Stanford ChEM-H solicited seed grant proposals from the Stanford community of postdoctoral fellows for exploratory projects that align with the Institute’s mission. Successful proposals combined the complementary expertise of two or more postdoctoral fellows seeking to collaboratively explore a potentially transformative new idea with the support of their mentors. The four proposals selected were each funded with $50,000 for one year.
"The Effect of Pediatric Specific Hypertrophic Cardiomyopathy Mutations on the Biomechanics of beta-Cardiac Myosin at the Molecular and Cellular Level"
Postdoctoral Scholars:
Arjun Adhikari, Spudich Lab
Kristina Bezold, Bernstein Lab
Alexandre Ribeiro, Pruitt Lab
Mentors:
Daniel Bernstein, Pediatrics - Cardiology
Beth Pruitt, Mechanical Engineering
James Spudich, Biochemistry
Project Summary:
Hypertrophic Cardiomyopathy (HCM) is a genetic disease that affects 1 in 500 people. In infants it is particularly severe and is the leading cause of sudden cardiac death in pediatric populations. A high percentage of HCM is attributed to mutations in ß-cardiac myosin, the protein responsible for heart contraction. This study explores how pediatric specific HCM mutations can alter the biochemical and biomechanical properties of ß-cardiac myosin at a molecular and cellular level. At a molecular level, this study aims to understand how HCM mutations affect the biomechanics of purified human ß-cardiac myosin. At a cellular level, cardiomyocytes derived from human induced pluripotent stem cells (iPSC) engineered to express each mutation will be used to study how HCM affects the power output and signaling pathways of these cells. The current standard care for patients with HCM involves beta-blockers and ACE inhibitors, which treat the symptoms, but not the disease. The results from this study will increase the understanding of how genetic mutations that cause HCM lead to the presentation of the disease, and will uncover potential targets for therapeutic design to treat the cause of the disease instead of the symptoms.
This project brings together three postdoctoral researchers to form an interdisciplinary team spanning the wide range of skill sets necessary to study this important problem in cardiac disease. Dr. Adhikari is an expert in biophysics and cell biomechanics and has solid knowledge of cell and molecular biology. During his doctoral work with Dr. Dunn (Stanford Chemical Engineering), he gained deep understanding of how mechanical forces can influence biological function at a molecular and cellular level. He is currently a postdoctoral researcher in Dr. James Spudich’s lab in the Biochemistry Department. Dr. Bezold is an expert in muscle biology and biomechanics and has significant experience in stem cell biology. She gained in depth knowledge of muscle biology while working in the lab of Dr. Samantha Harris (UC Davis). Currently, she is a postdoctoral fellow in the lab of Dr. Daniel Bernstein in Pediatrics/Cardiology, where she has led the effort to generate iPSC lines with HCM mutations using the CRISPR/Cas9 system. Dr. Ribeiro is a highly skilled mechanical engineer and has considerable experience in cell biology. During his graduate work, under the guidance of Dr. Kris Dahl (Chemical Engineer, Carnegie Mellon University), he studied nuclear mechanics during cell migration. During his postdoctoral work in the lab of Dr. Beth Pruitt in Mechanical Engineering, he has been instrumental in developing patterned platforms that lead to more mature phenotype of iPSC cardiomyocytes.
"Bioorthogonal Secretome Labeling for the Study of Mammalian Aging and Rejuvenation"
Postdoctoral Scholars:
Kyle Brewer, Wyss-Coray Lab
Justin Kim, Bertozzi Lab
Mentors:
Carolyn Bertozzi, Chemistry
Tony Wyss-Coray, Neurology & Neurological Sciences
Project Summary:
Aging is the largest risk factor for heart disease, cancer, stroke, dementia, and diabetes, as well as a major impediment to quality of life. Recently, protein factors present in young blood have been shown to rejuvenate the heart, brain, and muscle in aged mice. In addition, protein factors in old blood have been shown to impair cognitive function in mice. To find how these proteins can best be exploited to benefit human health, tissues in mice will be examined to determine where these factors are produced, as well as where they localize. To achieve this goal, recent developments in chemical biology will be used to incorporate non-natural amino acids (nnAAs) into the mouse proteome to target secreted factors for bioorthogonal labeling. The genetic system to incorporate the nnAAs will be introduced specifically into the brain and liver of young mice via lentiviral delivery, and the proteins secreted from these regions will be collected in plasma. After injecting the labeled, young plasma into unlabeled, old mice, whether these factors can cross the blood-brain barrier and where these factors distribute in the brain will be determined. This system can then be applied more widely to find other areas these proteins have specific effects to rejuvenate tissues.
This project leverages the complementary expertise of two postdoctoral researchers with knowledge of protein and synthetic chemistry to address an important question in human biology. Dr. Brewer has a background in biophysics including protein structure, protein dynamics, and protein-protein interactions and has extensive experience using novel compounds and biochemistry to produce labeled proteins for biophysical and biological purposes. Since joining the Wyss-Coray Lab as a postdoctoral researcher, he has been developing novel methods to study the mouse secretome, placing an emphasis on examining natively and in vitro the aging and rejuvenation factors being discovered in the lab. Dr. Kim’s graduate work was performed in the field of complex natural product total synthesis, and he has expertise in non-natural amino acid incorporation and bioorthogonal labeling. His postdoctoral research in the Bertozzi Lab involves the use of non-natural amino acids in conjunction with bioorthogonal chemistry to study the mechanism by which secreted virulence factors mediate the pathogenesis of tuberculosis in a Mycobacterium marinum-zebrafish model system.
