The document describes the ALTSORGHUM project which aims to develop molecular markers for aluminum tolerance in sorghum. It discusses the gene AltSB which encodes for organic acid release to chelate aluminum ions and reduce toxicity. The project seeks to identify alleles of this gene through association mapping and use markers in molecular breeding programs to develop aluminum tolerant sorghum varieties.
This document discusses the regulation of acetyl CoA carboxylase (ACC) by AMP-activated protein kinase (AMPK) and the role of liver kinase B1 (LKB1) in this process. It shows that LKB1 interacts with and is phosphorylated by the tyrosine kinase Fyn. Fyn promotes the nuclear export and activation of LKB1 leading to increased AMPK phosphorylation and inhibition of mTORC1 signaling. Mutation of specific LKB1 tyrosine residues involved in Fyn phosphorylation disrupts this process and LKB1 localization.
Determination of Oxygenates & BTX in Beverage Grade Carbon DioxideJennifer Maclachlan
In this paper, we describe a method for volatile oxygenates and demonstrate that it
can be combined with the previous BTX (1) method to perform multifunctional analyte
analyses. The photoionization detector (PID) has been improved to provide the sensitivity and
ppb detection limits required by the industry. The 301-B now has multiple valve switching
capability and multiple autozeros to smooth the baseline when columns are switched in or out.
With these changes and system optimizations, we have achieved some complex separations at
ppb levels with a minimum of operator time or training required.
The following method on the 301-B was developed for the analysis of ppb levels of oxygenates
(acetaldehyde, methanol & dimethyl ether) in beverage grade carbon dioxide. The detection
limits required were 100 ppb for acetaldehyde and dimethyl ether.
The analysis can be automated via a PLC that controls the 301-B (start, calibrate, print report).
An optional operating system is Windows XP. If the 301 GC is part of the plant network, the
system can be operated from any PC on the network. Of course the system has passwords.
The detection limit for this analysis was <100 ppb (acetaldehyde), & <50 ppb (dimethyl
ether) for a direct injection with no concentration of the sample. The analysis time for this
multicomponent analysis was < 10 minutes.
Table 1 describes various Bordetella strains and plasmids used in the study, including wild type strains and mutant derivatives lacking genes like bvgS, fla, and frlAB. Figures 1-5 show results of experiments examining the role of these genes in biofilm formation, flagella expression, and other phenotypes over time or under different conditions like shaking or static growth.
This document provides a summary of 10 artworks that provide context for the project "Waste in Place: The McRobies Gully Tip as Landscape". The artworks are divided into two sections: 1) "Framing Waste", which discusses works that have used waste as a device to critique social, economic and environmental perspectives; and 2) "Transformational Relations", which discusses works that have sought to provoke consideration of human relations with animals, objects or place. The works discussed use a variety of media including sculpture, photography, video and installation. They address issues such as waste and consumption, human impact on the environment, and complex relationships between humans, objects and sites. The document concludes that these artworks
11 Moments of Hitting the Sales Wall and How to Get Over ItMichael Lang
Warning: After viewing this presentation you may find yourself wanting to improve your sales skills.
11 different situations that lead to hitting the sales wall and how to climb over it to sales success. Learn how to avoid these sales situations and excel in your business.
If the chance to improve yourself is tempting, please visit www.sgpartners.com.au to find out more information on how we can train you and your sales team to a profitable tomorrow.
Lessons learned from early innovators in the field of intellectual capital. The field of IC is at risk of getting caught in the marketing chasm described by Geoffrey Moore in Crossing the Chasm. Presentation by Mary Adams and Peder Hofman-Bang at the European Conference on Intellectual Capital in April, 2009.
The document describes the ALTSORGHUM project which aims to develop molecular markers for aluminum tolerance in sorghum. It discusses the gene AltSB which encodes for organic acid release to chelate aluminum ions and reduce toxicity. The project seeks to identify alleles of this gene through association mapping and use markers in molecular breeding programs to develop aluminum tolerant sorghum varieties.
This document discusses the regulation of acetyl CoA carboxylase (ACC) by AMP-activated protein kinase (AMPK) and the role of liver kinase B1 (LKB1) in this process. It shows that LKB1 interacts with and is phosphorylated by the tyrosine kinase Fyn. Fyn promotes the nuclear export and activation of LKB1 leading to increased AMPK phosphorylation and inhibition of mTORC1 signaling. Mutation of specific LKB1 tyrosine residues involved in Fyn phosphorylation disrupts this process and LKB1 localization.
