Biology subjects

Lebedenko, C. G.

Publications and source records attributed to Lebedenko, C. G..

3 recordsLinked to original sources

bioRxiv2024

Cell surface ribonucleoproteins cluster with heparan sulfate to regulate growth factor signaling

Receptor-ligand interactions govern a wide array of biological pathways, facilitating a cells ability to interrogate and integrate information from the extracellular space. Here, using an unbiased genome-wide knockout screen, we identify heparan sulfate proteoglycans (HSPGs) as a major component in the organizational mechanism of cell surface glycoRNA and cell surface RNA binding proteins (csRBPs). Cleavage of mature heparan sulfate chains, knockout of N- and 6-O-sulfotransferases, overexpression of endo-6-O-sulfatases, or the addition of exogenous heparan sulfate chains with high 2-O sulfation result in marked loss in glycoRNA-csRBP clustering in U2OS cells. Functionally, we provide evidence that signal transduction by HS-dependent growth factors such as VEGF-A165 is regulated by cell surface RNAs, and in vitro VEGF-A165, selectively interacts with glycoRNAs. Our findings uncover a new molecular mechanism of controlling signal transduction of specific growth factors across the plasma membrane by the regulated assembly of glycoRNAs, csRBPs, and heparan sulfate clusters.

molecular biology↗

bioRxiv2023

The modified RNA base acp3U is an attachment site for N-glycans in glycoRNA

We recently identified glycoRNA--a previously undescribed glycoconjugate--which consists of RNAs modified with secretory N-glycans and presented on the cell surface. While previous work supported a covalent linkage between RNA and glycans, the direct chemical nature of the RNA-glycan connection was not described. Here we develop a sensitive and scalable protocol to detect and characterize native glycoRNAs. Leveraging periodate oxidation and aldehyde ligation (rPAL) and Sequential Window Acquisition of all Theoretical Mass Spectra (SWATH-MS), we identified the modified RNA base 3-(3-amino-3-carboxypropyl)uridine (acp3U) as a site of attachment of N-glycans in glycoRNA. The sensitivity and robustness of rPAL provided the first evidence of a direct glycan-RNA linkage, and its flexibility will enable further characterization of glycoRNA biology.

molecular biology↗

bioRxiv2023

Rapid and sensitive detection of native glycoRNAs

Chemical tools enable precise characterization of many biopolymers, including glycoconjugates. Metabolic chemical reporters enabled the discovery of glycoRNAs, however they have certain limitations due the requirement of having living cells to incorporate the modified sugar. Here we develop a periodate oxidation and aldehyde ligation method to detect and characterize native sialoglycoRNAs, termed rPAL. With optimized RNA biochemistry to enhance recovery and analysis of small RNAs, we show rPAL is at least an order of magnitude more sensitive than previous methods for detecting sialoglycoRNAs. These improvements allow rPAL to detect sialoglycoRNA from human clinical samples as demonstrated by defining the abundance and patterns of sialoglycoRNAs from sorted populations of peripheral blood mononuclear cells. The sensitivity, robustness, and flexibility of rPAL will allow greater access towards characterizing glycoRNA biology.

molecular biology↗