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vaccine design

Using R2D2 to Understand RNA Folding

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If you love learning more about biology at a fundamental level, I have a great video for you! It simulates the 3D folding of RNA. RNA is a single stranded molecule, but it is still capable of forming internal loops that can be stabilized by base pairing, just like its famously double-stranded parent, DNA. Understanding more about RNA folding may be valuable in many different areas of biomedical research, including developing ways to help people with RNA-related diseases, such as certain cancers and neuromuscular disorders, and designing better mRNA vaccines against infectious disease threats (like COVID-19).

Because RNA folding starts even while an RNA is still being made in the cell, the process has proven hugely challenging to follow closely. An innovative solution, shown in this video, comes from the labs of NIH grantees Julius Lucks, Northwestern University, Evanston, IL, and Alan Chen, State University of New York at Albany. The team, led by graduate student Angela Yu and including several diehard Star Wars fans, realized that to visualize RNA folding they needed a technology platform that, like a Star Wars droid, is able to “see” things that others can’t. So, they created R2D2, which is short for Reconstructing RNA Dynamics from Data.

What’s so groundbreaking about the R2D2 approach, which was published recently in Molecular Cell, is that it combines experimental data on RNA folding at the nucleotide level with predictive algorithms at the atomic level to simulate RNA folding in ultra-slow motion [1]. While other computer simulations have been available for decades, they have lacked much-needed experimental data of this complex folding process to confirm their mathematical modeling.

As a gene is transcribed into RNA one building block, or nucleotide, at a time, the elongating RNA strand folds immediately before the whole molecule is fully assembled. But such folding can create a problem: the new strand can tie itself up into a knot-like structure that’s incompatible with the shape it needs to function in a cell.

To slip this knot, the cell has evolved immediate corrective pathways, or countermoves. In this R2D2 video, you can see one countermove called a toehold-mediated strand displacement. In this example, the maneuver is performed by an ancient molecule called a single recognition particle (SRP) RNA. Though SRP RNAs are found in all forms of life, this one comes from the bacterium Escherichia coli and is made up of 114 nucleotides.

The colors in this video highlight different domains of the RNA molecule, all at different stages in the folding process. Some (orange, turquoise) have already folded properly, while another domain (dark purple) is temporarily knotted. For this knotted domain to slip its knot, about 5 seconds into the video, another newly forming region (fuchsia) wiggles down to gain a “toehold.” About 9 seconds in, the temporarily knotted domain untangles and unwinds, and, finally, at about 23 seconds, the strand starts to get reconfigured into the shape it needs to do its job in the cell.

Why would evolution favor such a seemingly inefficient folding process? Well, it might not be inefficient as it first appears. In fact, as Chen noted, some nanotechnologists previously invented toehold displacement as a design principle for generating synthetic DNA and RNA circuits. Little did they know that nature may have scooped them many millennia ago!

Reference:

[1] Computationally reconstructing cotranscriptional RNA folding from experimental data reveals rearrangement of non-naïve folding intermediates. Yu AM, Gasper PM Cheng L, Chen AA, Lucks JB, et. al. Molecular Cell 8, 1-14. 18 February 2021.

Links:

Ribonucleic Acid (RNA) (National Human Genome Research Institute/NIH)

Chen Lab (State University of New York at Albany)

Lucks Laboratory (Northwestern University, Evanston IL)

NIH Support: National Institute of General Medical Sciences; Common Fund


Antibody Points to Possible Weak Spot on Novel Coronavirus

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Credit: Meng Yuan and Nicholas Wu, Wilson Lab, The Scripps Research Institute, La Jolla, CA

Researchers are working hard to produce precise, 3D molecular maps to guide the development of safe, effective ways of combating the coronavirus disease 2019 (COVID-19) pandemic. While there’s been a lot of excitement surrounding the promise of antibody-based tests and treatments, this map you see above highlights another important use of antibodies: to inform efforts to design a vaccine.

This image shows the crystal structure of a human antibody (heavy chain in orange, light chain in yellow), which is a blood protein our immune systems produce to attack viruses and other foreign invaders. This particular antibody, called CR3022, is bound to a key surface protein of the novel coronavirus (white).

The CR3022 antibody actually doesn’t come from someone who has recovered from COVID-19. Instead, it was obtained from a person who, nearly two decades ago, survived a bout of severe acute respiratory syndrome (SARS). The SARS virus, which disappeared in 2004 after a brief outbreak in humans, is closely related to the novel coronavirus that causes COVID-19.

In a recent paper in the journal Science, the NIH-funded lab of Ian Wilson, The Scripps Research Institute, La Jolla, CA, along with colleagues at The University of Hong Kong, sought to understand how the human immune system interacts with and neutralizes this highly infectious virus [1]. The lab did so by employing high-resolution X-ray crystallography tools [2]. They captured the atomic structure of this antibody bound to its target by shooting X-rays through its crystallized form. (An antibody measures about 10 nanometers; a nanometer is 1 billionth of a meter.)

Other researchers had shown previously that CR3022 cross-reacts with the novel coronavirus, although the antibody doesn’t bind tightly enough to neutralize and stop it from infecting cells. So, Wilson’s team went to work to learn precisely where the antibody attaches to the novel virus. Those sites are of special interest because they highlight spots on a virus that are vulnerable to attack—and, as such, potentially good targets for vaccine designers.

A key finding in the new paper is that the antibody binds a highly similar site on both the SARS and novel coronaviruses. Those sites differ in each virus by just four amino acids, the building blocks of a protein.

This is particularly interesting because the antibody pictured above is bound to a spike protein, which is the appendage on both the SARS and novel coronavirus that enables them to bind to a key receptor protein on the surface of human cells, called ACE2. This binding activity marks the first step for these viruses in gaining entry into human cells and infecting them.

The human antibody shown in this image locks onto the virus’s spike protein at a different location than where the human ACE2 protein binds to the novel coronavirus. Intriguingly, the antibody binds to a spot on the novel coronavirus that is usually hidden, except for when virus shapeshifts its structure in order to infect a cell.

The findings suggest that a successful vaccine may be one that elicits antibodies that targets this same spot, but binds more tightly than the one seen above, thereby protecting human cells against the virus that causes COVID-19. However, Wilson notes that this study has just uncovered one potential vulnerability of the novel coronavirus, and it is likely the virus likely has many more that could be revealed with further study.

To continue in this quest to design a safe and effective vaccine, Wilson and his colleagues are now gathering blood samples to collect antibodies from people who’ve recovered from COVID-19. So, we can look forward to seeing some even more revealing images soon.

References:

[1] A highly conserved cryptic epitope in the receptor-binding domains of SARS-CoV-2 and SARS-CoV. Yuan M, Wu NC, Zhu X, Lee CD, So RTY, Lv H, Mok CKP, Wilson IA. Science. 2020 Apr 3.

[2] 100 Years Later: Celebrating the Contributions of X-ray Crystallography to Allergy and Clinical Immunology. Pomés A, Chruszcz M, Gustchina A, Minor W, Mueller GA, Pedersen LC, Wlodawer A, Chapman MD. J Allergy Clin Immunol. 2015 Jul;136(1):29-37.

Links:

Coronaviruses (National Institute of Allergy and Infectious Diseases/NIH)

Coronavirus (COVID-19) (NIH)

Ian Wilson (The Scripps Research Institute, La Jolla, CA)

NIH Support: National Institute of Allergy and Infectious Diseases; National Cancer Institute; National Institute of General Medical Sciences