Determination of Oxygenates & BTX in Beverage Grade Carbon DioxideJennifer Maclachlan
In this paper, we describe a method for volatile oxygenates and demonstrate that it
can be combined with the previous BTX (1) method to perform multifunctional analyte
analyses. The photoionization detector (PID) has been improved to provide the sensitivity and
ppb detection limits required by the industry. The 301-B now has multiple valve switching
capability and multiple autozeros to smooth the baseline when columns are switched in or out.
With these changes and system optimizations, we have achieved some complex separations at
ppb levels with a minimum of operator time or training required.
The following method on the 301-B was developed for the analysis of ppb levels of oxygenates
(acetaldehyde, methanol & dimethyl ether) in beverage grade carbon dioxide. The detection
limits required were 100 ppb for acetaldehyde and dimethyl ether.
The analysis can be automated via a PLC that controls the 301-B (start, calibrate, print report).
An optional operating system is Windows XP. If the 301 GC is part of the plant network, the
system can be operated from any PC on the network. Of course the system has passwords.
The detection limit for this analysis was <100 ppb (acetaldehyde), & <50 ppb (dimethyl
ether) for a direct injection with no concentration of the sample. The analysis time for this
multicomponent analysis was < 10 minutes.
Table 1 describes various Bordetella strains and plasmids used in the study, including wild type strains and mutant derivatives lacking genes like bvgS, fla, and frlAB. Figures 1-5 show results of experiments examining the role of these genes in biofilm formation, flagella expression, and other phenotypes over time or under different conditions like shaking or static growth.
This document provides a summary of 10 artworks that provide context for the project "Waste in Place: The McRobies Gully Tip as Landscape". The artworks are divided into two sections: 1) "Framing Waste", which discusses works that have used waste as a device to critique social, economic and environmental perspectives; and 2) "Transformational Relations", which discusses works that have sought to provoke consideration of human relations with animals, objects or place. The works discussed use a variety of media including sculpture, photography, video and installation. They address issues such as waste and consumption, human impact on the environment, and complex relationships between humans, objects and sites. The document concludes that these artworks
11 Moments of Hitting the Sales Wall and How to Get Over ItMichael Lang
Warning: After viewing this presentation you may find yourself wanting to improve your sales skills.
11 different situations that lead to hitting the sales wall and how to climb over it to sales success. Learn how to avoid these sales situations and excel in your business.
If the chance to improve yourself is tempting, please visit www.sgpartners.com.au to find out more information on how we can train you and your sales team to a profitable tomorrow.
Lessons learned from early innovators in the field of intellectual capital. The field of IC is at risk of getting caught in the marketing chasm described by Geoffrey Moore in Crossing the Chasm. Presentation by Mary Adams and Peder Hofman-Bang at the European Conference on Intellectual Capital in April, 2009.
The shift to a knowledge-based economy has created significant challenges for accounting and corporate reporting. This presentation gives an overview of the challenges and opportunities facing accountants and businesspeople today.
The Role of the Intangibles Information Gap in the Financialization of the A...Smarter-Companies
This document discusses how corporate value is increasingly derived from intangible assets like intellectual property, brand recognition, and organizational capital rather than tangible assets. It notes that intangible investments by US corporations now exceed tangible investments annually but are not fully captured by traditional accounting systems. To address short-termism and more accurately value firms, the "intangibles information gap" must be closed by better identifying, measuring, and reporting intangible capital and knowledge assets.
The document advertises and provides details about Rockers' League, an annual cricket tournament organized by IBS Rocks for students and alumni of ICFAI Business School. The 8th edition of the tournament will take place on December 14th and 21st, 2008 at Chiranjeev Bharti School Ground in Gurgaon. It will feature a 16-team knockout format with 8-over innings. The tournament aims to promote teamwork, leadership, and sportsmanship among participants.
Here's one of our most recent projects that was completed in 2011. This client had just finished a major remodel of their house in 2008 and were about to enjoy Christmas in their new home. At the time, Seattle was buried under several inches of snow (a rarity for us) and the entire region was paralyzed for a few days waiting for the thaw. Our client decided to take advantage of this opportunity and was in his driveway sledding when his neighbor rushed down the drive yelling that his house was on fire. Unfortunately, the house was already engulfed in flames. Equally unfortunate was that snowstorm and the delay it caused the fire department getting to the site. By the time they arrived, the house and contents were a total loss of more than $2.2 million.
