UCL Great Ormond Street Institute of Child Health


Great Ormond Street Institute of Child Health


Using yeast as a tractable cell system to better understand conserved disease proteins and inherited

Supervisors: Professor Sara Mole, Dr Chris Stefan

Using yeast as a tractable cell system to better understand conserved disease proteins and inherited paediatric disease

There are life-limiting inherited neurodegenerative disorders of children that are autosomal recessive lysosomal storage diseases (LSDs, combined incidence ~1:50 000). There is no treatment for most children with such as the neuronal ceroid lipofuscinoses (NCL, Batten disease) and there is a desperate need to understand the biology and disease mechanism to inform therapeutic development. Yeast are powerful unicellular organism for revealing the intricacies of conserved eukaryotic molecular pathways. 
Much insight into the function of CLN3, which causes the juvenile form that accounts for ~50% of all NCL cases, has been eluded from studying Btn1, its functional orthologue in the fission yeast Schizosaccharomyces pombe. Deletion or mutation of btn1 results in a complex array of phenotypes. The most common CLN3 mutation is an intragenic deletion, and modelling this in yeast has shown a functional transcript that loses and retains some functions but also acquires new functions. We have identified two FDA-approved drugs and other small molecules that rescue the yeast disease model and also a number of genetic suppressors of these phenotypes. We are currently using deep RNA sequencing to identify all variant CLN3 transcripts in healthy individuals and patients. Conditions of stress increase transcription of CLN3, with vacuole size dependent on the levels of Btn1. Very recent work supports Btn1/CLN3 as a voltage-gated ion transporter.

To model the function of variant CLN3 healthy and disease transcripts.
To extend understanding of the contribution of Btn1 and CLN3 to maintaining the homeostasis of endolysosomal organelles under conditions of stress, and consequences in disease. Ie. CLN3 function and disease mechanism.

The project will use yeast as the major model system, aiking to confirm key findings in mammalian cell. Specifically:
1. Use strains mutated in key residues or motifs to dissect the functional domains of Btn1 (eg ion pore, voltage sensing, interaction with intra-organelle proteins, link to cytoplasmic Ca2+ homeostasis) and expressing the equivalent of variant transcripts.
2. Exploit existing genetic interactor and small molecule leads, clarify the basis of their compensatory mechanism to the disease (for example slowing down autophagic input or membrane trafficking may prevent overburdening defective lysosomes/vacuoles).
3. Relate findings to human cells.

Timeline (if applicable):
Many missense yeast strains are all available in the lab each carrying a different disease missense mutation in the endogenous btn1 gene on a common genetic background. Strains expressing equivalent variant transcripts will be constructed. (Year 1)
Applying cell biology including recently constructed fluorometric probes that detect potassium ions and part of the project can validate these and assess whether candidate drugs improve anion transport phenotype. (Year 2)
Using patients’ fibroblast lines, CLN3-depleted lines, and access to iPS cells derived from these are available. (Year 3)


1.    Gachet, Y., Codlin, S., Hyams, J. S., & Mole, S. E. (2005). btn1, the Schizosaccharomyces pombe homologue of the human Batten disease gene CLN3, regulates vacuole homeostasis. J CELL SCI, 118(23), 5525-5536. PMID: 16291725 doi:10.1242/jcs.02656.
2.    Codlin, S., Haines, R. L., Burden, J. J. E., & Mole, S. E. (2008). btn1 affects cytokinesis and cell-wall deposition by independent mechanisms, one of which is linked to dysregulation of vacuole pH. J CELL SCI, 121(17), 2860-2870. PMID: 18697832 doi:10.1242/jcs.030122.
3.    Codlin, S., Haines, R. L., & Mole, S. E. (2008). btn1 affects endocytosis, polarization of sterol-rich membrane domains and polarized growth in Schizosaccharomyces pombe. TRAFFIC, 9(6), 936-950. PMC2440566 doi:10.1111/j.1600-0854.2008.00735.x
4.    Haines, R. L., Codlin, S., & Mole, S. E. (2009). The fission yeast model for the lysosomal storage disorder Batten disease predicts disease severity caused by mutations in CLN3. DIS MODEL MECH, 2(1-2), 84-92. PMC2615160 doi:10.1242/dmm.000851
5.    Bond, M. E., Brown, R., Rallis, C., Bähler, J., Mole, S. E. (2015). A central role for TOR signalling in a yeast model for juvenile CLN3 disease. Microbial Cell. 2(12): 466-480. PMC5354605 doi:10.15698/mic2015.12.241
6.    Minnis CJ, Townsend SJ, Petschnigg J, Tinelli, E, Bähler J, Russell C, Mole SE. (2021). Global network analysis in Schizosaccharomyces pombe reveals three distinct consequences of the common 1-kb deletion causing juvenile CLN3 disease. Scientific Reports. 11(1) 6332.  doi: 10.1038/s41598-021-85471-4
7.    UCL PhD Thesis of Yaxuan Lyu (2021)
8.    Sharaireh AM, Guevara-Ferrer M, Herranz-Martin S, Garcia-Macia M, Phillips A, Tierney A, Hughes MP, Coombe-Tennant O, Nelvagel H, Burrows AE, Fielding S, FitzPatrick LM, Thornton CD, Storch S, Mole SE, Dowsey A, Unwin R, Bolanos JP, Rahim AA, McKay TR. (2022). CLN7 mutation causes aberrant redistribution of protein isoforms and contributes to Batten disease pathobiology. bioRxiv. doi 10.1101/2022.04.21.488782