Contact
Biomedical Research Center251 Bayview Boulevard
Suite 200
Baltimore, MD 21224
Education
Ph.D - University of Maryland, Baltimore
Research Interests
Kathleen Trychta received her Ph.D. in 2019 from University of Maryland, Baltimore. As a part of Dr. Harvey’s lab at NIDA she has identified a cellular phenomenon, ER exodosis, in which there is a mass redistribution of normally ER resident proteins into the extracellular space following an ER calcium associated insult. Current research avenues include examining the consequences of ER exodosis, identifying biomarkers to monitor the composition of the ER proteome, and evaluating whether ER proteostasis exhibits relevant changes in the clinical context of disease.
Publications
Selected Publications
2018
Trychta, Kathleen A; Back, Susanne; Henderson, Mark J; Harvey, Brandon K
KDEL Receptors Are Differentially Regulated to Maintain the ER Proteome under Calcium Deficiency. Journal Article
In: Cell Rep, vol. 25, no. 7, pp. 1829–1840, 2018, ISSN: 2211-1247 (Electronic).
@article{Trychta:2018aab,
title = {KDEL Receptors Are Differentially Regulated to Maintain the ER Proteome under Calcium Deficiency.},
author = {Kathleen A Trychta and Susanne Back and Mark J Henderson and Brandon K Harvey},
url = {https://www.ncbi.nlm.nih.gov/pubmed/30428351},
doi = {10.1016/j.celrep.2018.10.055},
issn = {2211-1247 (Electronic)},
year = {2018},
date = {2018-11-13},
journal = {Cell Rep},
volume = {25},
number = {7},
pages = {1829--1840},
address = {Molecular Mechanisms of Cellular Stress and Inflammation, Intramural Research Program, National Institute on Drug Abuse, Baltimore, MD 21224, USA.},
abstract = {Retention of critical endoplasmic reticulum (ER) luminal proteins needed to carry out diverse functions (e.g., protein synthesis and folding, lipid metabolism) is mediated through a carboxy-terminal ER retention sequence (ERS) and its interaction with KDEL receptors. Here, we demonstrate that depleting ER calcium causes mass departure of ERS-containing proteins from cells by overwhelming KDEL receptors. In addition, we provide evidence that KDELR2 and KDELR3, but not KDELR1, are unfolded protein response (UPR) genes upregulated as an adaptive response to counteract the loss of ERS-containing proteins, suggesting previously unknown isoform-specific functions of the KDEL receptors. Overall, our findings establish that decreases in ER calcium change the composition of the ER luminal proteome and secretome, which can impact cellular functions and cell viability. The redistribution of the ER proteome from inside the cell to the outside has implications for dissecting the complex relationship of ER homeostasis with diverse disease pathologies.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
Retention of critical endoplasmic reticulum (ER) luminal proteins needed to carry out diverse functions (e.g., protein synthesis and folding, lipid metabolism) is mediated through a carboxy-terminal ER retention sequence (ERS) and its interaction with KDEL receptors. Here, we demonstrate that depleting ER calcium causes mass departure of ERS-containing proteins from cells by overwhelming KDEL receptors. In addition, we provide evidence that KDELR2 and KDELR3, but not KDELR1, are unfolded protein response (UPR) genes upregulated as an adaptive response to counteract the loss of ERS-containing proteins, suggesting previously unknown isoform-specific functions of the KDEL receptors. Overall, our findings establish that decreases in ER calcium change the composition of the ER luminal proteome and secretome, which can impact cellular functions and cell viability. The redistribution of the ER proteome from inside the cell to the outside has implications for dissecting the complex relationship of ER homeostasis with diverse disease pathologies.
Trychta, Kathleen A; Heathward, Emily J; Sulima, Agnieszka; Back, Susanne; Farokhnia, Mehdi; Richie, Christopher T; Leggio, Lorenzo; Rice, Kenner C; Harvey, Brandon K
Extracellular esterase activity as an indicator of endoplasmic reticulum calcium depletion. Journal Article
In: Biomarkers, pp. 1–10, 2018, ISSN: 1366-5804 (Electronic); 1354-750X (Linking).
