentially dependent on basic residues flanking putative transmembrane domains, demonstrating that DMPK tails form unique tail anchors. This work identifies DMPK as the first kinase in the class of tail-anchored proteins, with a possible role in organelle distribution and dynamics.

Determination of apparent calcium affinity for endogenously expressed human sarco(endo)plasmic reticulum calcium-ATPase isoform SERCA3

2009 ◽  
Vol 296 (5) ◽  
pp. C1105-C1114 ◽  
Author(s):  
P. Charukeshi Chandrasekera ◽  
Margaret E. Kargacin ◽  
Julie P. Deans ◽  
Jonathan Lytton
Keyword(s):  
Cell Types ◽  
Protein Isoforms ◽  
Inhibitory Antibody ◽  
Human Tonsil ◽  
Hek 293 ◽  
293 Cells ◽  
Plasmic Reticulum ◽  
Two Component ◽  
Calcium Affinity ◽  
First Time

The sarco(endo)plasmic reticulum Ca2+-ATPases (SERCAs) play a crucial role in regulating free cytosolic Ca2+ concentration in diverse cell types. It has been shown that recombinant SERCA3, when measured in heterologous systems, exhibits low apparent affinity for Ca2+; however, Ca2+ affinity of native SERCA3 in an endogenous setting has not been examined. Such a measurement is complicated, because SERCA3 is always coexpressed with the housekeeping isoform SERCA2b. We used a fluorescence-based assay for monitoring continuous Ca2+ uptake into microsomes to examine the properties of endogenous human SERCA3 and SERCA2b. The kinetic parameters were derived using a cooperative two-component uptake model for Ca2+ activation, and the values assigned to SERCA3 were confirmed using the highly specific human SERCA3 inhibitory antibody PL/IM430. First, using recombinant human SERCA3 and SERCA2b proteins transiently expressed in HEK-293 cells, we confirmed the previously observed low apparent Ca2+ affinity for SERCA3 compared with SERCA2b (1.10 ± 0.04 vs. 0.26 ± 0.01 μM), and using mixtures of recombinant protein isoforms, we validated the two-component uptake model. Then we determined apparent Ca2+ affinity for SERCA proteins present endogenously in cultured Jurkat T lymphocytes and freshly isolated human tonsil lymphocytes. The apparent Ca2+ affinity in these two preparations was 1.04 ± 0.07 and 1.1 ± 0.2 μM for SERCA3 and 0.27 ± 0.02 and 0.26 ± 0.01 μM for SERCA2b, respectively. Our data demonstrate, for the first time, that affinity for Ca2+ is inherently lower for SERCA3 expressed in situ than for other SERCA isoforms.

Prevalence of the pit and antipit in the genus Glycine

1987 ◽  
Vol 45 ◽  
pp. 986-987
Author(s):  
R. W. Yaklich ◽  
E. L. Vigil ◽  
W. P. Wergin
Keyword(s):  
Amino Acids ◽  
Endoplasmic Reticulum ◽  
Glycine Max ◽  
Seed Coat ◽  
Cell Types ◽  
Abaxial Surface ◽  
Legume Seed ◽  
Initial Report ◽  
Cone Cells ◽  
Glycine Max L

The legume seed coat is the site of sucrose unloading and the metabolism of imported ureides and synthesis of amino acids for the developing embryo. The cell types directly responsible for these functions in the seed coat are not known. We recently described a convex layer of tissue on the inside surface of the soybean (Glycine max L. Merr.) seed coat that was termed “antipit” because it was in direct opposition to the concave pit on the abaxial surface of the cotyledon. Cone cells of the antipit contained numerous hypertrophied Golgi apparatus and laminated rough endoplasmic reticulum common to actively secreting cells. The initial report by Dzikowski (1936) described the morphology of the pit and antipit in G. max and found these structures in only 68 of the 169 seed accessions examined.

scholarly journals Selective deletion of SHIP-1 in hematopoietic cells in mice leads to severe lung inflammation involving ILC2 cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Xujun Ye ◽  
Fengrui Zhang ◽  
Li Zhou ◽  
Yadong Wei ◽  
Li Zhang ◽  
...  
Keyword(s):  
Lung Inflammation ◽  
Airway Remodeling ◽  
Pulmonary Inflammation ◽  
Hematopoietic Cells ◽  
Cell Lineage ◽  
Allergic Inflammation ◽  
Cell Types ◽  
Lymphoid Cells ◽  
Whole Body

