Biology subjects

Mole, S. E.

Publications and source records attributed to Mole, S. E..

6 recordsLinked to original sources

bioRxiv2026

Bro1-Mediated Trafficking Couples TOR Signalling to Cellular Metabolism and Longevity

Adaptation to nutrient availability requires coordination between growth control, metabolism, and intracellular trafficking. In eukaryotes, inhibition of Target of Rapamycin (TOR) signalling robustly promotes stress resistance and longevity, yet how reduced growth signalling is coupled to organelle dynamics and proteome remodelling remains unclear. Here, we identify the conserved ESCRT-associated protein Bro1 as a central integrator of TOR signalling, vacuolar trafficking, and metabolic adaptation. Using fission yeast, we show that Bro1 is required for normal lifespan and for the global proteomic reprogramming that accompanies TOR inhibition. In Bro1 mutant cells, repression of ribosome biogenesis is uncoupled from activation of catabolic, vacuolar, and metabolic pathways, resulting in an altered metabolic state characterised by elevated lipid metabolism and increased abundance of nutrient transporters. Mechanistically, Bro1 promotes TOR-dependent cargo deubiquitination, vacuolar trafficking, and turnover of plasma membrane hexose transporters and enables appropriate nuclear relocalisation of the transcriptional repressor Scr1. In the absence of Bro1, nutrient transporters persist at the cell surface despite TOR inhibition, conferring resistance to TOR inhibitors while impairing stress responses and reducing lifespan. Together, our findings establish Bro1 as a key coordinator linking ESCRT-mediated endosomal-vacuolar trafficking to TOR-dependent metabolic control. By coupling growth suppression to enhanced recycling and cellular maintenance, Bro1 enables the transition from growth to longevity-promoting states, revealing a mechanism connecting intracellular trafficking, metabolism, and ageing.

genetics↗

bioRxiv2025

Targeted long-read RNA sequencing reveals the complexity of CLN3 transcription and the consequences of the most common 1-kb deletion in patients with juvenile CLN3 disease

Most genes are not yet fully annotated, and the extent of their transcript diversity and the roles and significance of specific isoforms is not understood. This information is therefore lacking for disease genes. The CLN3 gene underlies classic juvenile CLN3 disease, also known as juvenile neuronal ceroid lipofuscinosis, a rare paediatric neurodegenerative disorder. The most common cause of this biallelic disorder is a 1-kb intragenic deletion that removes two internal coding exons (exons 7 and 8). Here, we report findings from the first long-read RNA sequencing targeting CLN3 in blood samples derived from control individuals and from patients clinically and genetically diagnosed with juvenile CLN3 disease. We find that CLN3 transcription is complex, with >80 different transcripts encoding >35 different open reading frames (ORF) of different lengths, and no dominantly expressed transcript. The 1-kb deletion has direct consequences on this. This is consistent across patients, with total loss of some transcripts including those encoding the canonical 438 amino acid protein and other significant smaller isoforms. The highest expressed disease transcripts include those lacking exons 7 and 8 and encoding a 181 amino acid protein isoform, and other novel isoforms that lack additional exons and encode longer ORFs. The different effects on transcription of other CLN3 disease-causing variants are revealed in single patients. Together, these findings confirm the complexity of transcription at the CLN3 locus, reveal the impact of the 1-kb deletion and other variants on isoform abundance, and highlight the importance of understanding the contribution of these isoforms to CLN3 function in health and disease. Moreover, they impact the future design and development of personalised therapeutics and the design and generation of disease models. Finally, they underline the importance of full annotation for disease genes.

genetics↗

bioRxiv2023

CLN3 transcript complexity revealed by long-read RNA sequencing analysis

BackgroundBatten disease is a group of rare inherited neurodegenerative diseases. Juvenile CLN3 disease is the most prevalent type, and the most common mutation shared by most patients is the "1-kb" deletion which removes two internal coding exons (7 and 8) in CLN3. Previously, we identified two transcripts in patient fibroblasts homozygous for the "1-kb" deletion: the "major" and "minor" transcripts. To understand the full variety of disease transcripts and their role in disease pathogenesis, it is necessary to first investigate CLN3 transcription in "healthy" samples without juvenile CLN3 disease. MethodsWe leveraged PacBio long-read RNA sequencing datasets from ENCODE to investigate the full range of CLN3 transcripts across various tissues and cell types in human control samples. Then we sought to validate their existence using data from different sources. ResultsWe found that a readthrough gene affects the quantification and annotation of CLN3. After taking this into account, we detected over 100 novel CLN3 transcripts, with no dominantly expressed CLN3 transcript. The most abundant transcript has median usage of 42.9%. Surprisingly, the known disease-associated "major" transcripts are detected. Together, they have median usage of 1.51% across 22 samples. Furthermore, we identified 48 CLN3 ORFs, of which 26 are novel. The predominant ORF that encodes the canonical CLN3 protein isoform has median usage of 66.7%, meaning around one-third of CLN3 transcripts encode protein isoforms with different stretches of amino acids. The same ORFs could be found with alternative UTRs. Moreover, we were able to validate the translational potential of certain transcripts using public mass spectrometry data. ConclusionOverall, these findings provide valuable insights into the complexity of CLN3 transcription, highlighting the importance of studying both canonical and non-canonical CLN3 protein isoforms as well as the regulatory role of UTRs to fully comprehend the regulation and function(s) of CLN3. This knowledge is essential for investigating the impact of the "1-kb" deletion and rare mutations on CLN3 transcription and disease pathogenesis.