@article{Trychta:2018aa,
title = {Extracellular esterase activity as an indicator of endoplasmic reticulum calcium depletion.},
author = {Kathleen A Trychta and Emily J Heathward and Agnieszka Sulima and Susanne Back and Mehdi Farokhnia and Christopher T Richie and Lorenzo Leggio and Kenner C Rice and Brandon K Harvey},
url = {https://www.ncbi.nlm.nih.gov/pubmed/30095301},
doi = {10.1080/1354750X.2018.1490968},
issn = {1366-5804 (Electronic); 1354-750X (Linking)},
year = {2018},
date = {2018-08-10},
urldate = {2018-08-10},
journal = {Biomarkers},
pages = {1--10},
address = {a Molecular Mechanisms of Cellular Stress and Inflammation , Intramural Research Program, National Institute on Drug Abuse , Baltimore , MD , USA.},
abstract = {CONTEXT: Endoplasmic reticulum (ER) calcium depletion is associated with diverse diseases, including cardiac, hepatic, and neurologic diseases. OBJECTIVE: The aim of the present study was to identify and characterize an endogenous protein that could be used to monitor ER calcium depletion comparably to a previously described exogenous reporter protein. MATERIALS AND METHODS: The use of a selective esterase-fluorescein diester pair allowed for carboxylesterase activity in extracellular fluid to be measured using a fluorescent readout. Cell culture media from three different cell lines, rat plasma, and human serum all possess quantifiable amounts of esterase activity. RESULTS: Fluorescence produced by the interaction of carboxylesterases with a fluorescein diester substrate tracks with pharmacological and physiological inducers of ER calcium depletion. The fluorescence measured for in vitro and in vivo samples were consistent with ER calcium depletion being the trigger for increased esterase activity. DISCUSSION: Decreased luminal ER calcium causes ER resident esterases to be released from the cell, and, when assessed concurrently with other disease biomarkers, these esterases may provide insight into the role of ER calcium homeostasis in human diseases. CONCLUSION: Our results indicate that carboxylesterases are putative markers of ER calcium dysfunction.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
CONTEXT: Endoplasmic reticulum (ER) calcium depletion is associated with diverse diseases, including cardiac, hepatic, and neurologic diseases. OBJECTIVE: The aim of the present study was to identify and characterize an endogenous protein that could be used to monitor ER calcium depletion comparably to a previously described exogenous reporter protein. MATERIALS AND METHODS: The use of a selective esterase-fluorescein diester pair allowed for carboxylesterase activity in extracellular fluid to be measured using a fluorescent readout. Cell culture media from three different cell lines, rat plasma, and human serum all possess quantifiable amounts of esterase activity. RESULTS: Fluorescence produced by the interaction of carboxylesterases with a fluorescein diester substrate tracks with pharmacological and physiological inducers of ER calcium depletion. The fluorescence measured for in vitro and in vivo samples were consistent with ER calcium depletion being the trigger for increased esterase activity. DISCUSSION: Decreased luminal ER calcium causes ER resident esterases to be released from the cell, and, when assessed concurrently with other disease biomarkers, these esterases may provide insight into the role of ER calcium homeostasis in human diseases. CONCLUSION: Our results indicate that carboxylesterases are putative markers of ER calcium dysfunction.
2017
Wires, Emily S; Trychta, Kathleen A; Back, Susanne; Sulima, Agnieszka; Rice, Kenner C; Harvey, Brandon K
High fat diet disrupts endoplasmic reticulum calcium homeostasis in the rat liver. Journal Article
In: J Hepatol, vol. 67, no. 5, pp. 1009–1017, 2017, ISSN: 1600-0641 (Electronic); 0168-8278 (Linking).
