Doctor of Philosophy, Yale University (2018)
Bachelor of Arts, The University of Chicago, Biological Sciences (2012)
Mark Krasnow, Postdoctoral Faculty Sponsor
Serum proteome analysis of systemic JIA and related lung disease identifies distinct inflammatory programs and biomarkers.
Arthritis & rheumatology (Hoboken, N.J.)
OBJECTIVES: Recent observations in systemic Juvenile Idiopathic Arthritis (sJIA) suggest an increasing incidence of high-mortality interstitial lung disease (sJIA-LD) often characterized by a variant of pulmonary alveolar proteinosis (PAP). Co-occurrence of macrophage activation syndrome (MAS) and PAP in sJIA suggested a shared pathology, but sJIA-LD patients also commonly experience features of drug reaction such as atypical rashes and eosinophilia. We sought to investigate immunopathology and identify biomarkers in sJIA, MAS, and sJIA-LD.METHODS: We used SOMAscan to measure >1300 analytes in sera from healthy controls and patients with sJIA, MAS, sJIA-LD and other related diseases. We verified selected findings by ELISA and lung immunostaining. Because the proteome of a sample may reflect multiple states (sJIA, MAS, sJIA-LD), we used regression modeling to identify subsets of altered proteins associated with each state. We tested key findings in a validation cohort.RESULTS: Proteome alterations in active sJIA and MAS overlapped substantially, including known sJIA biomarkers like SAA and S100A9, and novel elevations of heat shock proteins and glycolytic enzymes. IL-18 was elevated in all sJIA groups, particularly MAS and sJIA-LD. We also identified an MAS-independent sJIA-LD signature notable for elevated ICAM5, MMP7, and allergic/eosinophilic chemokines, which have been previously associated with lung damage. Immunohistochemistry localized ICAM5 and MMP7 in sJIA-LD lung. ICAM5's ability to distinguish sJIA-LD from sJIA/MAS was independently validated.CONCLUSION: Serum proteins support an sJIA-to-MAS continuum, help distinguish sJIA, sJIA/MAS, and sJIA-LD and suggest etiologic hypotheses. Select biomarkers, such as ICAM5, could aid in early detection and management of sJIA-LD.
View details for DOI 10.1002/art.42099
View details for PubMedID 35189047
Cell types of origin of the cell-free transcriptome.
Cell-free RNA from liquid biopsies can be analyzed to determine disease tissue of origin. We extend this concept to identify cell types of origin using the Tabula Sapiens transcriptomic cell atlas as well as individual tissue transcriptomic cell atlases in combination with the Human Protein Atlas RNA consensus dataset. We define cell type signature scores, which allow the inference of cell types that contribute to cell-free RNA for a variety of diseases.
View details for DOI 10.1038/s41587-021-01188-9
View details for PubMedID 35132263
RNA splicing programs define tissue compartments and cell types at single cell resolution.
The extent splicing is regulated at single-cell resolution has remained controversial due to both available data and methods to interpret it. We apply the SpliZ, a new statistical approach, to detect cell-type-specific splicing in >110K cells from 12 human tissues. Using 10x data for discovery, 9.1% of genes with computable SpliZ scores are cell-type-specifically spliced, including ubiquitously expressed genes MYL6 and RPS24. These results are validated with RNA FISH, single-cell PCR, and Smart-seq2. SpliZ analysis reveals 170 genes with regulated splicing during human spermatogenesis, including examples conserved in mouse and mouse lemur. The SpliZ allows model-based identification of subpopulations indistinguishable based on gene expression, illustrated by subpopulation-specific splicing of classical monocytes involving an ultraconserved exon in SAT1. Together, this analysis of differential splicing across multiple organs establishes that splicing is regulated cell-type-specifically.
View details for DOI 10.7554/eLife.70692
View details for PubMedID 34515025
Phosphofructokinase Relocalizes into Subcellular Compartments with Liquid-like Properties In Vivo.
