Each year, postbacs from across the NIH gather at the annual Postbac Poster Day to share their research with the NIH community. We are excited to announce that a total of eight NICHD postbacs received an overall top 20 percent poster award (NIH-wide) and/or one of the three “Best Poster” NICHD awards at this year’s event. To honor this achievement, The NICHD Connection invited the winning postbacs to publish a synopsis of their research. Read below to learn more about the award-winning studies from several of our winners.
The 2015 postbac awardees include:
- Megan Bannon (Lilly lab, NIH-wide award)
- Nicket Dedhia (Yanovski lab, NIH-wide award)
- Vy Duong (Pfeifer lab, NIH-wide and NICHD award)
- Daniel Flores (Pfeifer lab, NIH-wide award)
- Robyn Kalwerisky (Schisterman & Mumford lab of the DIPHR, NIH-wide award)
- Jung Park (Hoffman lab, NIH-wide and NICHD award)
- Maya Sangesland (Levin lab, NIH-wide award)
- Nathan Thomas (Cashel lab, NICHD award)
Obesity-Linked Gene Unrelated to Bone Density
By Nicket Dedhia
Advisor: Dr. Jack Yanovski
Obesity is an increasing problem without a single solution. Researchers have hypothesized several contributors to obesity susceptibility, including genetic background, viral infections, and chemical exposures. In the NICHD Section on Growth and Obesity, we study the genetic contributions to obesity. In particular, we examine how a protein called the melanocortin 3 receptor (MC3R) relates to energy homeostasis and obesity.
Children who have specific mutations in both copies of the MC3R gene (which we call “double mutant” or DM) have greater BMI, body fat mass, body fat percentage, and insulin resistance than children who carry one mutated copy (heterozygous, HET) or only wildtype (WT) MC3R. According to previous work in our lab, mice carrying this human genetic variant also display increased adiposity, as well as reductions in bone mineral content and bone area. The objective of my current study was to investigate if humans with two copies of the obesity-linked MC3R variation also show reductions in bone parameters
We completed MC3R genotype analysis and measured bone mineral density using dual-energy X-ray absorptiometry imaging for 239 healthy adults. Consistent with previous research, the DM cohort displayed greater BMI, fat mass, and fat mass percentage when compared to WT and HET individuals. Additionally, African Americans constituted a greater percentage of the DM population, confirming our past observations that this polymorphism occurs more frequently in African Americans as compared with Caucasians.
However, unlike the mice, humans carrying two copies of the obesity-linked MC3R variant did not show any statistically significant changes in bone parameters. Although we report the largest cohort of DM-MC3R individuals studied to date, we hope to aggregate a larger number of individuals to better power further studies on their bone metrics, energy homeostasis, and brain-gut-adipose axis.
Cardiac Calcium Control Conundrum
By Vy Duong
Advisor: Dr. Karl Pfeifer
Every muscle in the human body requires calcium to contract, and the heart is no exception. The release of calcium in heart cells is tightly regulated. A disturbance in this process can lead to an abnormal heart rhythm known as an arrhythmia. If prolonged, arrhythmias can be fatal. But what factors control calcium balance in the cell, and how can we use this information to improve human health?
My current work focuses on Cardiac Calsequestrin (encoded by the gene Casq2), a calcium-binding protein that prevents the inappropriate release of calcium inside heart cells. Patients with genetic defects in Casq2 suffer from stress-induced cardiac arrhythmias (a condition called CPVT for catecholaminergic polymorphic ventricular tachycardia) and an abnormally slow basal heart rate.
In previous studies, our lab generated a Casq2 knockout mouse model. To assess whether this model effectively phenocopies the human disease, I and Daniel Flores, another postbac in the lab, recorded electrocardiograms (ECG) from Casq2 knockout and control mice under adrenaline-induced stress, and we measured basal heart rate.
The ECG data revealed that only Casq2 knockout mice respond with multiple arrhythmias when under stress. They also display a lower basal heart rate than control mice. These results indicate that our mouse model effectively phenocopies the overt symptoms of CPVT.
We noted, however, that not every mutant mouse showed stress-induced arrhythmias. We believe such variation within the mutants is not atypical to what one would see in human diseases with patient-to-patient phenotype variation. We hope to use this mouse model to further elucidate the role Casq2 plays in causing arrhythmias and to identify candidate genes and pathways to target for treatment.
Do Mice Make Up for Missing CASQ2?