Our role in this reconstruction of this home was two-fold. The first year of our involvement was spent working a team of forensic contractors gutting the house, cleansing the house of all particulate matter, and helping our client negotiate his insurance settlement. Once we got over these hurdles, the design work and reconstruction started. Maintaining the exisiting shell, we reworked the interior room arrangement to create classic great room house with a contemporary twist. Both levels of the home we opened up to take advantage of the waterfront views and flood the interiors with natural light. On the lower level, rearrangment of the walls more than tripled the size of the family room while creating an additional sitting/game room. The upper level was arranged with living spaces bookended by the Master Bedroom at one end the kitchen at the other. The open Great Room and wrap around deck create a relaxed and sophisticated living and entertainment space that is accentuated by a high level of trim and tile detail on the interior and by custom metal railings and light fixtures on the exterior.
Most of the value in mergers and acquisitions is ascribed to intangible (not tangible) capital. Ignoring these intangibles can be dangerous. This presentation covers how to identify and measure intangibles in traditional Accounting deals--and presents a more modern alternative: ICounting.
Este documento es una ficha de autoevaluación de una propuesta presentada por Ana Marisol Gonzales Pérez en Mindmeister. La ficha evalúa diversos aspectos de la propuesta como los objetivos, la justificación, los beneficiarios, las actividades planteadas, la evaluación, los requerimientos y el cronograma. La autoevaluación determina que la propuesta cumple con todos los aspectos evaluados, obteniendo un puntaje total de 20 sobre 20. El único inconveniente mencionado es que hubo problemas de conectividad con la plataforma PERUEDUCA
We are a Field Marketing & Media Agency with our Regional Headquarters in the UAE and have a presence in 12 countries across the Middle East & Africa. We specialize in BTL Field Marketing Solutions with a focus on execution in the retail trade space, man-power outsourcing and Media Solutions to many multi-nationals and large scale local clients. In addition, we have a Media division in KSA with exclusive media rights in Panda, the leading supermarket chain in Saudi Arabia.
La Unión Europea ha propuesto un nuevo paquete de sanciones contra Rusia que incluye un embargo al petróleo. El embargo prohibiría las importaciones de petróleo ruso por mar y por oleoducto, aunque se concederían exenciones temporales a Hungría y Eslovaquia. El objetivo es aumentar la presión económica sobre Rusia para que ponga fin a su invasión de Ucrania.
This document discusses the regulation of acetyl CoA carboxylase (ACC) by AMP-activated protein kinase (AMPK) and the role of liver kinase B1 (LKB1) in this process. It shows that LKB1 interacts with and is phosphorylated by the tyrosine kinase Fyn. Fyn promotes LKB1 export from the nucleus into the cytoplasm. Phosphorylation of specific tyrosine residues on LKB1 by Fyn is required for its nuclear export and activation of AMPK signaling. This pathway regulates cell growth and metabolism through AMPK and mTOR signaling.
GPE/TiO2 paste electrode was modified with Calix[6]arene ionophore to determine Pb2+ ions. The optimum modified electrode composition was determined to be 0.015g Calix[6]arene, which showed an anodic peak current of 226μA and cathodic peak current of -207μA. The detection limit for Pb2+ was 0.04702ppm. Tests showed the modified electrode had good repeatability, stability for 30 days, and was selective for Pb2+ even in the presence of interfering Cd2+ and Mn2+ ions. The electrode was also able to determine Pb2+ levels in a real sample.
Presentation of Fridtof Lund-Johansen in 1st International Antibody Validatio...St John's Laboratory Ltd
Fridtjof Lund-Johansen is an MDPhD who has worked for 30 years with antibodies and flow cytometry. He took his degree at the University of Bergen in Norway. He then did post-docs in California, first at Becton Dickinson in San Jose and then at DNAX instiute of Immunology in Palo Alto. He is now a PI at the Institute of Immunology at Oslo University Hospital where he leads an effort to develop antibody-based proteomics. His technology is called MAP for microsphere affinity proteomics. He will talk about how MAP can be used to test thousands of antibodies in parallel.
For more details about 1st international antibody validation forum please check on http://www.stjohnslabs.com/ac_cms/blog
This document summarizes research on developing a method for targeted delivery of interleukin-12 (IL-12) to tumors to enhance its anti-tumor effects while reducing systemic toxicity. Key points:
1) Separate administration and vascular targeting of the individual IL-12 subunits p35 and p40 could allow for their reassembly and activity specifically in tumors, avoiding toxicity from cytokine release syndrome.