@article{cite-key,
title = {High fat diet disrupts endoplasmic reticulum calcium homeostasis in the rat liver.},
author = {Emily S Wires and Kathleen A Trychta and Susanne Back and Agnieszka Sulima and Kenner C Rice and Brandon K Harvey},
url = {https://www.ncbi.nlm.nih.gov/pubmed/28596111},
doi = {10.1016/j.jhep.2017.05.023},
issn = {1600-0641 (Electronic); 0168-8278 (Linking)},
year = {2017},
date = {2017-07-17},
urldate = {2017-07-17},
journal = {J Hepatol},
volume = {67},
number = {5},
pages = {1009--1017},
address = {Molecular Mechanisms of Cellular Stress and Inflammation Unit, Intramural Research Program, National Institute on Drug Abuse, Baltimore, MD 21224, USA.},
abstract = {BACKGROUND & AIMS: Disruption to endoplasmic reticulum (ER) calcium homeostasis has been implicated in obesity, however, the ability to longitudinally monitor ER calcium fluctuations has been challenging with prior methodologies. We recently described the development of a Gaussia luciferase (GLuc)-based reporter protein responsive to ER calcium depletion (GLuc-SERCaMP) and investigated the effect of a high fat diet on ER calcium homeostasis. METHODS: A GLuc-based reporter cell line was treated with palmitate, a free fatty acid. Rats intrahepatically injected with GLuc-SERCaMP reporter were fed a cafeteria diet or high fat diet. The liver and plasma were examined for established markers of steatosis and compared to plasma levels of SERCaMP activity. RESULTS: Palmitate induced GLuc-SERCaMP release in vitro, indicating ER calcium depletion. Consumption of a cafeteria diet or high fat pellets correlated with alterations to hepatic ER calcium homeostasis in rats, shown by increased GLuc-SERCaMP release. Access to ad lib high fat pellets also led to a corresponding decrease in microsomal calcium ATPase activity and an increase in markers of hepatic steatosis. In addition to GLuc-SERCaMP, we have also identified endogenous proteins (endogenous SERCaMPs) with a similar response to ER calcium depletion. We demonstrated the release of an endogenous SERCaMP, thought to be a liver esterase, during access to a high fat diet. Attenuation of both GLuc-SERCaMP and endogenous SERCaMP was observed during dantrolene administration. CONCLUSIONS: Here we describe the use of a reporter for in vitro and in vivo models of high fat diet. Our results support the theory that dietary fat intake correlates with a decrease in ER calcium levels in the liver and suggest a high fat diet alters the ER proteome. Lay summary: ER calcium dysregulation was observed in rats fed a cafeteria diet or high fat pellets, with fluctuations in sensor release correlating with fat intake. Attenuation of sensor release, as well as food intake was observed during administration of dantrolene, a drug that stabilizes ER calcium. The study describes a novel technique for liver research and provides insight into cellular processes that may contribute to the pathogenesis of obesity and fatty liver disease.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
BACKGROUND & AIMS: Disruption to endoplasmic reticulum (ER) calcium homeostasis has been implicated in obesity, however, the ability to longitudinally monitor ER calcium fluctuations has been challenging with prior methodologies. We recently described the development of a Gaussia luciferase (GLuc)-based reporter protein responsive to ER calcium depletion (GLuc-SERCaMP) and investigated the effect of a high fat diet on ER calcium homeostasis. METHODS: A GLuc-based reporter cell line was treated with palmitate, a free fatty acid. Rats intrahepatically injected with GLuc-SERCaMP reporter were fed a cafeteria diet or high fat diet. The liver and plasma were examined for established markers of steatosis and compared to plasma levels of SERCaMP activity. RESULTS: Palmitate induced GLuc-SERCaMP release in vitro, indicating ER calcium depletion. Consumption of a cafeteria diet or high fat pellets correlated with alterations to hepatic ER calcium homeostasis in rats, shown by increased GLuc-SERCaMP release. Access to ad lib high fat pellets also led to a corresponding decrease in microsomal calcium ATPase activity and an increase in markers of hepatic steatosis. In addition to GLuc-SERCaMP, we have also identified endogenous proteins (endogenous SERCaMPs) with a similar response to ER calcium depletion. We demonstrated the release of an endogenous SERCaMP, thought to be a liver esterase, during access to a high fat diet. Attenuation of both GLuc-SERCaMP and endogenous SERCaMP was observed during dantrolene administration. CONCLUSIONS: Here we describe the use of a reporter for in vitro and in vivo models of high fat diet. Our results support the theory that dietary fat intake correlates with a decrease in ER calcium levels in the liver and suggest a high fat diet alters the ER proteome. Lay summary: ER calcium dysregulation was observed in rats fed a cafeteria diet or high fat pellets, with fluctuations in sensor release correlating with fat intake. Attenuation of sensor release, as well as food intake was observed during administration of dantrolene, a drug that stabilizes ER calcium. The study describes a novel technique for liver research and provides insight into cellular processes that may contribute to the pathogenesis of obesity and fatty liver disease.
