A dual-purpose real-time indicator and transcriptional integrator for calcium detection in living cells

2021 ◽  
Author(s):  
Elbegduuren Erdenee ◽  
Alice Y Ting
Keyword(s):  
Gene Expression ◽  
Real Time ◽  
T Cell Activation ◽  
Protein Interactions ◽  
Cell Activation ◽  
Modular Design ◽  
Stable Gene ◽  
Dual Purpose ◽  
Calcium Detection ◽  
Stable Gene Expression

Calcium is a ubiquitous second messenger in eukaryotes, correlated with neuronal activity and T-cell activation among other processes. Real-time calcium indicators such as GCaMP have recently been complemented by newer calcium integrators that convert transient calcium activity into stable gene expression. Here we introduce LuCID, a dual-purpose real-time calcium indicator and transcriptional calcium integrator which combines the benefits of both calcium detection technologies. We show that the calcium-dependent split luciferase component of LuCID provides real-time bioluminescence readout of calcium dynamics in cells, while the GI/FKF1 split GAL4 component of LuCID converts bioluminescence into stable gene expression. We also show that LuCID modular design enables it to also be used for dual-purpose detection of other cellular events such as protein-protein interactions. LuCID should facilitate the study of cell populations and circuits that utilize calcium for signaling.

Abstract 647: Identification of Platelet Derived Growth Factors A, B, C and D Specific Role in Vascular Remodeling

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Maria Gonzalez Diez ◽  
Anton Razuvaev ◽  
Ulf Hedin ◽  
Anders Hamsten
Keyword(s):  
Gene Expression ◽  
Cell Differentiation ◽  
Blood Vessel ◽  
Platelet Activation ◽  
T Cell Activation ◽  
Vascular Remodeling ◽  
Cell Activation ◽  
Opposite Effect ◽  
Specific Role ◽  
Biological Processes

Restenosis is a major complication after coronary angioplasty and stenting. The major cause of restenosis is neointimal hyperplasia, which results from an excessive proliferative response of vascular smooth muscle cells (VSMC) to mechanical injury. Platelet derived growth factor (PDGF) family members (A, B, C, D) are known to be related to vascular remodeling. However whether this role is specific for each one or overlapping remains to be elucidated. Aim: To assess the specific role of PDGF family members (A, B, C, D) in vascular remodeling after injury. Methods: We used an established model of balloon injury in rat carotid artery. The endothelium of the intima is mechanically removed. The animals (n=10/group) were sacrificed at different time points after injury (0-2-20 hours, 2-5-15 days, 6-12 weeks). mRNA from carotid arteries were isolated for gene expression studies using microarray gene expression. Results: PDGFs are differentially expressed in vascular remodeling (mRNA, A adj P val=3.28E-06, B adj P val=4.52E-8, C adj P val=5,91E-15, D adj P val=2,64E-18). Also the expression profile differs among them. We selected the genes highly correlated with each of the PDGFs (Spearman correlation, │rs >0.7│) and identified the most preeminent biological pathways associated to each one. PDGF-A positively correlates with program cell death. On the other hand, PDGF-B and C have some overlapping biological processes. There is positive correlation with blood vessel morphogenesis and angiogenesis (B), cell differentiation (B and C), DNA replication (B and C), antigen presentation and T-cell activation/differentiation (B and C). However, there is negative correlation with platelet activation (B) and cell adhesion (B and C). PDGF-D positively correlates with blood vessel morphogenesis and angiogenesis (like B) and cell differentiation (B, C), but is negatively correlated with T-cell activation/proliferation (opposite effect to B and C), apoptosis (opposite effect to A) and platelet activation (B). Conclusion: We identified specific biological processes for PDGF- A, B, C and D. Despite some overlapping, each one plays a specific role within vascular remodeling.

