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Supervisors: Professor Sara Mole, Dr Chris Stefan
Background:
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. Mutations in CLN3 cause the juvenile form that accounts for ~50% of all NCL cases. Yeast are powerful unicellular organism for revealing the intricacies of conserved eukaryotic molecular pathways. Much insight into the function of CLN3 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. 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. 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.
Aims/Objectives:
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.
Methods:
The project will use yeast as the major model system, confirming key findings in mammalian cell. Specifically:
1. Use strains mutated in key residues to dissect the functional domains of Btn1 (eg ion pore, voltage sensing, interaction with intra-organelle proteins, link to cytoplasmic Ca2+ homeostasis).
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).
References:
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
