(This page is under active update.)
Challenge Our laboratory is interested in understanding the role of lipids in physiology and pathology. The cellular lipidome contains a wide spectrum of structurally distinct lipids and collectively plays important roles in every aspect of cellular functions including membrane compartmentalization, signal transduction and energy homeostasis. Dysregulation of lipid metabolism is implicated in a large number of human diseases including obesity, neurodegeneration, cancer, and various organ failure.
While most cellular lipids contain multiple chemically reactive groups hence simultaneously belong to multiple lipid classes, there is a major gap in our understanding about what chemical properties of each lipid are contributing to a specific cellular activity and how. As a result, our knowledge about how cells organize and regulate their lipidome in response to developmental and environmental cues, and how the lipidome in turn supports cell type-specific structure and function remains scarce. Addressing these critical questions will help establish the missing link between the cellular lipidome and the genetic information flow (DNA/RNA/protein), and accelerate our therapeutic development against lipid metabolism-related diseases.
To gain insights on these questions, we are actively pursuing the following research directions:
Understanding and targeting ferroptosis in human diseases
All cells in our body are surrounded by a membrane that is mainly made of lipid molecules. Similar membranes also surround many of the organelles inside the cells. It is important for life that these membranes are impermeable to external molecules; but the membranes also need to be flexible and fluidic enough to permit cell division and membrane fission and fusion. These features are largely achieved by introducing unsaturated fatty acids to the membrane. According to the number of double bonds, fatty acids can be grouped into saturated, mono- or poly-unsaturated fatty acids. As the unsaturation level increases, these fatty acids display decreasing melting temperature and packing density, and increasing curvature and fluidity.
At the cellular level, the introduction of PUFA lipids into cellular membranes is very significant in evolution, because it is very effective in enabling motility, vesicle fission and fusion, and the formation of bioactive lipid signaling molecules. However, there is a major peril of including polyunsaturated or PUFA lipids in membranes, which is that the multiple double bonds organized in these pentadienyl structures make PUFA-lipids particularly susceptible to hydrogen abstraction and lipid peroxidation. One reason for lipid peroxidation to be particularly concerning is that in the presence of oxygen and reactive iron, lipid peroxyl radicals can self-propagate and leads to a burst in lipid radical species, which damage cellular membranes and proteins and cause ferroptosis.
At the organism level, lipid peroxidation contributes to various degenerative diseases in the kidney, liver and brain, as well as to the normal aging process. In order to balance this ferroptotic risk, eukaryotic cells have evolved a counter enzymatic system centered on glutathione peroxidase 4 or GPX4. GPX4 is a selenoprotein and is the only cellular enzyme that utilizes glutathione to reduce lipid hydroperoxides to non-toxic lipid alcohols to keep cells alive. Artificial inhibition of GPX4 can also lead to lipid hydroperoxide accumulation and ferroptosis. Therefore, our cells need to keep a fine balance when utilizing PUFA-lipids to avoid aberrant peroxidation and ferroptosis.
In the past few years, the field has made rapid progress in understanding the biochemical basis of ferroptosis, developing chemical tools for reporting this cell death pathway, and identifying ferroptosis-relevant disease contexts. However, key questions remain before the power of ferroptosis-targeting strategies can be utilized to treat human diseases:
1. What is the chemical basis of lipid peroxidation and what is its physiological role in normal tissues?
2. What are the death-executing molecules or events caused by lipid peroxidation, and what types of membrane damage are necessary or sufficient to cause cell death?
3. What are the genes and pathways involved in executing lipid peroxidation? Can we target such proteins to control cellular lipid peroxidation levels?
4. What are the physiological roles of ferroptotic cell death in early development, adulthood and aging, respectively?
Constructing a Mammalian Lipidome Atlas to understand lineage-specific roles of the lipidome
Our prior research highlighted that cellular differentiation, tumor progression and acquisition of therapy resistance are coupled with significant changes in the cellular lipid composition, making a
compelling case that the cellular lipidome exhibits a high degree of plasticity and is highly responsive to cell state transitions. Nonetheless, few cell type-specific lipid pathways have been identified despite the vast diversity of the chemical structures of cellular lipids. To accelerate the discovery of such lipids, we are developing a comprehensive “Mammalian Lipidome Atlas” (MLA) in major human and mouse cell types at representative developmental and aging stages. I envision that more novel functional lipid classes and pathways, like plasmalogens, triacylglycerides and cholesterols, will emerge from this analysis. To collaborate on this work, please contact the PI directly.