AbstractSrc homology 2 domain–containing inositol 5-phosphatase 1 (SHIP-1) regulates the intracellular levels of phosphotidylinositol-3, 4, 5-trisphosphate, a phosphoinositide 3–kinase (PI3K) product. Emerging evidence suggests that the PI3K pathway is involved in allergic inflammation in the lung. Germline or induced whole-body deletion of SHIP-1 in mice led to spontaneous type 2-dominated pulmonary inflammation, demonstrating that SHIP-1 is essential for lung homeostasis. However, the mechanisms by which SHIP-1 regulates lung inflammation and the responsible cell types are still unclear. Deletion of SHIP-1 selectively in B cells, T cells, dendritic cells (DC) or macrophages did not lead to spontaneous allergic inflammation in mice, suggesting that innate immune cells, particularly group 2 innate lymphoid cells (ILC2 cells) may play an important role in this process. We tested this idea using mice with deletion of SHIP-1 in the hematopoietic cell lineage and examined the changes in ILC2 cells. Conditional deletion of SHIP-1 in hematopoietic cells in Tek-Cre/SHIP-1 mice resulted in spontaneous pulmonary inflammation with features of type 2 immune responses and airway remodeling like those seen in mice with global deletion of SHIP-1. Furthermore, when compared to wild-type control mice, Tek-Cre/SHIP-1 mice displayed a significant increase in the number of IL-5/IL-13 producing ILC2 cells in the lung at baseline and after stimulation by allergen Papain. These findings provide some hints that PI3K signaling may play a role in ILC2 cell development at baseline and in response to allergen stimulation. SHIP-1 is required for maintaining lung homeostasis potentially by restraining ILC2 cells and type 2 inflammation.

EFFECT OF FK506 ON ENDOPLASMIC RETICULUM - MEDIATED APOPTOSIS OF JURKAT CELLS

2010 ◽  
Vol 90 ◽  
pp. 649
Author(s):  
S. Y. Chung ◽  
H. K. Lee ◽  
W. Y. Shin ◽  
S. J.N. Choi
Keyword(s):  
Endoplasmic Reticulum ◽  
Jurkat Cells

scholarly journals Effects of taurolithocholate, a Ca2+-mobilizing agent, on cell Ca2+ in rat hepatocytes, human platelets and neuroblastoma NG108-15 cell line

1991 ◽  
Vol 273 (1) ◽  
pp. 153-160 ◽  
Author(s):  
J F Coquil ◽  
B Berthon ◽  
N Chomiki ◽  
L Combettes ◽  
P Jourdon ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Bile Acid ◽  
Cell Line ◽  
Rat Liver ◽  
Neuronal Cell ◽  
Rat Hepatocytes ◽  
Cell Types ◽  
Liver Cells ◽  
Human Platelets ◽  
Rat Liver Cells

The monohydroxy bile acid taurolithocholate permeabilizes the endoplasmic reticulum to Ca2+ in rat liver cells. To assess whether this action on the endoplasmic reticulum was restricted to this tissue, the effects of bile acid were investigated in two cell types quite unrelated to rat hepatocyte, namely human platelets and neuronal NG108-15 cell line. The results showed that taurolithocholate (3-100 microM) had no effect on free cytosolic [Ca2+] in human platelets and NG108-15 cells. whereas it increased it from 180 to 520 nM in rat hepatocytes. In contrast, in cells permeabilized by saponin, taurolithocholate initiated a profound release of the stored Ca2+ from the internal Ca2+ pools in the three cell types. The bile acid released 90% of the Ca2+ pools, with rate constants of about 5 min-1 and half-maximal effects at 15-30 microM. The results also showed that, in contrast with liver cells, which displayed an influx of [14C]taurolithocholate of 2 nmol/min per mg, human platelets and the neuronal cell line appeared to be resistant to [14C]taurolithocholate uptake. The influx measured in these latter cells was about 100-fold lower than in rat liver cells. Taken together, these data suggest that human platelets and NG108-15 cells do not possess the transport system for concentrating monohydroxy bile acids into cells. However, they show that human platelets and neuronal NG108-15 possess, in common with liver cells, the intracellular system responsible for taurolithocholate-mediated Ca2+ release from internal stores.