2) Mutations were introduced to the subunits to allow their purification via different tags after co-expression and cleavage with TEV protease.
3) Surface plasmon resonance analysis showed the mutated subunits can still assemble into IL-12, and in vitro bioassays demonstrated the assembled IL-12 induces IFN
The document discusses how AP-2α induces apoptosis in cancer cells. It finds that AP-2α activates the intrinsic apoptosis pathway by binding to the Bcl-2 promoter and repressing its transcription. This allows Bax to translocate to mitochondria, disrupt membrane potential, and release cytochrome c, activating caspase-9 and caspase-3. Downregulation of anti-apoptotic Bcl-2 is important for AP-2α-induced apoptosis. Overexpressing AP-2α enhances cancer cell chemosensitivity to various drugs.
The document discusses the use of pluripotent stem cell-derived cardiomyocytes as substrates for automated ion channel and electrophysiological analysis systems. Specifically, it describes how cardiomyocytes can be selected from mouse embryonic stem cells using puromycin, and how these cells express appropriate cardiac ion channels. It also demonstrates that the cardiomyocytes can be used on automated patch clamp systems like PatchXpress and QPatch to measure cardiac ion currents like INa, ICa, and IK, and to study the effects of drugs on these currents. The document establishes pluripotent stem cell-derived cardiomyocytes as a physiologically relevant and reproducible model for high-throughput electrophysiology screening.
The shift to a knowledge-based economy has created significant challenges for accounting and corporate reporting. This presentation gives an overview of the challenges and opportunities facing accountants and businesspeople today.
The Role of the Intangibles Information Gap in the Financialization of the A...Smarter-Companies
This document discusses how corporate value is increasingly derived from intangible assets like intellectual property, brand recognition, and organizational capital rather than tangible assets. It notes that intangible investments by US corporations now exceed tangible investments annually but are not fully captured by traditional accounting systems. To address short-termism and more accurately value firms, the "intangibles information gap" must be closed by better identifying, measuring, and reporting intangible capital and knowledge assets.
The document advertises and provides details about Rockers' League, an annual cricket tournament organized by IBS Rocks for students and alumni of ICFAI Business School. The 8th edition of the tournament will take place on December 14th and 21st, 2008 at Chiranjeev Bharti School Ground in Gurgaon. It will feature a 16-team knockout format with 8-over innings. The tournament aims to promote teamwork, leadership, and sportsmanship among participants.
Here's one of our most recent projects that was completed in 2011. This client had just finished a major remodel of their house in 2008 and were about to enjoy Christmas in their new home. At the time, Seattle was buried under several inches of snow (a rarity for us) and the entire region was paralyzed for a few days waiting for the thaw. Our client decided to take advantage of this opportunity and was in his driveway sledding when his neighbor rushed down the drive yelling that his house was on fire. Unfortunately, the house was already engulfed in flames. Equally unfortunate was that snowstorm and the delay it caused the fire department getting to the site. By the time they arrived, the house and contents were a total loss of more than $2.2 million.
Our role in this reconstruction of this home was two-fold. The first year of our involvement was spent working a team of forensic contractors gutting the house, cleansing the house of all particulate matter, and helping our client negotiate his insurance settlement. Once we got over these hurdles, the design work and reconstruction started. Maintaining the exisiting shell, we reworked the interior room arrangement to create classic great room house with a contemporary twist. Both levels of the home we opened up to take advantage of the waterfront views and flood the interiors with natural light. On the lower level, rearrangment of the walls more than tripled the size of the family room while creating an additional sitting/game room. The upper level was arranged with living spaces bookended by the Master Bedroom at one end the kitchen at the other. The open Great Room and wrap around deck create a relaxed and sophisticated living and entertainment space that is accentuated by a high level of trim and tile detail on the interior and by custom metal railings and light fixtures on the exterior.
Most of the value in mergers and acquisitions is ascribed to intangible (not tangible) capital. Ignoring these intangibles can be dangerous. This presentation covers how to identify and measure intangibles in traditional Accounting deals--and presents a more modern alternative: ICounting.
