CLN5 and CLN3 function as a complex to regulate endolysosome function

Abstract

CLN5 is a soluble endolysosomal protein that regulates the itinerary of the lysosomal sorting receptor sortilin. Mutations in this protein cause neuronal ceroid lipofuscinosis, a rare neurodegenerative disorder, and have also been associated with Alzheimer’s disease, suggesting functional defects in a common pathway. We previously found that depletion of CLN5 leads to dysfunctional retromer, resulting in the degradation of the lysosomal sorting receptor, sortilin. However, how a soluble lysosomal protein can modulate the function of a cytosolic protein is not known. In this work, we show that deletion of CLN5 not only results in retromer dysfunction, but also in impaired endolysosome fusion events. This results in delayed degradation of endocytic proteins and in defective autophagy. CLN5 modulates these various pathways by regulating downstream interactions between CLN3, an integral membrane protein, Rab7A and a subset of Rab7A effectors. Mutations in CLN3 are also a cause of neuronal ceroid lipofuscinosis. Our data supports a model where CLN3 and CLN5 function as an endolysosome complex regulating several endosomal functions.Summary StatementWe have previously demonstrated that CLN3 is required for efficient endosome-to-trans Golgi Network (TGN) trafficking of sortilin by regulating retromer function. In this work, we show that CLN5, which interacts with CLN3, regulates retromer function by modulating key interactions between CLN3, Rab7A, retromer, and sortilin. Therefore, CLN3 and CLN5 serve as endosomal switch regulating the itinerary of the lysosomal sorting receptors.

Deficiency of the lysosomal protein CLN5 alters lysosomal function and movement

“…Although we did not find a change in lysosomal area and intensity with CLN5 inhibition in our iPSC-derived CLN5-deficient human neurons, we did detect an increased number of non-motile lysosomes (Figure 2I) in CLN5-deficient neurons with the overall number of lysosomes remaining the same (Figure 2E). The increased number of non-motile lysosomes explains the accumulation of lysosomes, seen in the CLN5-deficient neurons (Figure 2A, 2 nd , 3 rd and 4 th panels), and as reported by Uusi-Rauva et al Another study of CLN5 in HeLa cells by Yasa et al (2021) [19] suggested that CLN5 KO HeLa cells were not able to move lysosomes as efficiently as wild type HeLa cells. In our study, impaired anterograde movement in the CLN5-deficient human neurons could indicate that the neuronal projections are being exempt from getting their waste cleared, which could cause neuronal toxicity at the synapses.…”

“…Another study of CLN5 in HeLa cells by Yasa et al . (2021)[19] suggested that CLN5 KO HeLa cells were not able to move lysosomes as efficiently as wild type HeLa cells. In our study, impaired anterograde movement in the CLN5-deficient human neurons could indicate that the neuronal projections are being exempt from getting their waste cleared, which could cause neuronal toxicity at the synapses.…”

“…Lysosomes travel to distal ends of neurons to maintain local degradative activities, therefore making lysosomal movement crucial to maintain homeostasis in the neurons [18]. Some CLN proteins have been recently shown to alter lysosomal positioning [19,20], however whether inhibiting CLN5 changes the lysosomal movement/trafficking in human neurons is unknown. To assess the lysosomal movement in control vs CLN5i neurons, kymographs were generated from time lapse videos of LysoTracker Red stained lysosomes in control vs CLN5i neurons (Figure 2H).…”

The Batten disease protein CLN3 is important for stress granules dynamics and translational activity

“…NCL pathologies are linked to the dysfunction of several CLN proteins involved in intracellular trafficking (13,39). To assess if the delay in SG dissolution is CLN3 specific, we investigated SG dynamics in a HeLa KO model of CLN5 disease.…”

Cln5 represents a new type of cysteine-based S -depalmitoylase linked to neurodegeneration

“…The precursor Cln5 protein is a 407-amino acid-long membranelocated protein that is cleaved by signal peptide peptidase (SPP) and its homologs the SPP-like proteases (SPPL), resulting in the soluble Cln5 protein (7). Several studies have demonstrated Cln5 localized to lysosomes (8,9) and associated with endolysosomal dysfunction (10)(11)(12). However, the structure and molecular function of the Cln5 protein remains unknown (13), precluding a mechanistic understanding and potential treatment development for Cln5dependent NCL.…”