Decoding the contribution of lipid metabolism in tumorigenesis and metastasis
The development of metastatic disease accounts for >90% of cancer-associated mortality, yet few biological pathways can be exploited to inhibit metastasis. While previous studies have largely focused on the signaling and cytoskeleton changes required for metastasis such as epithelial-mesenchymal transitions (EMT), the membrane compositional changes required for supporting the enhanced motility and invasiveness of metastatic cells have not been fully examined. We are using mouse models and human patient tumor samples to investigate the molecular basis of cancer metastasis formation. We envision these studies will illuminate the membrane configurations required for supporting the enhanced motility and invasiveness of metastatic cells – an understudied aspect in metastasis research.
Developing novel lipid-detection technologies and lipid-metabolism targeting therapeutics
To empower efficient lipid characterizations, my lab will be actively engaged in developing novel probes and techniques for tracing, visualizing and quantifying specific lipids in vitro and in vivo. Moreover, other research areas in the lab will likely nominate novel metabolic pathways for therapeutic intervention in disease settings. Once an attractive target pathway is identified, we will integrate various chemical biology approaches, including high-throughput phenotypic screening, targeted protein degradation, and DNA-encoded small molecule screening, to develop therapeutic modalities. In addition to small molecule inhibitors, our lab is also interested in other therapeutic modalities including gene therapy and bioactive peptides. Currently, we are focusing on developing small molecule activators and inhibitors of ferroptosis for in vitro and in vivo use.
Dr. Zou's postdoc research in the Schreiber lab at the Broad Institute focused on identifying targetable vulnerabilities associated with cancer metabolism. He first explored the role of aberrant lipid metabolism in clear-cell carcinomas from the kidney and ovary, which together kills >15,000 patients annually in the US with the incidences on the rise. Using chemical profiling, he discovered a targetable dependency on glutathione peroxidase 4 (GPX4) in clear-cell cancers (Zou et al., Nature Communications 2019). GPX4 is a gate-keeper to protect cells from undergoing lipid peroxidation-induced ferroptosis, an iron-dependent non-apoptotic cell death program. He further demonstrated that this unique vulnerability is driven by the HIF-2ɑ pathway, doing so by selectively upregulating polyunsaturated lipids – the key substrates of lipid peroxidation and ferroptosis in clear-cell carcinomas. Using functional genomic screening and lipidomics profiling, he identified the hypoxia-induced, lipid droplet-associated protein (HILPDA), a poorly-characterized HIF-2ɑ target gene, as the executioner of HIF-2ɑ’s activity in selectively enriching polyunsaturated lipids and promoting ferroptosis. Together, this work depicts an “oncogene-induced ferroptosis sensitivity” in cancer, and suggests a potential therapeutic opportunity for the extremely lethal clear-cell cancers. Moreover, his findings place the HIF-2ɑ-HILPDA axis as one of the first transcriptional axis known to be dedicated to upregulate polyunsaturated lipids rather than saturated or monounsaturated fatty acids, advancing our understanding about selective regulation of polyunsaturated lipids. This work will fill a critical gap between cancer genetics and PUFA metabolism; and provide much-needed insights about the origin of ferroptosis sensitivity.
Using clear-cell carcinomas as models of intrinsic hypersensitivity to ferroptosis and genome-wide CRISPR screens as discovery tools, Dr. Zou identified the Cytochrome P450 Oxidoreductase (POR) enzyme as a necessary executioner of ferroptosis in various cancer lineages. Using cellular assays and lipidomic profiling, he found that POR specifically promotes lipid peroxidation by catalyzing the reduction of Fe (II) from Fe (III). This finding fills a major gap in our understanding about the biochemical nature of ferroptosis, and reveals a key regulatory node for modulating ferroptosis in cells (Zou*,#, Li* et al., Nature Chemical Biolology 2020). This finding enables specific pharmacological intervention of the ferroptosis pathway.