Este documento es una ficha de autoevaluación de una propuesta presentada por Ana Marisol Gonzales Pérez en Mindmeister. La ficha evalúa diversos aspectos de la propuesta como los objetivos, la justificación, los beneficiarios, las actividades planteadas, la evaluación, los requerimientos y el cronograma. La autoevaluación determina que la propuesta cumple con todos los aspectos evaluados, obteniendo un puntaje total de 20 sobre 20. El único inconveniente mencionado es que hubo problemas de conectividad con la plataforma PERUEDUCA
We are a Field Marketing & Media Agency with our Regional Headquarters in the UAE and have a presence in 12 countries across the Middle East & Africa. We specialize in BTL Field Marketing Solutions with a focus on execution in the retail trade space, man-power outsourcing and Media Solutions to many multi-nationals and large scale local clients. In addition, we have a Media division in KSA with exclusive media rights in Panda, the leading supermarket chain in Saudi Arabia.
La Unión Europea ha propuesto un nuevo paquete de sanciones contra Rusia que incluye un embargo al petróleo. El embargo prohibiría las importaciones de petróleo ruso por mar y por oleoducto, aunque se concederían exenciones temporales a Hungría y Eslovaquia. El objetivo es aumentar la presión económica sobre Rusia para que ponga fin a su invasión de Ucrania.
This document discusses the regulation of acetyl CoA carboxylase (ACC) by AMP-activated protein kinase (AMPK) and the role of liver kinase B1 (LKB1) in this process. It shows that LKB1 interacts with and is phosphorylated by the tyrosine kinase Fyn. Fyn promotes LKB1 export from the nucleus into the cytoplasm. Phosphorylation of specific tyrosine residues on LKB1 by Fyn is required for its nuclear export and activation of AMPK signaling. This pathway regulates cell growth and metabolism through AMPK and mTOR signaling.
GPE/TiO2 paste electrode was modified with Calix[6]arene ionophore to determine Pb2+ ions. The optimum modified electrode composition was determined to be 0.015g Calix[6]arene, which showed an anodic peak current of 226μA and cathodic peak current of -207μA. The detection limit for Pb2+ was 0.04702ppm. Tests showed the modified electrode had good repeatability, stability for 30 days, and was selective for Pb2+ even in the presence of interfering Cd2+ and Mn2+ ions. The electrode was also able to determine Pb2+ levels in a real sample.
Presentation of Fridtof Lund-Johansen in 1st International Antibody Validatio...St John's Laboratory Ltd
Fridtjof Lund-Johansen is an MDPhD who has worked for 30 years with antibodies and flow cytometry. He took his degree at the University of Bergen in Norway. He then did post-docs in California, first at Becton Dickinson in San Jose and then at DNAX instiute of Immunology in Palo Alto. He is now a PI at the Institute of Immunology at Oslo University Hospital where he leads an effort to develop antibody-based proteomics. His technology is called MAP for microsphere affinity proteomics. He will talk about how MAP can be used to test thousands of antibodies in parallel.
For more details about 1st international antibody validation forum please check on http://www.stjohnslabs.com/ac_cms/blog
This document summarizes research on developing a method for targeted delivery of interleukin-12 (IL-12) to tumors to enhance its anti-tumor effects while reducing systemic toxicity. Key points:
1) Separate administration and vascular targeting of the individual IL-12 subunits p35 and p40 could allow for their reassembly and activity specifically in tumors, avoiding toxicity from cytokine release syndrome.
2) Mutations were introduced to the subunits to allow their purification via different tags after co-expression and cleavage with TEV protease.
3) Surface plasmon resonance analysis showed the mutated subunits can still assemble into IL-12, and in vitro bioassays demonstrated the assembled IL-12 induces IFN
The document discusses how AP-2α induces apoptosis in cancer cells. It finds that AP-2α activates the intrinsic apoptosis pathway by binding to the Bcl-2 promoter and repressing its transcription. This allows Bax to translocate to mitochondria, disrupt membrane potential, and release cytochrome c, activating caspase-9 and caspase-3. Downregulation of anti-apoptotic Bcl-2 is important for AP-2α-induced apoptosis. Overexpressing AP-2α enhances cancer cell chemosensitivity to various drugs.
The document discusses the use of pluripotent stem cell-derived cardiomyocytes as substrates for automated ion channel and electrophysiological analysis systems. Specifically, it describes how cardiomyocytes can be selected from mouse embryonic stem cells using puromycin, and how these cells express appropriate cardiac ion channels. It also demonstrates that the cardiomyocytes can be used on automated patch clamp systems like PatchXpress and QPatch to measure cardiac ion currents like INa, ICa, and IK, and to study the effects of drugs on these currents. The document establishes pluripotent stem cell-derived cardiomyocytes as a physiologically relevant and reproducible model for high-throughput electrophysiology screening.
