GlycoRNA

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GlycoRNAs are small non-coding RNAs with sialylated glycans.

Contents

Glycans mediate inter- and intramolecular interactions by adding polysaccharide chains onto lipids and proteins. Similar to these other macromolecules, RNAs can undergo sialylation and bear glycan structures. Some examples include small nuclear (sn) RNAs, ribosomal (r) RNAs, small nucleolar (sno) RNAs, transfer (t) RNAs, and Y RNAs - the latter of which comprise the greatest percentage of glycosylated RNA species. [1]

Found primarily on the cell surface, these glycoRNAs can participate in the immune system and cell-to-cell communication. [2]

History of discovery

GlycoRNAs were discovered in May 2021 by a Stanford University research team led by Chemical Biologist Dr. Carolyn Bertozzi. The discovery came from an experiment where Post-doc Ryan Flynn used a metabolic tagging technique to label precursor sugars of glycan. What he discovered in the process was glycosylated, cell membrane-bound RNA.

Until now, lipids and proteins were the only kinds of similarly sugar-decorated macromolecules previously understood by science. Although glycoRNAs are not rare, it was understood that their discovery happened only relatively recently, as their presence defied accepted principles of well-established cellular biology. [2]

Biology of RNA glycolysation

The exact mechanism of GlycoRNA glycosylation is still unclear. It remains ambiguous whether it follows a similar process performed by glycolipids and glycoproteins in the ER-Golgi complex or through different machinery. Flynn and Bertozzi have offered the first description of glycoRNA, and suggest that RNAs are glycosylated using the same machinery and mechanism that produces N- and O-linked glycans in glycosylated proteins.

Previous experiments have not addressed where and how RNA is tagged with sugars in cells. Proteins and lipids acquire carbohydrate tags primarily in the endoplasmic reticulum and Golgi apparatus. Principles of cellular biology say the initial transfer of sugars to macromolecules occurs in the endoplasmic reticulum. Then, the subsequent addition of sugars in the Golgi results in the formation of mature glycan structures. In the Golgi, these newly made glycans are packaged into vesicles for transportation into organelles or secreted across the plasma membrane. However, RNA is not known to exist in these compartments. [2]

Proposed biosynthesis pathway of N-linked glycoRNA: The synthesis of glycans starts in the endoplasmic reticulum, continues in the Golgi before being transported to the plasma membrane, where the glycoRNA is either secreted or becomes embedded in the plasma membrane. This is very similar to the biosynthesis pathway of N-linked glycoproteins. Glycans and RNA.png
Proposed biosynthesis pathway of N-linked glycoRNA: The synthesis of glycans starts in the endoplasmic reticulum, continues in the Golgi before being transported to the plasma membrane, where the glycoRNA is either secreted or becomes embedded in the plasma membrane. This is very similar to the biosynthesis pathway of N-linked glycoproteins.

Using pharmacological and genetic inhibition approaches, Flynn and Bertozzi explored the effects of inhibiting key glycan biosynthetic machinery on the production of glycoRNA. They found that the production of glycoRNA is impaired in cells where the glycan biosynthetic machinery has been genetically altered, although adding exogenous glycan reverses the inhibition. [1] In another study, inhibition of oligosaccharyltransferase, a membrane protein that catalyzes the addition of glycans to asparagine residues, was also seen to diminish the production of glycoRNA. Flynn and Bertozzi's studies demonstrated that cellular glycoRNA is also produced by the glycan biosynthetic machinery. [2]

Some direct measurement of the glycan on RNA association would help disclose both the mechanism and its function. There are a few possible explanations for how the attachment proceeds. Bertozzi's lab conducted several experiments to understand what holds glycan RNA together. Running glycoRNAs through a sucrose gradient suggested that the molecule connecting the two entities was no larger than the small RNA itself. Researchers first thought that RNA could bind non-covalently to each other - a bond strong enough to avoid degradation unless digested by an RNase. However, this was very unlikely, as all evidence points to the RNA-glycan linkage via a covalent network. [2]

Another possibility is that glycans can bind proteins, which in turn bind RNA. Researchers have speculated that the RNA-glycan linkage is likely not due to the direct glycosylation of RNA bases. The sensitivity of glycoRNA moieties to PNGase F, which cleaves the glycosidic linkage between asparagine and the proximal GlcNAc of N-glycans, implies a non-nucleobase interaction. [2] However, a precursor nucleobase modification may be necessary for the glycosidic linkage to occur. The chemistry behind RNA glycosylation remains unknown and necessitates further research to define the chemical and structural features of the linkage.

