Posts classified under: Primary Faculty

Research Interests

Discovery and molecular mechanisms of novel ion transporters.
The Rao laboratory studies the role of novel ion transporters in human health and disease. One project focuses on the calcium signaling in breast cancer. We showed that an isoform of the secretory pathway Ca2+-ATPase, SPCA2, interacts with ion channels to drive tumor proliferation. We are currently investigating how downregulation of SPCA2 promotes epithelial to mesenchymal transition. A second project relates to the endosomal Na+/H+ exchangers NHE6 and NHE9 that are linked to autism, Christianson syndrome, ADHD and a growing list of neurodevelopmental and neurodegenerative disorders. We use a powerful PheWAS approach combined with model structure-driven evolutionary conservation analysis and functional screening of human variants to identify, evaluate and predict causality of autism-associated mutations. Loss of eNHE function results in hyperacidic endosomes, which increases amyloidogenic processing, and turnover of cell surface receptors and neurotransmitter transporters to impact neurotransmitter uptake and synaptic development.  On the other hand, overexpression of NHE9 in glioblastoma confers chemoradiation resistance due to “inside out” control of oncogenic signaling. Our lab also discovered NHA2, an unusual Na+/H+ exchanger that is implicated in essential hypertension and plays a role in salt handling in the kidney. We make extensive use of 3D/organoid cultures, confocal microscopy, live cell calcium and pH imaging, and phenotype complementation in yeast to investigate mammalian transport proteins.

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Research Interests

Osmolarity Sensing 

 Cell is composed of around 70% water with a plasma membrane also permeable to water. So keeping cell volume constant in response to osmotic challenges is fundamental to life. This is achieved in mammals by maintaining a stable blood plasma osmolarity (near 300 mOsm/L) and by possessing a variety of mechanisms that allow individual cells to monitor and recover their volume following osmotic swelling or shrinkage. Defective osmoregulation leads to various human disorders, including dehydration, hypertension, renal and neurological diseases. However, the identity of many key osmosensing molecules has been a long-standing mystery. Our goal is to elucidate the molecular mechanisms of mammalian osmotic regulation at both the cellular and whole body levels. We recently performed a genome-wide RNAi screen and co-discovered SWELL1 (LRRC8A) as an essential component of the elusive Volume-Regulated Anion Channel (VRAC). VRAC is required for maintaining cell volume in response to osmotic swelling. This discovery enables exciting studies elucidating the function of this important channel in cell volume regulation, fluid secretion, and diseases such as diabetes, stroke and traumatic brain injury.

 Deorphanizing the Human Transmembrane Genome: A Focus on Novel Ion Channels   

 The sequencing of the human genome has fueled the last two decades of work to functionally decipher genome content. An important subset (~25%) of genes encodes transmembrane proteins, which represent the targets of over half of known drugs. Despite recent progress, a large number (~1,500) of membrane proteins are still functionally uncharacterized. We focus on deorphanizing a particularly interesting functional class of membrane proteins, i.e. ion channels or transporters, many of which are well characterized biophysically yet lack underlying molecular identity. Toward this end, we are combining the powerful genomics tools (including bioinformatics, proteomics, single-cell RNA sequencing, and RNAi/CRISPR gene manipulation) with electrophysiology and imaging techniques. Our study will shed light on the molecular identity and physiological function of new pore-forming membrane proteins and may provide therapeutic strategies to target them for diseases with abnormal ion transport and homeostasis. 

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Research Interests

Copper homeostasis: mechanism of transport, protein and metabolic networks, integrative approaches to human disease

Copper plays an essential role in human physiology. It is an essential cofactor of enzymes that are required for respiration, biosynthesis of neurotransmitter, detoxification of radicals, blood clotting, connective tissue formation, and many others processes. Recent data suggest that coppers serves as a signaling molecules and regulates myelination of neurons, inflammatory response, and angiogenesis. Copper also alters the sensitivity of cancer cells to chemotherapeutic drugs, such as cisplatin.  The long-term goal of our research is provide a detailed understanding of human copper homeostasis in health and disease.

Structure, Function, and Regulation of Human Copper Transporters

In human cells, copper levels are controlled by two copper-transporting ATPases, ATP7A and ATP7B. Genetic mutations in ATP7A result in a systemic copper deficiency, particularly in the CNS, and Menkes disease. Menkes disease has a profound effect on human growth and development, and is mostly fatal. Mutations in another copper transporter, ATP7B, are associated with copper accumulation in tissues, particularly in the liver, and Wilson disease. Wilson disease affects primarily children and young adults and has hepatic, neurologic and psychiatric manifestations.

Our group studies the function and regulation of ATP7A and ATP7B in different cell types. We use a combination of biochemical, biophysical, and cell biological approaches to understand the mechanism of copper transport and effects of mutations on the structure and activity of copper transporters. The intracellular localization of ATP7A and ATP7B is regulated by changes in copper levels and hormones. By using site-directed mutagenesis and regulated expression of copper transporters in mammalian cells, we investigate how ATP7A and ATP7B sense changing copper levels and adjust their intracellular location and activity. Through a collaborative whole-genome screening we identified novel regulators of copper homeostasis and we are investigating their role in activity, stability, and trafficking of copper transporters.

Copper chaperones

The intracellular concentration of free copper is tightly controlled. In a cytosol, copper is carried around by small shuttle proteins called copper chaperones or metallochaperones. These proteins distribute copper to different cellular destinations (cytosol, mitochondria, secretory pathway) and modulate activity of their target enzymes. We have discovered that the copper-chaperone Atox1 is a redox sensitive molecule that serves as an important switch directing more copper towards the secretory pathway during neuronal differentiation.  We have developed a mouse model for targeted deletion of Atox1 and plan to better understand the role of Atox1 in copper metabolism in the brain

Molecular Mechanisms of Wilson disease

Copper misbalance has severe consequences for cell physiology. We are using the genetically engineered mice with a global and tissue specific deletions of ATP7B to better understand (i) the molecular and cellular events that trigger the onset of pathology in different tissues, (ii) the mechanism of disease progression, and (iii) the role of various metabolic pathways in specific disease manifestations. Our goal is to generate a comprehensive and predictive model for Wilson disease. To achieve this goal, we investigate the copper-induced changes at a single cell level and combine these studies with a large-scale analysis of a proteome, metallome and transcriptomes. We have identified lipid metabolism and liver nuclear receptors as important players in disease pathogenesis and are developing new therapeutic approaches for Wilson disease

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