Similar to Orai1-encoded store-operated channel and TRPC1-dependent IP3 receptor- operated channel (7)
Orai1-encoded store-operated channel and TRPC1-dependent IP3 receptor- operated channel
1. Separation of Siamese twins Orai1-encoded store-operated channel and TRPC1-dependent IP 3 receptor- operated channel Vladislav Zarayskiy, Ph.D. Ion Channel and Calcium Signaling Unit Boston University, Boston, MA
2. Ca 2+ influx pathways IP 3 IP 3 -sensitive Ca 2+ store Store-operated IP 3 receptor ? ? ? SERCA ? Ryanodine- sensitive store Ca 2+ Voltage-gated Ligand-gated Mechano-gated Other G PLC IP 3 Ca 2+ Physiological responses
4. CIF activates I CRAC CIF(-) CIF(+) CIF(-) CIF(+) RBL-2H3 mast cells Csutora et al., 2006 -2 -1 0 100 200 300 DES I (pA/pF) time (s) 0.1 BAPTA 20 Ca 2+ 130 Na + 165 Cs + -2 -1 -80 -40 0 40 80 V (mV) I (pA/pF)
5. iPLA 2 is crucial for CIF activation of I CRAC Csutora et al., 2006 0.1 BAPTA 20 Ca 2+ 130 Na + 165 Cs + a/s Control iPLA 2 ß I (pA/pF) -2 -1 0 100 200 300 DES time (s) -2 -1 -80 -40 0 40 80 V (mV) I (pA/pF) a/s Control a/s Control a/s Control iPLA 2 ß
6. iPLA 2 is crucial for I CRAC development Csutora et al., 2006 10 BAPTA 20 Ca 2+ 130 Na + 165 Cs + I (pA/pF) time (s) + rec- iPLA 2 ß -2 -1 0 100 200 300 + rec- iPLA 2 ß -2 -1 80 40 -40 -80 I (pA/pF) V (mV) 0 a/s a/s DES * 0 25 µM 25 µM 0 20 40 60 80 100 Current (%) ( S ) -BEL ( R ) -BEL Control -2 -1 -100 -50 50 V (mV) I (pA/pF) 0 0 ( S ) -BEL ( R ) -BEL
7. depleted Ca 2+ stores CIF Ca 2+ Store-Operated iPLA 2 CaM R G PLC SERCA IP 3 Ca 2+ IP 3 R physiological responses SOC (CRAC) ER/SR LysoPL Bolotina, Science STKE, July 2004 Ca 2+ influx pathways Ca 2+ IP 3 R IP 3 ROC IP 3 Receptor-Operated
8. Differences in the mechanism of activation : Store depletion versus IP 3 and the role of iPLA 2 β Zarayskiy et al., 2007 I CRAC 20 Ca 2+ 130 Na + 165 Cs + I IP3ROC IP 3 BAPTA 20 Ca 2+ 130 Na + 165 Cs + Control iPLA 2 a/s -2 -1 1 -80 -40 40 V (mV) I (pA/pF) -2 -1 0 0.01 0.1 1 10 100 IP 3 concentration (8) (3) (3) (4) (4) (5) µM I (pA/pF) a/s iPLA 2 I (pA/pF) -2 -1 0 100 200 Time (s) DES Control a/s iPLA 2 Control iPLA 2 a/s V (mV) I (pA/pF) -2 -1 1 -80 -40 40 -2 -1 0 100 200 Time (s) I (pA/pF) DES Control a/s iPLA 2 BAPTA
9. I CRAC and I IP3ROC : Biophysical and molecular discrimination Zarayskiy et al., 2007 Inwardly rectifying ~ Linear Inwardly rectifying Inwardly rectifying Current-voltage relationship (I/V) (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes Yes Inhibition by DES Yes Yes Inhibition by 2-APB Inhibition No effect TRPC1/IP 3 R peptide Inhibition No effect Calyculin A Present Absent iPLA 2 β-deficient cells Absent Present TRPC1-deficient cells Present Present STIM1-deficient cells Present Absent Orai1-deficient cells P Ca2+ : P Na+ <0.4 P Ca2+ : P Na+ >1000 Selectivity >+50 mV +5 mV > +50 mV >+50 mV Reversal potential: (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes No Requirement of IP 3 No Yes Activation by Ca 2+ store depletion I IP3ROC I CRAC