genetics↗

bioRxiv2022

CLN7 mutation causes aberrant redistribution of protein isoforms and contributes to Batten disease pathobiology

The variant late infantile form of the inherited neurodegenerative Batten disease (BD) is caused by mutations in the CLN7/MFSD8 gene and represents a strong candidate for gene therapy. Post-natal intracerebral administration of AAV9-hCLN7 to Cln7{Delta}ex2 knockout mice resulted in extended lifespan but dose escalation resulted in reduced acuity in neurophysiology tests, cerebral atrophy and elevated neuroinflammation. Comparing patient and control iPSC-derived neural progenitor cells (iNPC) we discovered that CLN7 localizes to the nucleus as well as the endolysosomal network and is differentially distributed in BD iNPC. Proteomics identified a profound nuclear defect in BD iNPC that compounds with mitochondrial and lysosomal metabolic defects resulting in elevated apoptosis. We further identified a 50kDa common nuclear CLN7 isoform and a 37kDa isoform that accumulates only in BD iNPC nuclei. Our findings suggest that successful treatment of CLN7 BD will require combinatorial therapies addressing both loss and aberrant gain of protein function.

cell biology↗

bioRxiv2020

Mitochondrial collapse links PFKFB3-promoted glycolysis with CLN7/MFSD8 neuronal ceroid lipofuscinosis pathogenesis

The neuronal ceroid lipofuscinoses (NCLs) are a family of monogenic life-limiting pediatric neurodegenerative disorders collectively known as Batten disease1. Although genetically heterogeneous2, NCLs share several clinical symptoms and pathological hallmarks such as lysosomal accumulation of lipofuscin and astrogliosis2,3. CLN7 disease belongs to a group of NCLs that present in late infancy4-6 and, whereas CLN7/MFSD8 gene is known to encode a lysosomal membrane glycoprotein4,7-9, the biochemical processes affected by CLN7-loss of function are unexplored thus preventing development of potential treatments1,10. Here, we found in the Cln7{Delta}ex2 mouse model11 of CLN7 disease that failure in the autophagy-lysosomal pathway causes accumulation of structurally and bioenergetically impaired, reactive oxygen species (ROS)-producing neuronal mitochondria that contribute to CLN7 pathogenesis. Cln7{Delta}ex2 neurons exhibit a metabolic shift mediated by pro-glycolytic enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (PFKFB3). PFKFB3 inhibition in Cln7{Delta}ex2 mice in vivo and in CLN7 patients-derived cells rectified key disease hallmarks. Thus, specifically targeting glycolysis may alleviate CLN7 pathogenesis.

neuroscience↗

bioRxiv2019

Neonatal brain-directed gene therapy rescues a mouse model of neurodegenerative CLN6 Batten disease

The neuronal ceroid lipofuscinoses (NCLs), more commonly referred to as Batten disease, are a group of inherited lysosomal storage disorders that present with neurodegeneration, loss of vision and premature death. There are at least 13 genetically distinct forms of NCL. Enzyme replacement therapies and preclinical studies on gene supplementation have shown promising results for NCLs caused by lysosomal enzyme deficiencies. The development of gene therapies targeting the brain for NCLs caused by defects in transmembrane proteins has been more challenging and only limited therapeutic effects in animal models have been achieved so far. Here, we describe the development of an adeno-associated virus (AAV)-mediated gene therapy to treat the neurodegeneration in a mouse model of CLN6 disease, a form of NCL with a deficiency in the membrane-bound protein CLN6. We show that neonatal bilateral intracerebroventricular injections with AAV9 carrying CLN6 increase lifespan by more than 90%, maintain motor skills and motor coordination and reduce neuropathological hallmarks of Cln6-deficient mice up to 23 months post vector administration. These data demonstrate that brain-directed gene therapy is a valid strategy to treat the neurodegeneration of CLN6 disease and may be applied to other forms of NCL caused by transmembrane protein deficiencies in the future.

neuroscience↗