Liang Feng - US grants

Alzheimer's disease is a debilitating and deadly disease that subverts our most basic mental functions including the ability to think and to recall memories and eventually to even care for our own basic bodily needs. It is not only the most common form of dementia but also the sixth leading cause of death in the United States. As the elderly population grows, the prevalence of Alzheimer's disease will increase dramatically, presenting a pressing need in medicine and in our society. Since its identification a century ago, considerable progress has been made in understanding the symptoms and risk factors of Alzheimer's disease. However, our understanding of its molecular mechanisms is still very limited, and as a result there are no disease-modifying interventions to prevent or even delay its progression. I propose to develop new tools and combine biochemical, biophysical, and bioengineering approaches to determine the molecular basis of Alzheimer's disease. Both the tools and the scientific results will then be used to develop new prototype therapeutic agents. The technology developed for this goal will also have far-reaching impact on elucidating mechanisms for a variety of proteins. 

? DESCRIPTION (provided by applicant): Sugar is a key source of energy for multicellular organisms, and its efflux across the membrane is critical to many physiological processes, including blood glucose maintenance and milk production. SWEET transporters (SWEETs) are novel membrane proteins that mediate sugar export. SWEETs are also prototypes of the large MtN3 membrane protein clan, which includes notable members such as mitochondrial pyruvate carrier, PQ-loop transporters and the KDEL receptor. Eukaryotic SWEETs, together with their half transporter bacterial homolog, SemiSWEET, are a unique model system to study the widely observed duplication-fusion in membrane protein evolution. Despite the importance of SWEETs in sugar utilization and MtN3 proteins in mitochondrial function, lysosomal amino acid homeostasis, and ER protein retention, we do not understand SWEET and MtN3 mechanisms at the molecular level. To overcome a major barrier to progress-the lack of a structural framework to guide our mechanistic understanding of transport-we have solved high-resolution structures of two SemiSWEET proteins, in two distinct conformational states: outward open and occluded. We will leverage this structural data to gain a detailed understanding of the structure and function of SWEETs and, more broadly, of MtN3s. Specific Aim 1: Elucidate the structural basis of sugar transport by SemiSWEET using X-ray crystallography and biophysical methods. These studies will help us understand the physical basis of sugar transport by SemiSWEETs. Specific Aim 2: Determine the first crystal structure of a eukaryotic SWEET. This structural information will provide a blueprint for the transport process and help elucidate the evolution of membrane transport proteins with internal symmetry. Specific Aim 3: Dissect the transport mechanism of SWEETs and MtN3s through functional studies. Fundamental aspects of SWEET-mediated transport will be elucidated and then extended to members of the MtN3 clan. OVERALL IMPACT: The proposed research will reveal the structural basis of sugar transport by SWEETs, elucidate their transport mechanism, and accelerate exploration of MtN3s as therapeutic targets to treat diabetes, cancer and lysosomal storage diseases.

Major advancements in the fields of electronics, photonics, electro-mechanical systems and wireless communication have enabled the development of compact wearable devices, with applications in diverse domains such as fitness, wellness and medicine. Despite their potential, existing wearable devices are only able to measure a few parameters (e.g., heart rate, breathing, temperature or blood pressure). In parallel to these efforts, nanotechnology is enabling the development of miniature sensors that can detect different types of human health events at the nanoscale with unprecedented accuracy. In-vivo nanosensing systems, which can operate inside the human body in real time, have been proposed as a way to provide faster and more accurate disease diagnosis over traditional technologies. Despite the potential of this technology, there are several limitations in the current systems, such as the cost and bulkiness of existing portable systems, which limit its real-world impact. The objective of this project is to develop a smart service system for advanced health monitoring and disease diagnosis based on wearable nano-biosensing networks. The system consists of three elements: 1) a nanoplasmonic biochip, to be implanted under the skin and designed to react to lung cancer biomarkers; 2) a wearable smart band, integrated by nanophotonic devices for excitation and measurement of the implant; and 3) a software platform to process the measured signals, extract the information, and formulate a diagnosis. This technology will significantly boost the applications of wearable devices, by providing the means to detect different types of diseases and, in particular, cancer. By partnering with two industry leaders and pioneers in the fields of solid-state electronics and advanced biomedical devices, this project is expected to enable cancer progression monitoring systems, with a broad societal impact. Importantly, integrating research and industry with education is a priority in this interdisciplinary effort, which will train the next generation of student scientists (6 doctoral students supported).