10. Zarayskiy et al., 2007 I CRAC I IP3ROC Differences in cation selectivity : Ca 2+ -selective I CRAC and cation-selective I IP3ROC. 20 Ca 2+ 165 Cs + 130 Na + 130 NMDG + 130 Cs + a/s iPLA 2 130 NMDG + 130 Cs + 0 50 100 Control NMDG + Cs + Current, (%) ( 6 ) ( 6 ) (5) 0 50 100 Current, (%) Control NMDG + Cs + ( 4 ) ( 4 ) ( 3 ) BAPTA IP 3 BAPTA 20 Ca 2+ 165 Cs + 130 Na +
11. Zarayskiy et al., 2007 I CRAC I IP3ROC Intracellular Cs + as a reason for inward rectification of I IP3ROC . 165 Cs + 20 Ca 2+ 130 Na + 130 Na + 130 Na + V (mV) I (pA/pF) -1 1 -80 -40 40 [Na + ] i [Cs + ] i [Na + ] i [Cs + ] i V( mV) I (pA/pF) -1 1 -80 -40 0 40 a/s iPLA 2 [Na + ] i [Cs + ] i BAPTA IP 3 BAPTA 20 Ca 2+ 130 Na + 165 Cs + 165 Cs +
12. I CRAC and I IP3ROC : Biophysical and molecular discrimination Zarayskiy et al., 2007 Inwardly rectifying ~ Linear Inwardly rectifying Inwardly rectifying Current-voltage relationship (I/V) (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes Yes Inhibition by DES Yes Yes Inhibition by 2-APB Inhibition No effect TRPC1/IP 3 R peptide Inhibition No effect Calyculin A Present Absent iPLA 2 β-deficient cells Absent Present TRPC1-deficient cells Present Present STIM1-deficient cells Present Absent Orai1-deficient cells P Ca2+ : P Na+ <0.4 P Ca2+ : P Na+ >1000 Selectivity >+50 mV +5 mV > +50 mV >+50 mV Reversal potential: (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes No Requirement of IP 3 No Yes Activation by Ca 2+ store depletion I IP3ROC I CRAC
13. I CRAC I IP3ROC Zarayskiy et al., 2007 Differences in molecular identity Orai1-encoded I CRAC and TRPC1-dependent I IP3ROC TRPC1 a/s Orai1 siRNA V (mV) I (pA/pF) -1 1 -80 -40 40 V (mV) I (pA/pF) -1 1 -80 -40 40 TRPC1 a/s Control Orai1 siRNA BAPTA IP 3 BAPTA
14. I CRAC I IP3ROC Zarayskiy et al., 2007 Differences in molecular identity Orai1-encoded I CRAC and TRPC1-dependent I IP3ROC Orai1 siRNA V (mV) I (pA/pF) -1 1 -80 -40 40 TRPC1 a/s V (mV) I (pA/pF) -1 1 -80 -40 40 IP 3 BAPTA Control CRAC+IP 3 ROC -1 1 -80 -40 40 80 V (mV) I (pA/pF)
15. I CRAC I IP3ROC Zarayskiy et al., 2007 Differences in molecular identity Orai1-encoded I CRAC and TRPC1-dependent I IP3ROC Orai1 siRNA V (mV) I (pA/pF) -1 1 -80 -40 40 TRPC1 a/s V (mV) I (pA/pF) -1 1 -80 -40 40 V (mV) I (pA/pF) -1 1 -80 -40 40 TRPC1 a/s Control Orai1 siRNA IP 3 BAPTA
16. I CRAC and I IP3ROC : Biophysical and molecular discrimination Zarayskiy et al., 2007 Inwardly rectifying ~ Linear Inwardly rectifying Inwardly rectifying Current-voltage relationship (I/V) (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes Yes Inhibition by DES Yes Yes Inhibition by 2-APB Inhibition No effect TRPC1/IP 3 R peptide Inhibition No effect Calyculin A Present Absent iPLA 2 β-deficient cells Absent Present TRPC1-deficient cells Present Present STIM1-deficient cells Present Absent Orai1-deficient cells P Ca2+ : P Na+ <0.4 P Ca2+ : P Na+ >1000 Selectivity >+50 mV +5 mV > +50 mV >+50 mV Reversal potential: (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes No Requirement of IP 3 No Yes Activation by Ca 2+ store depletion I IP3ROC I CRAC