Relations to disease

PYMOL: Three-dimensional structure of PNGase F, a glycosylasparaginase from Flavobacterium meningosepticum. (PDB: 1PGS) PDB 1pgs EBI.jpg
PYMOL: Three-dimensional structure of PNGase F, a glycosylasparaginase from Flavobacterium meningosepticum. (PDB: 1PGS)

The implication of glycoRNAs in RNA biology research is immense, especially when relating to disease progression and drug therapy. Aberrant glycosylation is a well-recognized hallmark of various human diseases. It has been a popular belief that diseases caused by abnormal glycosylation of proteins and lipids contribute to the pathology of those diseases. The discovery of glycoRNAs points researchers in the direction of an alternative molecule that may be responsible for the disrupted glycosylation network in human diseases.

Fractionation and immunohistochemical imaging studies revealed that glycoRNAs are primarily associated with the cell surface, as evidenced by their loss from the cell surface after treatment with a sialic acid-cleaving enzyme. [3] This suggests that they may be involved in extracellular molecular interactions. Cell surface glycoRNAs were found to interact with members of the sialic acid-binding immunoglobulin-like lectin (Siglec) receptor family in addition to binding an anti-RNA antibody used to detect RNA virus-infected cells. These receptors are known to play a role in immune response modulation, specifically host-pathogen interactions, immune evasion in cancer, and genetic links to autoimmune illnesses. The Siglec receptor family ligand partners are largely unknown, at least in part because previous research assumed they bind glycoproteins or glycolipids. As a result, glycoRNAs may be potential ligands for these and other orphan glycan-binding receptors. [4]

Autoimmune disorders are linked to a myriad of RNA and RNA-associated autoantigens. Flynn and Bertozzi employed soluble Siglec-Fc reagents and flow cytometry to examine the binding of human Siglec receptors to cell surface glycoRNA. They found that 9 of the 12 marketed Siglec-Fc reagents bind to HeLa cells, and two of which, Siglec-11 and Siglec-14, had binding that was vulnerable to RNase A treatment. [5] These data support the hypothesis that cell surface glycoRNAs might act as direct ligands for Siglec receptors.

Biomedical applications

GlycoRNAs have the potential to revolutionize the field of RNA sequencing (RNA-Seq) and improve our understanding of the human genome. By performing (qRT-PCR) arrays and RNA-Seq, it is possible to identify glycosylated RNA biomarkers that can be used to develop targeted therapies. By detecting altered expression of glycosyltransferases, we can see precisely which RNA is glycosylated and what proteins carry it out. [6] In diseases such as cancer and cystic fibrosis, driver mutations can cause variable RNA glycosylation. [7] To fully comprehend the genetic landscapes and heterogeneity of these disorders, more study in this field of glycoproteomics is required.

GlycoRNAs may also have a role in immune signal transduction. With their ability to bind to Siglec receptors and anti-dsRNA antibodies, glycoRNAs have the potential to be used as biomarkers, enabling rapid diagnosis and prognosis for specific diseases. Furthermore, modifying RNA with glycans may be sensitive to immunotherapy medicines and thus could serve as drug targets. In addition to immunotherapy, several other therapies, including radiotherapy and chemotherapy, have the potential to alter the glycan structure of glycoRNAs on the surface of the cell. Thus, these are expected to be used as a type of immune response signal receiver in many drug therapy. [4]

Related Research Articles

Glycomics is the comprehensive study of glycomes, including genetic, physiologic, pathologic, and other aspects. Glycomics "is the systematic study of all glycan structures of a given cell type or organism" and is a subset of glycobiology. The term glycomics is derived from the chemical prefix for sweetness or a sugar, "glyco-", and was formed to follow the omics naming convention established by genomics and proteomics.

<span class="mw-page-title-main">Glycoprotein</span> Protein with oligosaccharide modifications

Glycoproteins are proteins which contain oligosaccharide chains covalently attached to amino acid side-chains. The carbohydrate is attached to the protein in a cotranslational or posttranslational modification. This process is known as glycosylation. Secreted extracellular proteins are often glycosylated.

<span class="mw-page-title-main">Glycome</span>

The glycome is the entire complement of sugars, whether free or present in more complex molecules, of an organism. An alternative definition is the entirety of carbohydrates in a cell. The glycome may in fact be one of the most complex entities in nature. "Glycomics, analogous to genomics and proteomics, is the systematic study of all glycan structures of a given cell type or organism" and is a subset of glycobiology.