Wires, Emily S; Henderson, Mark J; Yan, Xiaokang; Bäck, Susanne; Trychta, Kathleen A; Lutrey, Molly H; Harvey, Brandon K
Longitudinal monitoring of Gaussia and Nano luciferase activities to concurrently assess ER calcium homeostasis and ER stress in vivo Journal Article
In: PLOS ONE, vol. 12, no. 4, pp. 1-17, 2017.
@article{10.1371/journal.pone.0175481,
title = {Longitudinal monitoring of Gaussia and Nano luciferase activities to concurrently assess ER calcium homeostasis and ER stress in vivo},
author = {Emily S Wires and Mark J Henderson and Xiaokang Yan and Susanne Bäck and Kathleen A Trychta and Molly H Lutrey and Brandon K Harvey},
url = {https://pubmed.ncbi.nlm.nih.gov/28403212/},
doi = {10.1371/journal.pone.0175481},
year = {2017},
date = {2017-01-01},
urldate = {2017-01-01},
journal = {PLOS ONE},
volume = {12},
number = {4},
pages = {1-17},
publisher = {Public Library of Science},
abstract = {The endoplasmic reticulum (ER) is essential to many cellular processes including protein processing, lipid metabolism and calcium storage. The ability to longitudinally monitor ER homeostasis in the same organism would offer insight into progressive molecular and cellular adaptations to physiologic or pathologic states, but has been challenging. We recently described the creation of a Gaussia luciferase (GLuc)-based secreted ER calcium-modulated protein (SERCaMP or GLuc-SERCaMP) to longitudinally monitor ER calcium homeostasis. Here we describe a complementary tool to measure the unfolded protein response (UPR), utilizing a UPRE-driven secreted Nano luciferase (UPRE-secNLuc) to examine the activating transcription factor-6 (ATF6) and inositol-requiring enzyme 1 (IRE1) pathways of the UPR. We observed an upregulation of endogenous ATF6- and XBP1-regulated genes following pharmacologically-induced ER stress that was consistent with responsiveness of the UPRE sensor. Both GLuc and NLuc-based reporters have favorable properties for in vivo studies, however, they are not easily used in combination due to overlapping substrate activities. We describe a method to measure the enzymatic activities of both reporters from a single sample and validated the approach using culture medium and rat blood samples to measure GLuc-SERCaMP and UPRE-secNLuc. Measuring GLuc and NLuc activities from the same sample allows for the robust and quantitative measurement of two cellular events or cell populations from a single biological sample. This study is the first to describe the in vivo measurement of UPRE activation by sampling blood, using an approach that allows concurrent interrogation of two components of ER homeostasis.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
The endoplasmic reticulum (ER) is essential to many cellular processes including protein processing, lipid metabolism and calcium storage. The ability to longitudinally monitor ER homeostasis in the same organism would offer insight into progressive molecular and cellular adaptations to physiologic or pathologic states, but has been challenging. We recently described the creation of a Gaussia luciferase (GLuc)-based secreted ER calcium-modulated protein (SERCaMP or GLuc-SERCaMP) to longitudinally monitor ER calcium homeostasis. Here we describe a complementary tool to measure the unfolded protein response (UPR), utilizing a UPRE-driven secreted Nano luciferase (UPRE-secNLuc) to examine the activating transcription factor-6 (ATF6) and inositol-requiring enzyme 1 (IRE1) pathways of the UPR. We observed an upregulation of endogenous ATF6- and XBP1-regulated genes following pharmacologically-induced ER stress that was consistent with responsiveness of the UPRE sensor. Both GLuc and NLuc-based reporters have favorable properties for in vivo studies, however, they are not easily used in combination due to overlapping substrate activities. We describe a method to measure the enzymatic activities of both reporters from a single sample and validated the approach using culture medium and rat blood samples to measure GLuc-SERCaMP and UPRE-secNLuc. Measuring GLuc and NLuc activities from the same sample allows for the robust and quantitative measurement of two cellular events or cell populations from a single biological sample. This study is the first to describe the in vivo measurement of UPRE activation by sampling blood, using an approach that allows concurrent interrogation of two components of ER homeostasis.