Electric field-mediated DNA transfer: transient and stable gene expression in human and mouse lymphoid cells

1986 ◽  
Vol 6 (2) ◽  
pp. 703-706
Author(s):  
F Toneguzzo ◽  
A C Hayday ◽  
A Keating
Keyword(s):  
Gene Expression ◽  
Cell Lines ◽  
Lymphoid Cell ◽  
Simian Virus 40 ◽  
Lymphoid Cells ◽  
Dna Transfer ◽  
Stable Gene ◽  
Regulatory Sequences ◽  
Stable Gene Expression ◽  
Lymphoid Cell Lines

The technique of DNA transfer by electroporation was investigated in an effort to evaluate its utility for the identification of developmentally controlled regulatory sequences. Transient and stable gene expression was detected in a variety of lymphoid cell lines subjected to electroporation. No correlation existed between the levels of chloramphenicol acetyltransferase (acetyl-CoA; chloramphenicol 3-O-acetyltransferase, EC 2.3.1.28) expression and stable transfection frequency. In all lymphoid cell lines tested, the simian virus 40 early region was a better promoter than was the Rous sarcoma virus long terminal repeat.

scholarly journals Downregulation of Prdm16 mRNA is a specific antileukemic mechanism during HOXB4-mediated HSC expansion in vivo

Blood ◽  
2014 ◽  
Vol 124 (11) ◽  
pp. 1737-1747 ◽  
Author(s):  
Hui Yu ◽  
Geoffrey Neale ◽  
Hui Zhang ◽  
Han M. Lee ◽  
Zhijun Ma ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stable Gene ◽  
Self Renewal ◽  
Key Points ◽  
Stable Gene Expression

Key Points HOXB4 induces stable gene expression changes in transplanted HSCs that drive balanced self-renewal and differentiation divisions. Marked downregulation of Prdm16 occurs concurrently with HOXB4-mediated HSC expansion and functions to prevent leukemia in vivo.

T.69. T Cell Activation and Th2-related Gene Expression Correlate with Disease Activity in Glatiramer Acetate-treated Multiple Sclerosis

2009 ◽  
Vol 131 ◽  
pp. S70
Author(s):  
Finn Sellebjerg ◽  
Martin Krakauer ◽  
Dan Hesse ◽  
Henrik Lund ◽  
Signe Limborg ◽  
...  
Keyword(s):  
Gene Expression ◽  
Multiple Sclerosis ◽  
T Cell ◽  
Disease Activity ◽  
Glatiramer Acetate ◽  
T Cell Activation ◽  
Cell Activation ◽  
Related Gene

scholarly journals Nanoconfinement of Microvilli Alters Gene Expression and Boosts T cell Activation

2021 ◽  
Author(s):  
Morteza Aramesh ◽  
Diana Stoycheva ◽  
Ioana Sandu ◽  
Stephan J. Ihle ◽  
Tamara Zund ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Cell Signaling ◽  
T Cell Activation ◽  
Cell Activation ◽  
Local Environment ◽  
T Cell Signaling ◽  
Sense And Respond ◽  
Altered Gene

T cells sense and respond to their local environment at the nanoscale by forming small actin-rich protrusions, called microvilli, which play critical roles in signaling and antigen recognition, particularly at the interface with the antigen presenting cells. However, the mechanisms by which microvilli contribute to cell signaling and activation is largely unknown. Here, we present a tunable engineered system that promotes microvilli formation and T cell signaling via physical stimuli. We discovered that nanoporous surfaces favored microvilli formation, and markedly altered gene expression in T cells and promoted their activation. Mechanistically, confinement of microvilli inside of nanopores leads to size-dependent sorting of membrane-anchored proteins, specifically segregating CD45 phosphatases and T cell receptors (TCR) from the tip of the protrusions when microvilli are confined in 200 nm pores, but not in 400 nm pores. Consequently, formation of TCR nanoclustered hotspots within 200 nm pores, allows sustained and augmented signaling that prompts T cell activation even in the absence of TCR agonists. The synergistic combination of mechanical and biochemical signals on porous surfaces presents a straightforward strategy to investigate the role of microvilli in T cell signaling as well as to boost T cell activation and expansion for application in the growing field of adoptive immunotherapy.

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