Crowdfunded whole-genome sequencing of the celebrity cat Lil BUB identifies causal mutations for her osteopetrosis and polydactyly

Author's response

Dario G. Lupiañez, Orsolya Simmons and Daniel M. Ibrahim shared

1. Lil Bub’s celebrity status was clearly important for a successful crowdfunding campaign. Do you think it would be possible to fund a project like this without the existing fanbase that Lil Bub brought with her?

Lil Bub’s fanbase was undeniably crucial to the success of our funding campaign. This is clearly depicted on Figure 2 of our manuscript, where page views and donations spiked when our crowdfunding campaign was mentioned on Lil BUB’s social media pages. We always knew that her popularity was going to help us raise the funds needed for the project, but getting a broad audience interested in the outcome of your project is inherent to the concept of a crowdfunding campaign. Each crowdfunding campaign must have an idea of who is going to donate, and LilBUB was our door-opener.

For this reason it is difficult for us to imagine how our funding campaign would have played out without the ‘celebrity’ aspect, but there have been other crowdfunded science projects that successfully raised more money than we did without having such a well-known internet phenomenon at the heart of their research.  Some of our favorites include the Kakapo project, which aims to sequence the genomes of all 125 remaining endangered kakapo birds in New Zealand (which raised $45,400) [4] and the Bighorn Basin Dinosaur Project that raised money for its field expeditions between 2015-2018 [5]. This demonstrates a commitment from the public to know more about and to participate in scientific endeavors, and shows that it is in our hand, as scientists, to engage this audience in our research and to provide appropriate tools for a better understanding of the scientific method. When scientists can get the public interested in the science and spend enough time on outreach it is possible to raise large amounts of money. However, we also believe that crowd-funded research requires extensive time-engagement on behalf of scientists, and that can be difficult to reconcile with such a demanding career.

2. Are you considering crowdfunding as a periodic way of funding some of your research projects? Would there be a potential danger for projects to become too biased on mediatic research?

Through our experience with the Lilbubome, we learned that crowdfunding has a lot of potential for funding of small-scale projects with targeted questions. We believe that this combination of factors creates a very specific niche that nicely complements traditional funding, which is normally focused on larger-budget research. So if a funding agency doesn’t support a really cool experiment, crowdfunding might be a way to get the money. But we’re also aware of the hard work it takes to raise funds this way, so we would likely only consider this option under special circumstances in the future.

Since crowdfunding is generally targeted towards a broad audience, it is of course easier to raise money for projects that broadly appeal the public. We don’t think there’s an inherent problem with media-friendly research. In fact, we feel that it’s rather the opposite: crowdfunding research is a unique opportunity to bring science out of the lab, and it also enables us to tackle questions very scientifically that non-scientists might ask during their normal lives .

3. After the success of this research, how do the authors envision the future of crowdfunding in science? Could there be too many projects trying to use this strategy and, thus, many of them fail to succeed?

Before we consider the financial aspect of crowdfunding, we want to emphasize that a crowdfunding campaign provides a great framework to reach out to an audience and discuss science with them. It is a campaign, which means that it focuses your attention on outreach over a restricted period and gives you a goal to achieve – instead of leaving a lot of loose ends. If one would want to reach out to an existing community (e.g. outsource “data analysis” in a gamefication project) a crowdfunding campaign might be just the right thing to focus these efforts.

From the financial perspective, we anticipate that crowdfunding will continue to fill a small but important niche in science funding. The amounts raised are typically low (median $3,015, average $12,652 on experiment.com [6]) compared to traditional routes of funding, but it can be a way to empower people (e.g. PhD students, citizen scientists) who otherwise might not have access to funding or it could be used to fund purely curiosity-driven research that might not be supported by government funding. However, given the large time commitment needed for a successful campaign and the small sums that are involved, we predict that most scientists will not use crowdfunding as a routine way to obtain financial support for their research.

