Abstract
The study of growth and possible metastasis in animal models of tumors would benefit from reliable cell labels for noninvasive whole-organism imaging techniques such as magnetic resonance imaging. Genetically encoded cell-tracking reporters have the advantage that they are contrast-selective for viable cells with intact protein expression machinery. Besides, these reporters do not suffer from dilution during cell division. Encapsulins, which are bacterial protein nanocompartments, can serve as genetically controlled labels for multimodal detection of cells. Such nanocompartments can host various guest molecules inside their lumen. These include, for example, fluorescent proteins or enzymes with ferroxidase activity leading to biomineralization of iron oxide inside the encapsulin nanoshell. The aim of this work was to implement heterologous expression of encapsulin systems from Quasibacillus thermotolerans using the fluorescent reporter protein mScarlet-I and ferroxidase IMEF in the human hepatocellular carcinoma cell line HepG2. The successful expression of self-assembled encapsulin nanocompartments with functional cargo proteins was confirmed by fluorescence microscopy and transmission electron microscopy. Also, coexpression of encapsulin nanoshells, ferroxidase cargo, and iron transporter led to an increase in T2-weighted contrast in magnetic resonance imaging of HepG2 cells. The results demonstrate that the encapsulin cargo system from Q. thermotolerans may be suitable for multimodal imaging of cancer cells and could contribute to further in vitro and in vivo studies.
Highlights
IntroductionMany advances in cancer treatment would come from a better understanding of tumor biology, the elucidation of carcinogenesis mechanisms in preclinical studies
We investigated the heterologous expression of encapsulin reporter genes from Quasibacillus thermotolerans in human cancer cells, starting with the coexpression of encapsulin nanocompartments with the fluorescent reporter cargo protein
HepG2 cells were transiently cotransfected with the DNA of QtEncFLAG nanoshell and
Summary
Many advances in cancer treatment would come from a better understanding of tumor biology, the elucidation of carcinogenesis mechanisms in preclinical studies. Sensitive molecular imaging of living cells could provide the means to study the formation and growth of metastases in a whole-body context in animal models [1,2,3]. The orthotopic transplantation approach is widely used to simulate, for example, cancer invasion and metastases when cancer cells interact with stromal components, including extracellular matrices, endothelial cells, fibroblasts, and various types of immune cells [4]. The primary method of live-cell imaging is direct labeling of cells with a probe or contrast agent before transplantation [5,6]. Quantum dots and fluorophores can be used for
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