| 1. |
- Kerr, A. G., et al.
(author)
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The long noncoding RNA ADIPINT regulates human adipocyte metabolism via pyruvate carboxylase
- 2022
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In: Nature Communications. - : Springer Nature. - 2041-1723. ; 13:1
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Journal article (peer-reviewed)abstract
- The pleiotropic function of long noncoding RNAs is well recognized, but their direct role in governing metabolic homeostasis is less understood. Here, we describe a human adipocyte-specific lncRNA, ADIPINT, that regulates pyruvate carboxylase, a pivotal enzyme in energy metabolism. We developed an approach, Targeted RNA-protein identification using Orthogonal Organic Phase Separation, which identifies that ADIPINT binds to pyruvate carboxylase and validated the interaction with electron microscopy. ADIPINT knockdown alters the interactome and decreases the abundance and enzymatic activity of pyruvate carboxylase in the mitochondria. Reduced ADIPINT or pyruvate carboxylase expression lowers adipocyte lipid synthesis, breakdown, and lipid content. In human white adipose tissue, ADIPINT expression is increased in obesity and linked to fat cell size, adipose insulin resistance, and pyruvate carboxylase activity. Thus, we identify ADIPINT as a regulator of lipid metabolism in human white adipocytes, which at least in part is mediated through its interaction with pyruvate carboxylase.
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| 2. |
- Wang, Zuoneng, 1991-, et al.
(author)
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Optimizing purification of the peripheral membrane protein FAM92A1 fused to a modified spidroin tag
- 2022
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In: Protein Expression and Purification. - : Elsevier. - 1046-5928 .- 1096-0279. ; 189, s. 105992-105992
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Journal article (other academic/artistic)abstract
- Cryo-electron microscopy has revolutionized structural biology. In particular structures of proteins at themembrane interface have been a major contribution of cryoEM. Yet, visualization and characterization of peripheralmembrane proteins remains challenging; mostly because there is no unified purification strategy forthese proteins. FAM92A1 is a novel peripheral membrane protein that binds to the mitochondrial inner membrane.There, FAM92A1 dimers bind to the membrane and play an essential role in regulating the mitochondrialultrastructure. Curiously, FAM92A1 has also an important function in ciliogenesis. FAM92A1 is part of themembrane bending Bin1/Amphiphsyin/RVS (BAR) domain protein family. Currently, there is no structure ofFAM92A1, mostly because FAM92A1 is unstable and insoluble at high concentrations, like many BAR domainproteins. Yet, pure and concentrated protein is a necessity for screening to generate samples suitable for structuredetermination. Here, we present an optimized purification and expression strategy for dimeric FAM92A1. To ourknowledge, we are the first to use the spidroin tag NT* to successfully purify a peripheral membrane protein. Ourresults show that NT* not only increases solubility but stabilizes FAM92A1 as a dimer. FAM92A1 fused to NT* isactive because it is able to efficiently bend membranes. Taken together, our strategy indicates that this is apossible avenue to express and purify other challenging BAR domain proteins.
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