Role of Snord123 in the translational regulation of gene expression in high endothelial venules – SNORD123-HEV

High endothelial venules (HEVs) are specialized blood vessels for lymphocyte trafficking found in lymph nodes and other lymphoid organs. HEVs play an essential role in the immune system because they are critical for immune surveillance and recirculation (homing) of lymphocytes throughout the different lymphoid organs of the body. Blood vessels related to HEVs are present in tissues undergoing chronic inflammation, as well as in solid tumors. A better understanding of HEV regulation could thus have important implications for immunity, inflammation and cancer.
It is therefore important to understand how HEVs work and what distinguishes them from conventional blood vessels. HEV endothelial cells (HECs) have a plump, almost cuboidal morphology, and exhibit the characteristics of metabolically active secretory-type cells, with a prominent Golgi complex, abundant mitochondria closely associated with rough endoplasmic reticulum, many ribosomes frequently found in polyribosomes clusters, and a large rounded nucleus with one or two nucleoli. We recently described differences between HECs and endothelial cells of conventional blood vessels using single-cell RNA-seq (Cell Rep 2019). Importantly, we identified several genes co-regulated or correlated with critical HEV genes, as candidate regulators of lymphocyte homing. Among these, the box C/D small nucleolar RNA Snord123 and the long non-coding RNA produced by its host gene Snhg18 are highly expressed in HECs compared to conventional endothelial cells. We showed that Snord123 guides a site-specific rRNA 2’-O-methylation in 28S rRNA and that deletion of Snord123 in mouse abrogates this site-specific rRNA modification, and results in an inhibition of HEV-mediated lymphocyte homing to lymph nodes. These reults are highly original and highlight a previously unsuspected involvement of rRNA chemically modified nucleotides in the physiology of HEVs.
The major objective of the current proposal is to investigate the role of Snord123 in the translational regulation of gene expression in HECs and the modulation of HEV function. Using various approaches, we will determine whether Snord123, and its associated rRNA 2’-O-methylation site, act by fine-tuning the global efficiency and fidelity of translation in HECs, or by modulating specifically the translation of a subset of HEV mRNAs required for lymphocyte trafficking. For that purpose, we will use multidisciplinary approaches : mouse KO and KI models, molecular and cellular biology, flow cytometry, immunohistofluorescence, high-throughput proteomics, RNA-seq, Ribo-seq, hybrid in vitro translation assays, intravital microscopy, in vivo homing assays.
Based on the strong preliminary results already obtained, we believe that the project will lead to an important scientific and conceptual advance, the role of a non-coding RNA (Snord123) in HEV mediated-lymphocyte trafficking to lymph nodes. Indeed, the project could provide the first genetic evidence for a critical role of site-specific rRNA methylation (guided by Snord123), ribosome heterogeneity and translational regulation, in the control of lymphocyte homing to lymph nodes, an important physiological process for immune surveillance.
The long-standing expertise of the two partners in the molecular and functional characterization of HEVs, and the functional analyses of snoRNAs, respectively, is a key asset for the project. We are thus in an ideal position to successfully pursue the current project.