<span class="mw-page-title-main">Consortium for Functional Glycomics</span>

The Consortium for Functional Glycomics (CFG) is a large research initiative funded in 2001 by a glue grant from the National Institute of General Medical Sciences (NIGMS) to “define paradigms by which protein-carbohydrate interactions mediate cell communication”. To achieve this goal, the CFG studies the functions of:

Defined in the narrowest sense, glycobiology is the study of the structure, biosynthesis, and biology of saccharides that are widely distributed in nature. Sugars or saccharides are essential components of all living things and aspects of the various roles they play in biology are researched in various medical, biochemical and biotechnological fields.

Glycosylation is the reaction in which a carbohydrate, i.e. a glycosyl donor, is attached to a hydroxyl or other functional group of another molecule in order to form a glycoconjugate. In biology, glycosylation usually refers to an enzyme-catalysed reaction, whereas glycation may refer to a non-enzymatic reaction.

An oligosaccharide is a saccharide polymer containing a small number of monosaccharides. Oligosaccharides can have many functions including cell recognition and cell adhesion.

<span class="mw-page-title-main">Glycolipid</span> Class of chemical compounds

Glycolipids are lipids with a carbohydrate attached by a glycosidic (covalent) bond. Their role is to maintain the stability of the cell membrane and to facilitate cellular recognition, which is crucial to the immune response and in the connections that allow cells to connect to one another to form tissues. Glycolipids are found on the surface of all eukaryotic cell membranes, where they extend from the phospholipid bilayer into the extracellular environment.

The terms glycans and polysaccharides are defined by IUPAC as synonyms meaning "compounds consisting of a large number of monosaccharides linked glycosidically". However, in practice the term glycan may also be used to refer to the carbohydrate portion of a glycoconjugate, such as a glycoprotein, glycolipid, or a proteoglycan, even if the carbohydrate is only an oligosaccharide. Glycans usually consist solely of O-glycosidic linkages of monosaccharides. For example, cellulose is a glycan composed of β-1,4-linked D-glucose, and chitin is a glycan composed of β-1,4-linked N-acetyl-D-glucosamine. Glycans can be homo- or heteropolymers of monosaccharide residues, and can be linear or branched.

Glycoconjugates are the classification family for carbohydrates – referred to as glycans – which are covalently linked with chemical species such as proteins, peptides, lipids, and other compounds. Glycoconjugates are formed in processes termed glycosylation.

Siglecs(Sialic acid-binding immunoglobulin-type lectins) are cell surface proteins that bind sialic acid. They are found primarily on the surface of immune cells and are a subset of the I-type lectins. There are 14 different mammalian Siglecs, providing an array of different functions based on cell surface receptor-ligand interactions.

<span class="mw-page-title-main">CD22</span> Lectin molecule

CD22, or cluster of differentiation-22, is a molecule belonging to the SIGLEC family of lectins. It is found on the surface of mature B cells and to a lesser extent on some immature B cells. Generally speaking, CD22 is a regulatory molecule that prevents the overactivation of the immune system and the development of autoimmune diseases.

<span class="mw-page-title-main">Galectin</span> Protein family binding to β-galactoside sugars

Galectins are a class of proteins that bind specifically to β-galactoside sugars, such as N-acetyllactosamine, which can be bound to proteins by either N-linked or O-linked glycosylation. They are also termed S-type lectins due to their dependency on disulphide bonds for stability and carbohydrate binding. There have been about 15 galectins discovered in mammals, encoded by the LGALS genes, which are numbered in a consecutive manner. Only galectin-1, -2, -3, -4, -7, -7B, -8, -9, -9B, 9C, -10, -12, -13, -14, and -16 have been identified in humans. Galectin-5 and -6 are found in rodents, whereas galectin-11 and -15 are uniquely found in sheep and goats. Members of the galectin family have also been discovered in other mammals, birds, amphibians, fish, nematodes, sponges, and some fungi. Unlike the majority of lectins they are not membrane bound, but soluble proteins with both intra- and extracellular functions. They have distinct but overlapping distributions but found primarily in the cytosol, nucleus, extracellular matrix or in circulation. Although many galectins must be secreted, they do not have a typical signal peptide required for classical secretion. The mechanism and reason for this non-classical secretion pathway is unknown.

The mannose receptor is a C-type lectin primarily present on the surface of macrophages, immature dendritic cells and liver sinusoidal endothelial cells, but is also expressed on the surface of skin cells such as human dermal fibroblasts and keratinocytes. It is the first member of a family of endocytic receptors that includes Endo180 (CD280), M-type PLA2R, and DEC-205 (CD205).