"A Chemical-Genetic Investigation of Non-Apoptotic Cell Death Mechanisms"
Postdoctoral Scholars:
Jennifer Yinuo Cao, Dixon Lab
Cole Dovey, Carette Lab
Mentors:
Jan Carette, Microbiology & Immunology
Scott Dixon, Biology
Project Summary:
Regulated cell death enables the elimination of old, damaged or unwanted cells from the body. This process is essential to maintain homeostasis and prevent disease. A central aim of the Carette and Dixon laboratories is to better understand the structure and function of different cell death pathways which will ultimately allow substantial improvements in human health. The goal of this proposal is to better understand the operation of two recently described non-apoptotic cell death pathways, termed necroptosis and ferroptosis. The regulation of these pathways at the genetic and biochemical levels remains largely a mystery. To address this problem, this proposal combines state-of-the-art methods in human genetics and chemical biology pioneered in the Carette and Dixon laboratories. Specifically, a powerful, high-throughput chemical screening approach will be applied to identify small molecule modulators of necroptosis. Concurrently, an innovative, genome-wide genetic screening platform will be employed to investigate the genetic architecture of ferroptosis. This research aims to clarify the regulation of non-apoptotic cell death pathways and identify new druggable targets that ultimately will enable more precise therapeutic control of cell death.
Dr. Cao completed her doctoral studies in the Frappier Lab at the University of Toronto, where she used proteomic and biochemical techniques to identify novel herpes virus-host protein interactions which revealed novel connections between herpes virus and reactive oxygen species (ROS) production. She is currently studying the regulation of ferroptosis under the mentorship of Dr. Scott Dixon and has helped develop a novel high-throughput, time-lapse imaging platform that can be used to examine the effects of various bioactive small molecules on cell death. Dr. Dovey has over a decade of experience in molecular pathogenesis research, with expertise in genetics and biochemistry. During his graduate training with Jeffery Cox at UCSF, he applied a biochemical and bacterial genetic approach toward understanding the regulation of virulence in Mycobacterium tuberculosis. As a postdoctoral scholar in the Carette Lab, he has expanded his field of expertise by investigating the cell biological and genetic mechanisms of regulated cell death pathways including necroptosis. Dr. Cao and Dr. Dovey’s expertise and research interests will be combined synergistically in the effort to understand cell death mechanisms that are critical for human health.
Publications:
A Genome-wide Haploid Genetic Screen Identifies Regulators of Glutathione Abundance and Ferroptosis Sensitivity. J.Y. Cao, A. Poddar, L. Magtanong, J.H. Lumb, T.R. Mileur, M.A. Reid, C.M. Dovey, J. Wang, J.W. Locasale, E. Stone, S.P.C. Cole, J.E. Carette, S.J. Dixon. Cell Reports. 2019, 26, 1544–1556.
MLKL Requires the Inositol Phosphate Code to Execute Necroptosis. C.M. Dovey, J. Diep, B.P. Clarke, A.T. Hale, D.E. McNamara, H. Guo, N.W. Brown Jr, J.Y. Cao, C.R. Grace, P.J. Gough, J. Bertin, S.J. Dixon, D. Fiedler, E.S. Mocarski, W.J. Kaiser, T. Moldoveaunu, J.D. York, J.E. Carette. Molecular Cell. 2018, 70, 936–948.
"Lanthanide Resonance Energy Transfer (LRET)-Based Imaging of GPR126 Signaling In Vivo"
Postdoctoral Scholars:
Paulina Ciepla, Chen Lab
Ana Meireles de Sousa, Talbot Lab
Mentors:
James Chen, Chemical and Systems Biology & Developmental Biology
William Talbot, Developmental Biology
Project Summary:
G-protein coupled receptors (GPCR) are essential for many aspects of human development, physiology, and behavior. These receptors are also important drug targets. There is intense interest in understanding how different signals activate GPCRs, but progress has been hindered by a lack of methods to visualize active receptors in living organisms. The goal of this project is to establish a new imaging method using lanthanides (metallic chemical elements) to visualize the interactions of GPCRs and other proteins in cells, tissues, and embryos. The aims are to develop an imaging approach with much greater sensitivity than current methods and to use it to visualize interactions of a particular receptor, GPR126. GPR126 controls the formation of the myelin sheath in nerves of humans and other vertebrates. GPR126 role was first discovered in zebrafish, a vertebrate model organism with fundamental similarities to humans. In addition, the zebrafish embryo is transparent, making it ideally suited for this application. Understanding the mechanisms that activate GPR126 may lead to new ideas for nerve disease therapies. Additionally, development of lanthanide-based imaging as a general approach to study a wide range of molecular interaction in living organisms would have a broad impact in many fields of the biomedical sciences.
Dr. Ciepla is a postdoctoral fellow in Chemical and Systems Biology and an expert in designing and optimizing chemical probes of protein function. During her PhD at Imperial College London (UK), Paulina’s research focused on the development of imaging tools to study posttranslational modifications of Hedgehog protein. Realizing that zebrafish provide a powerful tool in which to study the functional effects of drugs and chemical probes, she chose to further develop her skills in the lab of Dr. James Chen. Dr. Sousa is a postdoctoral fellow in Developmental Biology, with an excellent track record in biochemistry, cell biology and animal genetics. During her PhD at the Wellcome Trust Centre for Cell Biology (Scotland/Portugal), she studied the mechanisms that control microtubule dynamics and spindle formation during cell division through the combined use of forward and reverse genetics and varied biochemical approaches. During her time at in the Talbot Lab, Ana has focused on nervous system development and neuroimmune interactions. Dr. Sousa’s knowledge of GPR126 biology and zebrafish genetics together with Dr. Ciepla’s background in chemical probe-based imaging provides a unique combination of expertise to enable this important mechanistic study.