Although much is known about the biochemical regulation of glycolytic enzymes, less is understood about how they are organized inside cells. We systematically examine the dynamic subcellular localization of glycolytic protein phosphofructokinase-1/PFK-1.1 in Caenorhabditis elegans. We determine that endogenous PFK-1.1 localizes to subcellular compartments in vivo. In neurons, PFK-1.1 forms phase-separated condensates near synapses in response to energy stress from transient hypoxia. Restoring animals to normoxic conditions results in cytosolic dispersion of PFK-1.1. PFK-1.1 condensates exhibit liquid-like properties, including spheroid shapes due to surface tension, fluidity due to deformations, and fast internal molecular rearrangements. Heterologous self-association domain cryptochrome 2 promotes formation of PFK-1.1 condensates and recruitment of aldolase/ALDO-1. PFK-1.1 condensates do not correspond to stress granules and might represent novel metabolic subcompartments. Our studies indicate that glycolytic protein PFK-1.1 can dynamically form condensates in vivo.
View details for DOI 10.1016/j.bpj.2020.08.002
View details for PubMedID 32853565
Activity and structure of human acetyl-CoA carboxylase targeted by a specific inhibitor.
2018; 592 (12): 2048-2058
We have studied a series of human acetyl-CoA carboxylase (ACC) 1 and ACC2 proteins with deletions and/or Ser to Ala substitutions of the known phosphorylation sites. In vitro dephosphorylation/phosphorylation experiments reveal a substantial level of phosphorylation of human ACCs produced in insect cells. Our results are consistent with AMPK phosphorylation of Ser29 , Ser80 , Ser1,201 , and Ser1,216 . Phosphorylation of the N-terminal regulatory domain decreases ACC1 activity, while phosphorylation of residues in the ACC central domain has no effect. Inhibition of the activity by phosphorylation is significantly more profound at citrate concentrations below 2 mm. Furthermore, deletion of the N-terminal domain facilitates structural changes induced by citrate, including conversion of ACC dimers to linear polymers. We have also identified ACC2 amino acid mutations affecting specific inhibition of the isozyme by compound CD-017-0191. They form two clusters separated by 60-90 Å: one located in the vicinity of the BC active site and the other one in the vicinity of the ACC1 phosphorylation sites in the central domain, suggesting a contribution of the interface of two ACC dimers in the polymer to the inhibitor binding site.
View details for DOI 10.1002/1873-3468.13097
View details for PubMedID 29772612
Glycolytic Enzymes Localize to Synapses under Energy Stress to Support Synaptic Function.
2016; 90 (2): 278-91
Changes in neuronal activity create local and transient changes in energy demands at synapses. Here we discover a metabolic compartment that forms in vivo near synapses to meet local energy demands and support synaptic function in Caenorhabditis elegans neurons. Under conditions of energy stress, glycolytic enzymes redistribute from a diffuse localization in the cytoplasm to a punctate localization adjacent to synapses. Glycolytic enzymes colocalize, suggesting the ad hoc formation of a glycolysis compartment, or a "glycolytic metabolon," that can maintain local levels of ATP. Local formation of the glycolytic metabolon is dependent on presynaptic scaffolding proteins, and disruption of the glycolytic metabolon blocks the synaptic vesicle cycle, impairs synaptic recovery, and affects locomotion. Our studies indicate that under energy stress conditions, energy demands in C. elegans synapses are met locally through the assembly of a glycolytic metabolon to sustain synaptic function and behavior. VIDEO ABSTRACT.
View details for DOI 10.1016/j.neuron.2016.03.011
View details for PubMedID 27068791
View details for PubMedCentralID PMC4840048
Resistance to herbicides caused by single amino acid mutations in acetyl-CoA carboxylase in resistant populations of grassy weeds.
The New phytologist
2013; 197 (4): 1110-6
Eleven spontaneous mutations of acetyl-CoA carboxylase have been identified in many herbicide-resistant populations of 42 species of grassy weeds, hampering application of aryloxyphenoxypropionate, cyclohexadione and phenylpyrazoline herbicides in agriculture. IC(50) shifts (resistance indices) caused by herbicide-resistant mutations were determined using a recombinant yeast system that allows comparison of the effects of single amino acid mutations in the same biochemical background, avoiding the complexity inherent in the in planta experiments. The effect of six mutations on the sensitivity of acetyl-CoA carboxylase to nine herbicides representing the three chemical classes was studied. A combination of partially overlapping binding sites of the three classes of herbicides and the structure of their variable parts explains cross-resistance among and between the three classes of inhibitors, as well as differences in their specificity. Some degree of resistance was detected for 51 of 54 herbicide/mutation combinations. Introduction of new herbicides targeting acetyl-CoA carboxylase will depend on their ability to overcome the high degree of cross-resistance already existing in weed populations.
View details for DOI 10.1111/nph.12117
View details for PubMedID 23301879