2015
Henderson, Mark J AU -; Wires, Emily S AU -; Trychta, Kathleen A AU -; Yan, Xiaokang AU -; Harvey, Brandon K AU -
Monitoring Endoplasmic Reticulum Calcium Homeostasis Using a Gaussia Luciferase SERCaMP Journal Article
In: JoVE, no. 103, pp. e53199, 2015, ISBN: 1940-087X.
@article{AU---Henderson:2015aa,
title = {Monitoring Endoplasmic Reticulum Calcium Homeostasis Using a Gaussia Luciferase SERCaMP},
author = {Mark J AU - Henderson and Emily S AU - Wires and Kathleen A AU - Trychta and Xiaokang AU - Yan and Brandon K AU - Harvey},
url = {https://pubmed.ncbi.nlm.nih.gov/26383227/},
doi = {doi:10.3791/53199},
isbn = {1940-087X},
year = {2015},
date = {2015-01-01},
journal = {JoVE},
number = {103},
pages = {e53199},
publisher = {MyJoVE Corp},
abstract = {The endoplasmic reticulum (ER) contains the highest level of intracellular calcium, with concentrations approximately 5,000-fold greater than cytoplasmic levels. Tight control over ER calcium is imperative for protein folding, modification and trafficking. Perturbations to ER calcium can result in the activation of the unfolded protein response, a three-prong ER stress response mechanism, and contribute to pathogenesis in a variety of diseases. The ability to monitor ER calcium alterations during disease onset and progression is important in principle, yet challenging in practice. Currently available methods for monitoring ER calcium, such as calcium-dependent fluorescent dyes and proteins, have provided insight into ER calcium dynamics in cells, however these tools are not well suited for in vivo studies. Our lab has demonstrated that a modification to the carboxy-terminus of Gaussia luciferase confers secretion of the reporter in response to ER calcium depletion. The methods for using a luciferase based, secreted ER calcium monitoring protein (SERCaMP) for in vitro and in vivo applications are described herein. This video highlights hepatic injections, pharmacological manipulation of GLuc-SERCaMP, blood collection and processing, and assay parameters for longitudinal monitoring of ER calcium.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
The endoplasmic reticulum (ER) contains the highest level of intracellular calcium, with concentrations approximately 5,000-fold greater than cytoplasmic levels. Tight control over ER calcium is imperative for protein folding, modification and trafficking. Perturbations to ER calcium can result in the activation of the unfolded protein response, a three-prong ER stress response mechanism, and contribute to pathogenesis in a variety of diseases. The ability to monitor ER calcium alterations during disease onset and progression is important in principle, yet challenging in practice. Currently available methods for monitoring ER calcium, such as calcium-dependent fluorescent dyes and proteins, have provided insight into ER calcium dynamics in cells, however these tools are not well suited for in vivo studies. Our lab has demonstrated that a modification to the carboxy-terminus of Gaussia luciferase confers secretion of the reporter in response to ER calcium depletion. The methods for using a luciferase based, secreted ER calcium monitoring protein (SERCaMP) for in vitro and in vivo applications are described herein. This video highlights hepatic injections, pharmacological manipulation of GLuc-SERCaMP, blood collection and processing, and assay parameters for longitudinal monitoring of ER calcium.