Recently, Dr. Zou and collaborators used genome-wide CRISPR-Cas9 suppressor screens to identify the oxidative organelles peroxisomes as critical contributors to ferroptosis sensitivity in human renal and ovarian carcinoma cells (Zou*,#, Henry*, Ricq* et al., Nature 2020). Using lipidomic profiling, the team showed that peroxisomes contribute to ferroptosis by synthesizing polyunsaturated ether phospholipids (PUFA-ePLs), which act as substrates for lipid peroxidation that, in turn, results in the induction of ferroptosis. Carcinoma cells that are initially sensitive to ferroptosis can switch to a ferroptosis-resistant state in vivo in mice, which is associated with extensive downregulation of PUFA-ePLs. The team further found that the pro-ferroptotic role of PUFA-ePLs can be extended beyond neoplastic cells to other cell types, including neurons and cardiomyocytes. Together, this work revealed roles for the peroxisome-ether-phospholipid axis in driving susceptibility to and evasion from ferroptosis, highlighted PUFA-ePL as a distinct functional lipid class that is dynamically regulated during cell-state transitions, and suggested multiple regulatory nodes for therapeutic interventions in diseases that involve ferroptosis.
Dr. Zou obtained his Ph.D. in Dr. Joan Massagué’s group at Memorial Sloan Kettering Cancer Center. In his graduate studies, he focused on characterizing the transcriptional regulation mechanisms of cell fate transitions during stem cell differentiation and cancer metastasis. He identified a set of lineage-restricted enhancers that act as signal-integration platforms. He then used gene-editing tools to depict a base-pair resolution map of transcription factor binding sites at these enhancers. Furthermore, we discovered that the TGF-β pathway diverts from pluripotency to differentiation roles by switching from interacting with pluripotency factors to with Wnt/Tcf, a process controlled by p53-mediated Wnt3 production (Wang*, Zou* et al., Cell Stem Cell 2017). In addition, he discovered that a unique exon in Smad2 underlies the strict requirement of Smad2, but not Smad3, in vertebrate development (Aragón*, Wang*, Zou* et al., Genes&Dev 2019).
Technology and Software Development
Along with Dr. Zou's deep interest in molecular biology and disease mechanisms, he has a long-term interest in developing technologies that enable better understanding of cell states in vitro and in vivo. In his postdoc,
He co-developed the photochemical activation of lipid peroxidation (PALP) technique, which enables non-invasive visualization and quantification of polyunsaturated lipids in live cells (Zou*, Graham*, et al., bioRxiv 2020). This method also allows rapid assessment of cellular sensitivity to ferroptosis.
Together with Paul A. Clemons’ (Broad Institute) group, Dr. Zou co-developed a computational package named Gene-List Network Enrichment Analysis (GeLiNEA) for identifying enriched pathways in a given gene list (Zou*, Henry*, Ricq* et al., in press). This method integrates protein-protein (gene-gene) interaction information into the widely used Gene Set Enrichment Analysis (GSEA) method (Subramanian et al., PNAS 2005), and is more powerful than GSEA for pathway discovery.
In his PhD, Dr. Zou optimized and extended the translating ribosome affinity purification and sequencing (TRAP-Seq) method for interrogating the behaviors of metastatic cancer cells in vivo, and established a computational pipeline for its data analysis. TRAP-Seq bypasses the need to isolate cells of interest and can characterize gene expression of cells in situ. Key findings enabled by TRAP-Seq include:
Identification of the role of the PI3K pathway in melanoma and lung tumor relapse stimulated by therapy-induced tumor debulking (Obenauf, Zou et al., Nature 2015).
Identification of the Src pathway as the downstream mediator of mesenchymal stem cell-stimulated pro-bone metastasis effect in triple-negative breast cancers (Zhang, Jin, Malladi, Zou et al., Cell 2013).
Identification of the Complement component 3 (C3) factor as a driver of cancer metastasis to the leptomeningeal space (Boire, Zou et al., Cell 2017).
Identification of the YAP pathway as a mediator of the pro-tumor-initiating role of L1CAM, a potent mediator of metastasis in multiple cancers (Er, Valiente, Ganesh, Zou, et al., Nature Cell Biol 2017).