Diversity, localization and (patho)physiology of mature lymphocyte populations in the bone marrow

Blood ◽  
2021 ◽  
Author(s):  
Christian M. Schürch ◽  
Chiara Caraccio ◽  
Martijn A. Nolte
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Plasma Cells ◽  
Cell Types ◽  
Lymphoid Organ ◽  
Lymphoid Cells ◽  
Term Survival ◽  
Hematopoietic Stem ◽  
Mature Lymphocyte ◽  
Lymphocyte Populations

The bone marrow (BM) is responsible for generating and maintaining lifelong output of blood and immune cells. Besides its key hematopoietic function, the BM acts as an important lymphoid organ, hosting a large variety of mature lymphocyte populations, including B-cells, T-cells, NK(T)-cells and innate lymphoid cells (ILCs). Many of these cell types are thought to only transiently visit the BM, but for others, like plasma cells and memory T-cells, the BM provides supportive niches that promote their long-term survival. Interestingly, accumulating evidence points towards an important role for mature lymphocytes in the regulation of hematopoietic stem cells (HSCs) and hematopoiesis in health and disease. In this review, we describe the diversity, migration, localization and function of mature lymphocyte populations in murine and human BM, focusing on their role in immunity and hematopoiesis. We also address how various BM lymphocyte subsets contribute to the development of aplastic anemia and immune thrombocytopenia, illustrating the complexity of these BM disorders, but also the underlying similarities and differences in their disease pathophysiology. Finally, we summarize the interactions between mature lymphocytes and BM resident cells in HSC transplantation and graft-versus-host disease. A better understanding of the mechanisms by which mature lymphocyte populations regulate BM function will likely improve future therapies for patients with benign and malignant hematological disorders.

scholarly journals Phenotype of recovering lymphoid cell populations after marrow transplantation.

1985 ◽  
Vol 161 (6) ◽  
pp. 1483-1502 ◽  
Author(s):  
K A Ault ◽  
J H Antin ◽  
D Ginsburg ◽  
S H Orkin ◽  
J M Rappeport ◽  
...  
Keyword(s):  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Nk Cells ◽  
Cell Types ◽  
Lymphoid Cells ◽  
The Other ◽  
Hla Dr ◽  
T Cell Antigens ◽  
Leu 7

Four patients who received bone marrow transplants were studied sequentially during the posttransplant period to define the pattern of recovering lymphoid cell types. Three patients received T cell-depleted, HLA-matched marrow, and one received untreated marrow from an identical twin. Blood lymphoid cells were labeled with 25 different pairs of monoclonal antibodies. In each sample, one antibody was conjugated to fluorescein and one to phycoerythrin, thus allowing simultaneous assessment of the expression of the two markers using the fluorescence activated cell sorter. A total of 14 antibodies were used, routinely including HLE, Leu-M3, Leu-4, Leu-1, Leu-5, Leu-9, Leu-6, Leu-2, Leu-3, HLA-DR, Leu-7, Leu-11, Leu-15, and Leu-12. Other antibodies were used to further define some populations. This study has allowed us to define six distinct cell types that have appeared in all four patients by day 90 posttransplantation, and which account for 90-100% of all circulating lymphoid cells. These cell types are (a) T helper cells expressing Leu-1, Leu-4, Leu-9, Leu-5, Leu-3, and variable amounts of HLA-DR; (b) T suppressor cells expressing Leu-1, Leu-4, Leu-9, Leu-5, Leu-2, and variable amounts of HLA-DR; (c) B cells expressing Leu-12, B1, HLA-DR, IgD, and IgM, but none of the T cell antigens; (d) an unusual B cell phenotype (Leu-1 B) expressing all of the B cell markers, and also having low amounts of Leu-1, but none of the other T cell antigens; (e) natural killer (NK) cells expressing Leu-11, Leu-15, Leu-5 but none of the other T cell or B cell markers; (f) NK cells expressing Leu-11, Leu-15, Leu-5, and low levels of Leu-2. Both NK types also express Leu-7 on some, but not all cells. The relative frequencies of these cell types varied among the patients and with time, but the striking findings were the presence of relatively few mature T cells, large numbers of NK cells, and the preponderance of the unusual Leu-1 B cell over conventional B cells in all three patients who developed B cells. Sorting experiments confirmed the NK activity of the major NK cell phenotypes, and DNA analysis confirmed that all of the cells studied were of donor origin. In addition, analysis of Ig genes in one patient showed that the Leu-1 B cells were not clonally rearranged.(ABSTRACT TRUNCATED AT 400 WORDS)

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