By Daniel Flores
Advisor: Dr. Karl Pfeifer
Patients lacking functional Cardiac Calsequestrin (CASQ2) protein experience episodes of irregular heart muscle contractions (arrhythmia) while under stress, a condition known as catecholaminergic polymorphic ventricular tachycardia (CPVT). Most individuals with CPVT experience their first heart attack by twenty years of age—often without warning.
CASQ2, encoded by the Casq2 gene, is a heart-specific protein responsible for binding and releasing calcium ions in the sarcoplasmic reticulum. Preliminary data from electrocardiogram readings using Casq2 loss-of-function mouse models suggests that symptoms are less severe in mice that inherit a bad copy of Casq2 compared to mice that have induced Casq2 loss. From these findings, we hypothesize that mice with inherited loss of CASQ2 undergo a developmental adaptation to compensate for this deletion, reducing the severity of cardiac arrhythmia.
I began to address my hypothesis in September 2014, working alongside Vy Duong, another postbaccalaureate fellow in our laboratory. We discovered a number of challenges with our experimental plan. Specifically, RT-QPCR data of Casq2 expression revealed that our tamoxifen inducible Cre recombinase system is active even without the administration of tamoxifen to the mouse. This leads to a gradual, unintended deletion of Casq2. We also found that this accidental deletion appears to increase as the mice age.
To address these findings in the short term, we are working with juvenile mice, in which the uncontrolled Casq2 deletion is at a minimum. In the long term, we are exploring the use of an Adeno-associated virus vector that will deliver Cre recombinase to cardiomyocytes for Casq2 deletion in our mouse model. Obtaining a precise method of deleting Casq2 in adult mice will provide us with a reliable method to determine if genomic plasticity occurs in mice with an inherited loss of CASQ2.
Novel Modification to Alzheimer’s-linked Ion Channel
By Jung Park
Advisor: Dr. Dax Hoffman
Billions of interconnected neurons in the brain communicate with each other via trillions of synaptic connections. To make sense of this complex neuronal network, our lab specializes in the synaptic ion channels that regulate the excitability of individual neurons. Specifically, I am interested in the potassium voltage-gated channel Kv4.2 found in the CA1 neurons of the hippocampus—a region of the brain responsible for learning and memory.
Understanding Kv4.2’s regulation may provide new insight into how we tackle cognitive neurodegenerative disorders. For example, increased dendritic excitability induced by Kv4.2 deficiency contributes to neuronal dysfunction and memory deficits found in early stages of Alzheimer’s disease. Characterization of Kv4.2 trafficking and degradation mechanisms will help us appreciate the role of synaptic ion channel regulation in psychiatric illnesses.
Due to its dynamic regulation upon neuronal stimulation, Kv4.2 most likely undergoes post-translational modifications in response to changes in the environment. Currently, there are gaps in the literature on how such channels are modified by the ubiquitination pathway, a process that directs substrates towards various cellular fates, including lysosomal and proteasomal degradation, endosomal sorting, and the internalization of surface proteins.
We have identified that Kv4.2 trafficking and degradation is regulated by ubiquitin. Our optimized protocol allows for an unprecedented, high-resolution visualization of Kv4.2 ubiquitination. However, our most recent findings indicate that Kv4.2 may undergo a novel ubiquitination pathway. Should our data hold true, our discovery would be among the first of its kind for synaptic ion channels.
Who’s Helping Retrotransposons Fit In?
By Maya Sangesland
Advisor: Dr. Henry Levin
Retrotransposons are mobile portions of DNA that are able to move from one location to another in a host genome through an RNA intermediate. Our lab uses the yeast retrotransposon Tf1— of particular interest as it mimics the life cycle of many mammalian retroviruses, such as HIV-1— to study host factors responsible for mediating integration of this retrotransposon into the genome.
Last year*, we conducted a systematic screen of each non-essential gene (roughly 3000 genes) in Schizosaccharomyces pombe yeast and identified 116 candidate genes that might contribute to integration of this transposable element. Through gene ontology analysis, we have grouped the identified candidates based on function, which includes DNA repair, chromatin modification, nucleosome assembly, transcription, as well as cell-cycle regulation.
Building upon my previous work, this year we focused on a specific histone variant, H2A.Z, which we believe may be directly or indirectly interacting with the Tf1 integrase protein and therefore facilitating integration of Tf1 at promoter regions. Currently, I am working on affinity pull-down experiments, which may reveal any Tf1-related binding partners with H2A.Z. We hope that our work will help elucidate the mechanisms employed during retroviral propagation and integration site targeting during infection.
*Editor’s Note: To read about Maya Sangesland’s award-winning work from last year, check out her Postbac Poster Award summary in our July 2014 issue.