4. What would be the advice for young motivated researchers who want to get both funding and public engagement?

First, be willing to dedicate a substantial amount of your time – it takes time to have thoughtful interactions with people and to build momentum for a successful crowdfunding campaign.  Crowdfunding relies heavily on reaching people, generally through social media. Therefore it is important to create interest during the entire funding campaign. This means that for the 30 or 40 days of the funding period, you will spend A LOT of time generating and posting content regularly, engaging with the public more or less constantly, and – if you’re lucky – interacting with media outlets  

Second, think carefully about your target audience and the message you want to convey: it is easy to get lost in the details of your ideas, so having a central theme to which you can return and using language that fits your audience is key to the success of both engagement and funding.

5. Many different mutations affecting the ZRS limb enhancer cause ectopic anterior expression of the Shh gene and UK1, UK2 and Hemingway mutations are examples of that [1]. Is it known which transcription factor/s bind the ZRS enhancer? Do the authors think the different mutations affect the same putative binding site/s?

The most convincing data about the transcription factors that function of the ZRS in the limb are from detailed in another paper by Lettice et al, who specifically highlight the opposing roles of different ETS factor family members [7]. Some polydactyly-causing ZRS mutations directly affect the binding sites of these factors, but it is unclear whether all different mutations would act via the same mechanism. Interestingly, both the Hemingway mutation and the UK2 mutation are immediately adjacent to one of the sites identified by Lettice et al as an ETS binding site (ETS site 2 in their paper), so we can speculate (but have no evidence) that these mutations somehow change binding at the adjacent site.

6. ZMPSTE24 is a metalloprotease involved, among other things, in Lamin A maturation and knockout mice have been studied due to the similarities with phenotypes of patients with Progeria syndrome, showing strong phenotypes in mice when in homozygosis [2]. In Hutchinson-Gilford Progeria Syndrome (HGPS), a heterozygous point mutation affecting a splicing site in LMNA gene that impairs correct Lamin A maturation and causes premature aging. However, while in humans this mutation appears spontaneously and causes severe phenotypes in heterozygosis, the equivalent mutation in mice confers the strong phenotype when the animals are homozygous [3]. Due to the differences in the phenotype effect of heterozygous and homozygous mutations in human and mouse, we were wondering whether the heterozygous loss-of-function allele of ZMPSTE24 has been reported in cats and if the allele found in Lil BUB’s genome could be contributing to her congenital malformations.

To date, we are not aware of any known mutations in ZMPSTE24 in cats. However,  we considered RANK a much more likely causal candidate for Bub´s osteopetrosis, given the close resemblance between the human and mouse RANK mutation phenotypes, and that ZMPSTE24-deficient mice develop loss of bone mass, rather than increased calcification [8,9]. We cannot exclude, though, that the heterozygous ZMPSTE24 mutation could potentially modify some of Lil BUB’s traits, including the osteopetrosis.

Supplementary References

4. Crowdfunding campaign: Iorns D, Dighy A, Robertson Bruce and Howard JT. Sequencing the genome of all known kakapo.

https://experiment.com/projects/sequencing-the-genomes-of-all-known-kakapo

5. Crowdfunding campaign: Schein JP. Help us excavate a dinosaur bonebed in Wyoming’s bighorn basin.

https://experiment.com/users/scheijp

6. Sauermann H, Franzoni C, Shafi K. 2019. Crowdfundingcientific research: Descriptive insights and correlates of funding success. PLoS ONE 14(1): e0208384.

7. Lettice L et al. 2012. Opossing functions of the ETS factor family define shh spatial expression in limb buds and underlie polydactyly. Dev Cell. 22, 459-467.

8. Rivas D et al. 2009. Accelerated features of age-related bone loss in Zmpste24 metalloproteinase deficient mice. The Journal of Gerontology. 64A, 1015-1024.

9. Bergo MO et al. 2002. Zmpste24 deficiency in mice causes spontaneous bone fractures, muscle weakness and prelamin A processing defect. PNAS. 99(20), 13049-13054.