Microbes are key players in global carbon and nitrogen cycles that fuel the biosphere and modulate atmospheric composition. The ‘household’ economy of energy budgeting in a microbial cell strongly affects its fitness, and thus its rise and fall in ecosystems. However, cellular energy intake and spending cannot be quantified by current approaches.

I propose to develop a novel experimental framework to dissect the energy budgeting strategies of different microbes based on their growth-dependent heat dissipation. I will first use calorimetry and isotope labeling to respectively measure heat and anabolic rates of a laboratory strain for establishing a strategy to extract energy usage for maintenance and growth. I will apply this approach to studying environmentally-relevant microbes with different anabolic capacities from diverse geographical origins, including archaea and bacteria that produce and consume methane (a potent green-house gas) and that fixes atmospheric nitrogen gas. Furthermore, I will develop a novel microfluidics-based calorimeter and combine it with nanoscale secondary ion mass spectrometry to enable parallel, long-term, single-cell analyses on microbial metabolism in environments that differ in nutrient abundance, nutrient composition, temperature, and acidity.

The goal is to unveil: (i) the fitness tradeoffs between microbial assimilations of different carbon, nitrogen, and energy sources, and (ii) different energy budgeting strategies that govern bacterial and archaean adaptation. The obtained knowledge will advance our prediction on how microbe-based natural carbon and nitrogen cycling accommodates to manmade global geochemical and climate changes.

In Hh signaling pathway, activity of Smo, a seven-transmembrane (7-TM) protein belonging to G-protein coupled receptor (GPCR) family, is repressed by Patched (Ptch). After Hh binds to Ptch, the repression of Smo is relieved and the derepression consequently results in the activation of transcription factor Ci/Gli and then transcription of Hh target genes related to embryo development and postnatal tissue homeostasis. Smo plays a central role and mediates almost all biological activities of Hedgehog proteins. Dysfunction of Smo can lead to phenotypes ranging from severe malformations of the head and limbs to embryonic lethality, and even various tumors. The molecular mechanisms of Smo activation and its interactions with the downstream effector are still not fully understood, albeit with the available structures of full-length Smo and isolated domains bound to the respective small-molecule ligands. Compelling evidences suggest coupling of Smo to G protein during activation, specifically Gi protein. Inspired by recent technological breakthrough in single particle cryo-electron microscopy (cryo-EM), I propose to determine the structures of Smo-Gi protein complex by single particle cryo-EM. I will use conformational specific fragment antigen binding (Fabs) as fiducial marks to facilitate cryo-EM image analysis. Furthermore, I will use lipid nanodisc or salipro system to stabilize the Smo-Gi complex and aim to analyze the roles of lipids on Smo activation. The ultimate goal of this study is to facilitate structure-based drug design with better treatments for Smo-related cancers.

Cancer is not a single disease and individual tumors have different molecular profiles. Glioblastoma multiforme (GBM), the most aggressive form of glioma, can be grouped into four different molecular subtypes including proneural, neural, mesenchymal and classical, based on distinct gene expression signatures. GBM has an invariably terminal prognosis. Most patients die within 14 months following diagnosis, even with surgery, radiation and chemotherapy, stressing the urgency for new therapies. Non-cancerous cells in the tumor microenvironment (TME), including different populations of immune cells, fibroblasts and endothelial cells, represent genetically stable therapeutic targets since they are less likely to develop acquired resistance as a result of genetic changes, in contrast to tumor cells. One immune cell type that has been successfully targeted in preclinical trials of proneural GBM are tumor-associated recruited macrophages and resident microglia (TAMs). However, it remains to be determined whether targeting TAMs also results in clinical responses in other GBM subtypes with different genetic alterations. As a consequence of a distinct molecular profile it may be expected that different populations and/or amounts of stromal cells are recruited and differentially educated in the TME. Therefore, I will determine how specific genetic alterations in different GBM subtypes sculpt the TME. I also aim to assess the effect of targeting TAMs in these GBM subtypes and to gain insights into potential resistance mechanisms. To achieve this, I will employ the latest genetic mouse models for GBM, extensive flow cytometry analyses, RNA sequencing and intravital microscopy.