2014
Henderson, Mark J; Wires, Emily S; Trychta, Kathleen A; Richie, Christopher T; Harvey, Brandon K
SERCaMP: a carboxy-terminal protein modification that enables monitoring of ER calcium homeostasis. Journal Article
In: Mol Biol Cell, vol. 25, no. 18, pp. 2828–2839, 2014, ISSN: 1939-4586 (Electronic); 1059-1524 (Linking).
@article{Henderson2014,
title = {SERCaMP: a carboxy-terminal protein modification that enables monitoring of ER calcium homeostasis.},
author = {Mark J Henderson and Emily S Wires and Kathleen A Trychta and Christopher T Richie and Brandon K Harvey},
url = {https://www.ncbi.nlm.nih.gov/pubmed/25031430},
doi = {10.1091/mbc.E14-06-1141},
issn = {1939-4586 (Electronic); 1059-1524 (Linking)},
year = {2014},
date = {2014-07-16},
urldate = {2014-07-16},
journal = {Mol Biol Cell},
volume = {25},
number = {18},
pages = {2828--2839},
address = {Intramural Research Program, National Institute on Drug Abuse, Baltimore, MD 21224.},
abstract = {Endoplasmic reticulum (ER) calcium homeostasis is disrupted in diverse pathologies, including neurodegeneration, cardiovascular diseases, and diabetes. Temporally defining calcium dysregulation during disease progression, however, has been challenging. Here we describe secreted ER calcium-monitoring proteins (SERCaMPs), which allow for longitudinal monitoring of ER calcium homeostasis. We identified a carboxy-terminal modification that is sufficient to confer release of a protein specifically in response to ER calcium depletion. A Gaussia luciferase (GLuc)-based SERCaMP provides a simple and sensitive method to monitor ER calcium homeostasis in vitro or in vivo by analyzing culture medium or blood. GLuc-SERCaMPs revealed ER calcium depletion in rat primary neurons exposed to various ER stressors. In vivo, ER calcium disruption in rat liver was monitored over several days by repeated sampling of blood. Our results suggest that SERCaMPs will have broad applications for the long-term monitoring of ER calcium homeostasis and the development of therapeutic approaches to counteract ER calcium dysregulation.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
Endoplasmic reticulum (ER) calcium homeostasis is disrupted in diverse pathologies, including neurodegeneration, cardiovascular diseases, and diabetes. Temporally defining calcium dysregulation during disease progression, however, has been challenging. Here we describe secreted ER calcium-monitoring proteins (SERCaMPs), which allow for longitudinal monitoring of ER calcium homeostasis. We identified a carboxy-terminal modification that is sufficient to confer release of a protein specifically in response to ER calcium depletion. A Gaussia luciferase (GLuc)-based SERCaMP provides a simple and sensitive method to monitor ER calcium homeostasis in vitro or in vivo by analyzing culture medium or blood. GLuc-SERCaMPs revealed ER calcium depletion in rat primary neurons exposed to various ER stressors. In vivo, ER calcium disruption in rat liver was monitored over several days by repeated sampling of blood. Our results suggest that SERCaMPs will have broad applications for the long-term monitoring of ER calcium homeostasis and the development of therapeutic approaches to counteract ER calcium dysregulation.