The project encompasses four intertwined research thrusts. The first thrust is focused on the development of the nanoplasmonic biosensing technology at the basis of this smart health system. This includes an implantable nanoplasmonic biochip composed of multiplexed sensor arrays for lung cancer detection from biomarkers in blood, as well as the optical nano-sources and nano-photodetectors needed to respectively excite and measure the biosensing signals through reflection, both integrated in a wearable device. The second thrust is focused on the development of the software algorithms to dynamically calibrate and operate the nano-sources, collect and post-process the measured signals at the nano-photodetectors by considering the intra-body wireless channel, extract the diagnose information and securely share the collected data with the healthcare provider. The third thrust is focused on the human factors that impact the design of the entire system, including the study of the impact and optimization of the nanoplasmonic biochip in biological tissues, the development of biochip regeneration techniques for continued operation of the implant, the investigation of the photothermal effects introduced by the nanophotonic excitation platform and the implant, and the processing and distribution of sensitive data related to the users' health. Finally, the fourth thrust will create an integrated testbed for the entire proposed system, involving in-vitro testing of the biochips with blood samples of lung cancer patients, ex-vivo testing with biochips implanted in tissue-equivalent phantoms with blood microcirculation networks, and testing in cadaver specimens.

The project is led by an interdisciplinary team of researchers at the University at Buffalo with participation of the Departments of Electrical Engineering, Chemical and Biomedical Engineering and Orthopedics. Two industry partners contribute and support the development of this project, Intel Labs (Hillsboro, Oregon, large business partner) and Garwood Medical Devices (Buffalo, NY, start-up partner). In addition, the Roswell Park Cancer Institute (Buffalo, NY), a cancer research and treatment center, serves as a broader context partner and consultant to the team.

PROJECT SUMMARY Mitochondrial calcium (Ca2+) uptake is central to many fundamental physiological processes. It stimulates ATP production during times of increased metabolic need and provides a Ca2+ sink to modulate Ca2+-mediated signaling locally within a cell. Mitochondrial Ca2+ concentrations also regulate apoptosis and dysregulation?? specifically, Ca2+ overload??is a hallmark of pathologies ranging from neuronal excitotoxicity to heart failure and some epilepsies to muscular dystrophies. Yet despite the importance of mitochondrial Ca2+ uptake in normal physiology and disease, the molecular machinery mediating this process is relatively recently identified and many fundamental questions remain to be answered. The main route of Ca2+ influx to mitochondria is a channel called mitochondrial calcium uniporter, which includes the ubiquitous pore-forming subunit MCU and, depending on the species, several regulatory subunits (termed ?uniplex? when in complex). This novel channel is highly selective for Ca2+, and its activity is tightly regulated by cytosolic Ca2+ concentration. My group recently determined a high-resolution crystal structure for a fungal MCU that defined a novel channel architecture and revealed a high-affinity Ca2+-binding site. Moreover, our cryo-EM structure of the human uniplex holocomplex revealed its architecture and hints at the mechanisms by which it is regulated. With these structures and the methods we developed, my lab is uniquely poised to embark on the mechanistic understanding of the mitochondrial calcium uniporter. Here, we propose to: 1) elucidate the structural and biophysical basis of ion selectivity, conduction and inhibition; 2) understand mechanisms of the channel gating and the long-range modulation; and 3) probe the molecular basis of Ca2+-dependent regulation of the uniplex. These results will give us much needed mechanistic insights into the activity and regulation of mitochondrial calcium uniporter, expanding our understanding of general principles of Ca2+ channels. In addition, they should provide a strong framework to aid the design of MCU inhibitors, which may represent promising treatments for diseases and pathologies marked by MCU dysregulation and mitochondrial Ca2+ overload. !