Polysialic acid is an unusual posttranslational modification that occurs on neural cell adhesion molecules (NCAM). Polysialic acid is considerably anionic. This strong negative charge gives this modification the ability to change the protein's surface charge and binding ability. In the synapse, polysialation of NCAM prevents its ability to bind to NCAMs on the adjacent membrane.

Glycopeptides are peptides that contain carbohydrate moieties (glycans) covalently attached to the side chains of the amino acid residues that constitute the peptide.

<i>N</i>-linked glycosylation Attachment of an oligosaccharide to a nitrogen atom

N-linked glycosylation, is the attachment of an oligosaccharide, a carbohydrate consisting of several sugar molecules, sometimes also referred to as glycan, to a nitrogen atom, in a process called N-glycosylation, studied in biochemistry. The resulting protein is called an N-linked glycan, or simply an N-glycan.

O-linked glycosylation is the attachment of a sugar molecule to the oxygen atom of serine (Ser) or threonine (Thr) residues in a protein. O-glycosylation is a post-translational modification that occurs after the protein has been synthesised. In eukaryotes, it occurs in the endoplasmic reticulum, Golgi apparatus and occasionally in the cytoplasm; in prokaryotes, it occurs in the cytoplasm. Several different sugars can be added to the serine or threonine, and they affect the protein in different ways by changing protein stability and regulating protein activity. O-glycans, which are the sugars added to the serine or threonine, have numerous functions throughout the body, including trafficking of cells in the immune system, allowing recognition of foreign material, controlling cell metabolism and providing cartilage and tendon flexibility. Because of the many functions they have, changes in O-glycosylation are important in many diseases including cancer, diabetes and Alzheimer's. O-glycosylation occurs in all domains of life, including eukaryotes, archaea and a number of pathogenic bacteria including Burkholderia cenocepacia, Neisseria gonorrhoeae and Acinetobacter baumannii.

Translational glycobiology or applied glycobiology is the branch of glycobiology and glycochemistry that focuses on developing new pharmaceuticals through glycomics and glycoengineering. Although research in this field presents many difficulties, translational glycobiology presents applications with therapeutic glycoconjugates, with treating various bone diseases, and developing therapeutic cancer vaccines and other targeted therapies. Some mechanisms of action include using the glycan for drug targeting, engineering protein glycosylation for better efficacy, and glycans as drugs themselves.

<span class="mw-page-title-main">SIGLEC6</span> Protein-coding gene in the species Homo sapiens

Sialic acid-binding Ig-like lectin 6 is a protein that in humans is encoded by the SIGLEC6 gene. The gene was originally named CD33L (CD33-like) due to similarities between these genes but later became known as OB-BP1 due to its ability to bind to this factor and, finally, SIGLEC6 as the sixth member of the SIGLEC family of receptors to be identified. The protein has also been given the CD designation CD327.

References

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  2. 1 2 3 4 5 6 Flynn RA, Pedram K, Malaker SA, Batista PJ, Smith BA, Johnson AG, et al. (June 2021). "Small RNAs are modified with N-glycans and displayed on the surface of living cells". Cell. 184 (12): 3109–3124.e22. doi:10.1016/j.cell.2021.04.023. PMC   9097497 . PMID   34004145.
  3. ↑ Disney MD (June 2021). "A glimpse at the glycoRNA world". Cell. 184 (12): 3080–3081. doi: 10.1016/j.cell.2021.05.025 . PMID   34115968. S2CID   235385901.
  4. 1 2 Clyde D (August 2021). "Sugar-coated RNAs". Nature Reviews. Genetics. 22 (8): 480. doi:10.1038/s41576-021-00388-y. PMID   34168329. S2CID   235635316.
  5. ↑ Smith BA, Bertozzi CR (March 2021). "The clinical impact of glycobiology: targeting selectins, Siglecs and mammalian glycans". Nature Reviews. Drug Discovery. 20 (3): 217–243. doi:10.1038/s41573-020-00093-1. PMC   7812346 . PMID   33462432.
  6. ↑ Angata K, Sawaki H, Tsujikawa S, Ocho M, Togayachi A, Narimatsu H (2020-07-28). "Glycogene Expression Profiling of Hepatic Cells by RNA-Seq Analysis for Glyco-Biomarker Identification". Frontiers in Oncology. 10: 1224. doi: 10.3389/fonc.2020.01224 . PMC   7402167 . PMID   32850363.
  7. ↑ Nishimura S (2011). "Toward automated glycan analysis". Advances in Carbohydrate Chemistry and Biochemistry. Elsevier. 65: 219–271. doi:10.1016/B978-0-12-385520-6.00005-4. ISBN   9780123855206. PMID   21763513.
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