17. I CRAC I IP3ROC Zarayskiy et al., 2007 I IP3ROC but not I CRAC depends on conformational coupling with IP 3 R : differential effects of TRPC1 / IP 3 R binding peptide TRPC1 a/s Peptide (+) (-) V (mV) I (pA/pF) -1 1 -80 -40 40 Orai1 siRNA Peptide (+) (-) V (mV) I (pA/pF) -1 1 -80 -40 0 40 BAPTA IP 3 BAPTA
18. I CRAC and I IP3ROC : Biophysical and molecular discrimination Zarayskiy et al., 2007 Inwardly rectifying ~ Linear Inwardly rectifying Inwardly rectifying Current-voltage relationship (I/V) (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes Yes Inhibition by DES Yes Yes Inhibition by 2-APB Inhibition No effect TRPC1/IP 3 R peptide Inhibition No effect Calyculin A Present Absent iPLA 2 β-deficient cells Absent Present TRPC1-deficient cells Present Present STIM1-deficient cells Present Absent Orai1-deficient cells P Ca2+ : P Na+ <0.4 P Ca2+ : P Na+ >1000 Selectivity >+50 mV +5 mV > +50 mV >+50 mV Reversal potential: (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes No Requirement of IP 3 No Yes Activation by Ca 2+ store depletion I IP3ROC I CRAC
19. I CRAC and I IP3ROC : Biophysical and molecular discrimination Zarayskiy et al., 2007 Inwardly rectifying ~ Linear Inwardly rectifying Inwardly rectifying Current-voltage relationship (I/V) (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes Yes Inhibition by DES Yes Yes Inhibition by 2-APB Inhibition No effect TRPC1/IP 3 R peptide Inhibition No effect Calyculin A Present Absent iPLA 2 β-deficient cells Absent Present TRPC1-deficient cells Present Present STIM1-deficient cells Present Absent Orai1-deficient cells P Ca2+ : P Na+ <0.4 P Ca2+ : P Na+ >1000 Selectivity >+50 mV +5 mV > +50 mV >+50 mV Reversal potential: (Na + ,Ca 2+ ) out / (Cs + ) in (Na + ,Ca 2+ ) out / (Na + ,Cs + ) in Yes No Requirement of IP 3 No Yes Activation by Ca 2+ store depletion I IP3ROC I CRAC
20. depleted Ca 2+ stores CIF Ca 2+ iPLA 2 CaM R G PLC SERCA Ca 2+ Na + IP 3 IP 3 Receptor-Operated Ca 2+ IP 3 R IP 3 R physiological responses IP 3 ROC SOC ER/SR LysoPL Ca 2+ influx pathways Orai1 TRPC1 (+other TRPCs?) Store-Operated
21. I IP3ROC depends on Ca 2+ buffering Zarayskiy et al., 2007 0.5 1 1.5 20 40 60 80 100 I (pA/pF) sec 0 +80 mV 10 µM IP 3 + 1 mM BAPTA (4) (4) IP 3 BAPTA
22. BAPTA + OAG OAG activates additional current besides I CRAC 20 Ca 2+ 130 Na + 165 Cs + 2 1 -4 -2 0 2 Current (pA/pF) ( 2-1) -2 -1 0 1 2 0 100 200 2 1 Current (normalized) Time (sec) +80mV -80mV BAPTA
23. Calyculin A inhibits I IP3ROC but not I CRAC Zarayskiy et al., 2007 V (mV) CalyA a/s iPLA 2 Control I (pA/pF) V (mV) I (pA/pF) Control CalyA -1 1 -80 -40 40 -1 1 -80 -40 40 I CRAC I IP3ROC BAPTA IP 3 BAPTA
24. I IP3ROC can be inhibited from inside and outside Zarayskiy et al., 2007 I (pA/pF) -2 -1 1 2 -80 -40 40 80 V (mV) IP 3 + BAPTA -2 -1 0 1 2 Control 1 µM La 3+ 500 µg/ml Heparin iPLA 2 a/s iPLA 2 a/s Control La 3 + Heparin IP 3 BAPTA
25.
26. depleted Ca 2+ stores CIF Ca 2+ iPLA 2 CaM R G PLC SERCA Ca 2+ Na + IP 3 IP 3 Receptor-Operated Ca 2+ IP 3 R IP 3 R physiological responses IP 3 ROC SOC ER/SR LysoPL Ca 2+ influx pathways Orai1 TRPC1 (+other TRPCs?) Store-Operated